Download Cardiovascular Research Center Amsterdam

1st Amsterdam Cardiovascular Research Institute
SYMPOSIUM
ARTIS 9-10-2015
The mission of the Amsterdam Cardiovascular Research Institute (ACI) is to design knowledgebased treatment strategies to prevent and cure cardiovascular disease. For this purpose,
education, research and clinical activities are focused within 4 Centers of Excellence that not
only translate experiments to clinical interventions, but also use clinical observations to develop
and validate novel biological concepts. Support programs on infrastructure, education, talent
training, branding and industry and valorization will provide a common basis for translational
research within the Centers of Excellence. In this framework, we interconnect excellence in
clinical and basic research and provide unique infrastructure to train and attract talented
cardiovascular researchers to Amsterdam. As already successfully achieved a number of times,
we expect that this translational research approach will lead to discovery and proof of concept
of innovative, effective strategies that improve cardiovascular health.
Research themes within the 4 Centers of Excellence & support programs
HEART FAILURE
&
ARRHYTHMIAS
& THROMBOSIS
DIASTOLIC HEART
THROMBOSIS
FAILURE
CARDIOMYOPATHIES
ARRHYTHMIAS
ATRIAL FIBRILLATION
PULMONARY
HYPERTENSION
PULMONARY
ATHEROSCLEROSIS
& ISCHEMIC
SYNDROMES
DIABETES
&
METABOLISM
ATHEROSCLEROSIS
PATHOGENESIS DM2
NEUROVASCULAR
NEW DM2 THERAPIES
SYNDROMES
CIRCULATION AND RIGHT
HEART FAILURE
ANEURYSMS
RESPIRATORY MUSCLE
CORONARY ISCHEMIC
STRESS IN THE
SYNDROMES
CRITICALLY ILL
PERIPHERAL ARTERY
COMPLICATIONS OF
DIABETES
ORGAN FAILURE-INDUCED
CARDIOVASCULAR
DISEASE
DISEASE
I. INFRASTRUCTURE & EXPERTISE
II. EDUCATION PROGRAM
III. TALENT PROGRAM & OUT OF THE BOX IDEAS
IV. BRANDING & COMMUNICATION
V. INDUSTRY & VALORIZATION
By combining our expertise within ACI, we will strengthen and streamline our education,
research and clinical activities and build a common strategy to master current challenges in
cardiovascular research. During the 1st ACI symposium we highlight themes from the first
CoE Heart Failure & Arrhythmias and discuss support programs I (imaging) and III.
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Key lecturers
Thomas Eschenhagen:
Thomas Eschenhagen, MD, is Professor of Pharmacology and Director of the
Department of Experimental Pharmacology and Toxicology at the University
Medical Center Hamburg Eppendorf, Germany. He is also chairman of board
of directors of the German Centre of Cardiovascular Research (DZHK).
Dr. Eschenhagen has concentrated his research efforts on understanding
molecular mechanisms of heart failure with a focus on 
-adrenergic signaling,
its adaptation in heart failure and consequences on contractile function. He is best known for
his pioneering work on 3-dimensional engineered heart tissue (EHT) from primary cardiac cells,
starting 1994 in collaboration with Elliot Elson, St. Louis, USA. Originally designed as an
improved in vitro model for drug testing and target validation, the EHT technology has been
expanded to an automated 24-well screening platform. In combination with human embryonic
and induced pluripotent stem cell-derived cardiomyocyte, this technique opens new
perspectives in biomedicine, e.g. medium throughput drug screening, LQT and cardiotoxicity
testing, disease modeling and cardiac regeneration. Dr. Eschenhagen and his group have
recently shown that EHTs survive after implantation on injured guinea-pig hearts, can couple to
host myocardium and improve cardiac function.
For his contributions to science, he received the Fraenkel Award of the German Society of
Cardiology (1997) and the Outstanding Investigator Award of the ISHR (2012). He is member of
the German Academy of Science Leopoldina (2008) and received an ERC Advanced Grant (2013).
Connie Bezzina
Connie Bezzina graduated in Pharmacy from the University of Malta in 1992
and obtained her PhD in Genetics from the same university in 1998. In 1997
she joined the Department of Experimental Cardiology at the Academic
Medical Center (University of Amsterdam) and was appointed Professor of
Molecular Cardiogenetics in 2012. Her research focuses on the genetics of
cardiac arrhythmias associated with sudden cardiac death. Her group
employs an integrative approach, utilizing a range of genetic and functional methodologies
aimed at the identification of genetic factors involved in susceptibility to cardiac arrhythmia and
the understanding the underlying mechanisms. In 2005 she was appointed Established
Investigator of the Netherlands Heart Foundation and in 2013 she received the Outstanding
Achievement Award from the European Society of Cardiology Council on Basic Cardiovascular
Science.
Bianca Brundel:
Bianca Brundel is currently Established Investigator, funded by the Dutch Heart
Foundation (2014-2019) and Consortium leader of HALT&REVERSE
(collaboration with Natasja de Groot EMC; 2014-2019). Bianca Brundel is
project leader, together with Prof. Dr. J. van der Velden of the CVON project
'Dosis' (2015-2019). This project aims to identify determinants for onset of
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inherited cardiomyopathy. She opened up new directions for research on protein homeostasis
and the protein quality control system underlying atrial fibrillation with the development of a
Drosophila model. She is founder of NYKEN BV (2009), a bio-pharmaceutical company to
develop innovative therapies for the treatment of atrial fibrillation, using its proprietary
platform of Heat Shock Factor activators (www.nyken.com). Brundel has collaborations with
(inter)national top researchers (Stanley Nattel, Ivor Benjamin, Edward O'Brien, Rolf Bodmer).
Bianca will work fulltime at the VUmc,after the move to theO2 building in January 2016
Key Lectures:
ERC Project: Individualized risk prediction with iPSC-derived engineered heart tissue
(IndivuHeart)
By Thomas Eschenhagen
Heart failure is the consequence of very diverse medical conditions, but it is treated by a the
same 3-5 drugs. This “one for all” approach has to be advanced by an individualized approach
allowing early patient- or group-specific treatment. Inherited cardiomyopathies can serve as
paradigmatic examples of different heart failure pathogenesis. Both gain- and loss-of-function
mutations of the same gene cause disease, calling for disease-specific agonism or antagonism of
this gene´s function. However, mutations alone do not predict the severity of cardiomyopathies
nor therapy, because their impact on cardiomyocyte function is modified by numerous factors,
including the genetic context. Today, the function of patient-specific cardiomyocytes can be
assessed via human induced pluripotent stem cells (hiPSC). Yet, unfolding the true potential of
this technology requires robust quantitative high content assays. Our method to generate 3Dengineered heart tissue (EHT) from hiPSC provides a fully automated, high content analysis of
heart muscle function and the response to stressors in the dish. The aim of the ERC project
IndivuHeart is to make the technology a clinically applicable test. Major steps are (i) in depths
clinical phenotyping of patients with cardiomyopathies or HFpEF, (ii) panel sequencing of all
relevant cardiac genes, (iii) follow-up of clinical course, (iv) generation of hiPSC cell lines and
EHTs from 40 patients and 40 healthy controls, (v) extensive functional and molecular
phenotyping of hiPSC-EHT under basal conditions and various stressors. The product of the
study will be an SOP-controlled assay with normal values±SD for hiPSC-EHT from healthy
volunteers and patients with heart diseases. The project could change clinical practice and be a
step towards individualized risk prediction and therapy of heart failure.
Genetics of Sudden Cardiac Death: A Shift From Mendelian to Complex Inheritance Paradigms
By Connie Bezzina
Sudden cardiac death from ventricular arrhythmias occurs in a broad spectrum of cardiac
pathologies and is an important cause of mortality. In the young sudden cardiac death usually
occurs in the setting of inherited familial cardiac disorders, which are classically divided into the
primary electrical disorders and the cardiomyopathies. Genetic studies have during the past 20
years markedly illuminated the genetic basis of these disorders, resulting in improvements in
patient care and an increased understanding of the underlying pathophysiological mechanisms.
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Yet, considerable challenges remain. These relate primarily to the large variability that typically
exists in disease severity among mutation carriers in affected families, which may point to a
more complex inheritance than previously appreciated and the necessity of new approaches to
further our understanding of these disorders.
Keeping up the balance: Role of HDACs in cardiac proteostasis and therapeutic implications
for atrial fibrillation
By Bianca Brundel
Cardiomyocytes are long-lived post-mitotic cells with limited regenerative capacity. Therefore,
its proper function depends critically on the maintenance of a healthy homeostasis of protein
expression, post-translational modification, function and degradation, together commonly
referred to as proteostasis. Impairment of proteostasis has a prominent role in the
pathophysiology of ageing-related neurodegenerative diseases including Huntington’s,
Parkinson’s, and Alzheimer’s disease. Emerging evidence reveals also a role for impaired
proteostasis in the pathophysiology of common human cardiac diseases including atrial
fibrillation (AF). Histone deacetylases (HDACs) have recently been recognized as key modulators
which control cardiac proteostasis by deacetylating various proteins. By deacetylating chromatin
proteins, including histones, they modulate epigenetic regulation of pathological gene
expression. Also, HDACs exert a broad range of functions outside the nucleus by deacetylating
structural and contractile proteins. The cytosolic actions of HDACs result in changed protein
function through post-translational modifications and/or modulation of their degradation. This
presentations summarizes mechanisms underlying the derailment of proteostasis in AF, and
subsequently focuses on the role of HDACs herein. In addition, the therapeutic potential of
HDAC inhibition to maintain a healthy proteostasis resulting in a delay in AF onset and
progression, is discussed.
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EVEN posters are presented in first session (lunch)
ODD posters in second session (tea)
List of Abstracts in alphabetical order of the presenting author. See page 7 for location of posters.
NR
presenting author
Title
22
Aman, Jurjan
6
100
Arik, Bahar
Aziz, Y.
7
84
Bedussi, Beatrice
Beishuizen, Cathrien
17
Bennis, Frank
62
47
Berg, Susan van den
Berkhemer, Olvert
57
85
Bezzina, Connie
Blom, M.T.
64
Bogaards, Silvia
66
Bollen, Ilse
56
Boly, Chantal
74
34
Boon, Reinier
Boshuizen, Marieke
The tyrosine kinase Arg/Abl2 mediates endothelial barrier
disruption by impairing integrin α5β1-mediated cell adhesion
Angiogenesis in the retina: studying tip cell regulation
Off-pump Left Ventricle Reconstruction using the Revivent
Myocardial Transcatheter Anchoring System in Patients with
severe Ischaemic Cardiomyopathy
Clearance of the brain by interstitial drainage into the ventricles
Internet-interventions targeting cardiovascular risk factors in
older people: a systematic review and meta-analysis
A machine-learning based analysis of arterial pressure waveform
derived parameters and thoracic bioimpedance for the
determination of central blood volume
Immune cell infiltration in obese brown adipose tissue
A randomized trial of intraarterial treatment for acute ischemic
stroke – MR CLEAN
Sudden cardiac arrest and rare genetic variants in the community
Improved survival after out-of-hospital cardiac arrest and use of
automated external defibrillators.
STORM superresolution microscopy to measure Z-disk and Aband width in human diaphragm muscle
Commonalities and differences between dilated and peripartum
cardiomyopathy; titin, fibrosis and myomesin
Impaired insulin-mediated myocardial microvascular recruitment
and cardiac function in early obesity are associated with
decreased activity of AMPK
Non-coding RNAs in cardiac aging
27
Bossche, Jan van den HDAC inhibition improves atherogenic macrophage function and
affects cellular metabolism
Bronzwaer, AnneEffect of neurohormonal blockade on postural hemodynamic
Sophie
response in patients with chronic heart failure
Bruggen, Cathelijne
Bone Morphogenetic Protein Receptor Type 2 Mutation in
van der
Pulmonary Arterial Hypertension, A view on the Right Ventricle
Brundel, Bianca
Exploration of the function of Class I and Class IIa HDACs in
experimental Atrial Fibrillation
Casadonte, Lorena
Variability in IV Adenosine-Induced Coronary Microvascular
Resistance and Systemic Pressure Compromises FFR
Determination despite Stable Stenosis Hemodynamics
68
12
71
32
MACROPHAGE LIPID LOADING LEADS TO TYPE I INTERFERON
HYPORESPONSIVENESS
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List of Abstracts in alphabetical order of the presenting author. See page 7 for location of posters.
NR
presenting author
Title
104
Casini, Simona
89
Cocera Ortega, Lucia
16
Da Silva Consalves
Bos, Denielli
24
37
Dalen, Jan Willem
van
Dallinga, Marchien
67
Damanafshan, Amin
82
44
Deel, Elza van
Dekker, Nicole
Electrophysiological effects of late sodium current inhibitor
GS967 in Scn5a-1798insD mouse and SCN5A-1795insD hiPSCderived cardiomyocytes
Optimizing cellular delivery of HCN2/SkM1 to induce long-term
biological pacing
Parasympathetic Nervous System Stimulation by Pyridostigmine
Improves Survival and Cardiac Function in Experimental
Pulmonary Arterial Hypertension
Cerebral perfusion and white matter hyperintensities in
hypertension older people
CD34 marks angiogenic tip cells in human vascular endothelial cell
cultures
An alternative splice variant of myotonic dystrophy protein kinase
induces loss of sarcomere structure and cardiomyopathy
29
Diemen, Jeske van
86
Elias, Joëlle
36
Ellatar, Mustafa
49
60
Emmens, Reindert
Eurelings, L.
63-2
Fuijkschot, Wessel
9
94-2
Happe, Chris
Heer, Frederieke de
95
Helmes, Michiel
42
Hermans, M.C.
25
Hibender, Stijntje
14
Hollander, Maurits
CARDIAC SUBSTRATE STIFFNESS AFFECTS SARCOMERE FUNCTION
Derangements of microcirculatory perfusion after cardiopulmonary bypass continue in the early postoperative period
EVENING INTAKE OF ASPIRIN IS ASSOCIATED WITH A MORE STABLE 24
HOUR PLATELET INHIBITION COMPARED TO MORNING INTAKE: A
STUDY IN CHRONIC ASPIRIN USERS
The Impact of the Location of a Chronic Total Occlusion in a noninfarct Related Artery on Long-term Mortality in ST-Elevation
Myocardial Infarction Patients.
Automatic Aortic Root Landmarks Detection in CTA images for
Transcatheter Aortic Valve Implantation Planning
Colchicine aggravates Coxsackievirus B3 infection in mice
The Prevention of Dementia by Intensive Vascular Care (preDIVA)
trial –preliminary blood pressure findings
Atherosclerotic plaque area increases after orthopedic surgery in
Apo E -/- mice
Characterization of TGF-β/BMP-axis in rat models of PH
AVIATOR Initiative: Aortic Valve Insufficiency and Ascending Aorta
Aneurysm International Registry.
Measuring the power generated by a cardiac myocyte in the
cardiac cycle
Continuous vital signs monitoring using the VitalConnect MD
Healthpatch
Resveratrol Inhibits Aortic Root Dilatation in the Fbn1C1039G/+
Marfan Mouse Model
Galectin-2 polarizes macrophages into an non-arteriogenic phenotype.
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List of Abstracts in alphabetical order of the presenting author. See page 7 for location of posters.
NR
presenting author
Title
79
Horst, Ester ter
81
Hu, Xu
2
Huis in 't Veld, A.E.
72
Jensen, Bjarke
31
Jong, Willeke
70
Jongsma, Susan
73
Joureau, Barbara
61
Kakkar, Vaishali
10
Kappers, Tessa
20
Klaassen, Ingeborg
56
Koenis, Duco
21
Kovacevic, Igor
99
Krijnen, Paul.
103
Kuster, Diederik
45
Lachkar, Nadia
96
Lahrouchi, N.
39
Liem, Madieke
p47phox-dependent reactive oxygen species stimulate nuclear
translocation of the transcription factor FOXO1 in ischemiachallenged H9C2 rat cardiomyocytes
The PARP1 inhibitor ABT888 accelerates recovery from contractile
dysfunction in experimental Atrial Fibrillation
The potential use of LA size and LA/RA ratio in discriminating
IPAH from PH-HFpEF
Title: A specified atrioventricular conduction system in the
American alligator (Alligator mississippiensis) heart.
Nlrp3 plays no role in acute cardiac infarction due to low cardiac
expression
Trial protocol: Healthy Ageing Through Internet Counselling in the
Elderly – the HATICE randomized controlled trial for the
prevention of cardiovascular disease, cognitive impairment and
dementia
Muscle fiber contractile measurements in nemaline myopathy
patients affected by ACTA1 mutations
Understanding the role of KBTBD13 in striated muscle function
and in nemaline myopathy
Dissecting cardiac and vascular contributions to pressure wave
travel and blood flow distribution
A role for PLVAP in the regulation of endothelial permeability and
angiogenesis
Genome-wide profiling of nuclear receptor Nur77 by ChIP-seq
reveals interplay with NF-kB and YY1 in regulation of
inflammatory signaling
The role of ubiquitination in regulation of endothelial barrier
function
The inflammatory infiltrate in the endomyocardial biopsy area is
increased significantly compared to the entire myocardium in
patients with lymphocytic myocarditis
In vivo pathophysiology of hypertrophic cardiomyopathy in a
highly prevalent 25 bp deletion mutation in MYBPC3
Optical Clearing Of Brain Tissue For Three-Dimensional
Ultramicroscopic Imaging Of The Cerebral Vasculature
Exome sequencing identifies a homozygous SLC22A5 mutation
leading to primary carnitine deficiency in a consanguineous
Moroccan family affected by hypertrophic cardiomyopathy and
sudden cardiac death
Use of antiplatelet agents is associated with intraplaque
hemorrhage on carotid MRI
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List of Abstracts in alphabetical order of the presenting author. See page 7 for location of posters.
NR
presenting author
Title
38
Lier, Monique van
78-1
Lodder, Elisabeth
76
Marchal, Gerard
33
Marcus, Tim
77
Marion, D.M.S. van
18
Marquering, Henk
90
Medzjkovic, Lejla
26
Meekel, Jorn
40
Minneboo, M.
35-1
Mohan, Rajiv
69
Najafi, Aref
83
Nederlof, Rianne
4
11
Neele, A.E.
Oost. Elco
53
Plug, Tom
Endocardial Prevalence of Coronary Collaterals Becomes
Transmurally More Uniform in a Human Heart with Left
Ventricular Hypertrophy
Homozygous Mutations in GNB5 and SYDE2 in patients with
autosomal recessive bradycardia
Electrical and structural abnormalities in homozygous Scn5a1798insD mutant embryos.
Increased native T1-values at the interventricular insertion
regions of precapillary pulmonary hypertension patients
Novel HSP-inducing Compounds to Restore Cardiomyocyte
Function in Experimental Atrial Fibrillation
Difference of Thrombus Attenuation between CT and CT
Angiography Measured in Patients with Acute Ischemic Stroke: An
expression of Thrombus Porosity?
Orphan nuclear receptor Nur77 affects cardiomyocyte calcium
homeostasis and adverse cardiac remodelling.
Establishment of viable ex vivo human and calf aortic tissues as
basis for research on the pathophysiology of aortic aneurysms.
Secondary prevention for cardiovascular disease is challenging
and differs between ethnic groups: the HELIUS study
Early specification and progressive restriction of the cardiac
conduction system lineage during heart development
Selective phosphorylation of PKA targets after β-adrenergic
receptor stimulation impairs myofilament function in Mybpc3targeted HCM mouse model
Cyclophilin D ablation protects the heart against infarction and is
associated with increased end-ischemic mitochondrial hexokinase
activity
H3K27 methylation in macrophages and atherosclerosis
Microvasculature Segmentation in Cardiac Cryomicrotome
Images
65
Podliesna, Svitlana
101
Portero, Vincent
19
Preckel, Benedikt
THROMBIN-ACTIVATABLE FIBRINOLYSIS INHIBITOR (TAFI) PEPTIDES
SELECTIVELY MODULATE THROMBOMODULIN-DEPENDENT TAFI
ACTIVATION
HEY2, a novel susceptibility gene for Brugada Syndrome, controls
depolarization and repolarization gradients in the RVOT and
across the ventricular wall
Microtubule-associated protein RP/EB family member 1
modulates cardiac conduction
Effect of helium pre- or postconditioning on signal transduction kinases
in patients undergoing coronary artery bypass graft surgery
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List of Abstracts in alphabetical order of the presenting author. See page 7 for location of posters.
NR
presenting author
Title
1
Radic, Ozren
28
92
Rijswijk, Jan Willem
van
Rivaud, Mathilde
Identification of endothelial progenitor cells (EPC) in in relation to
angiogenic response of vascular malformations and in angiomas
46
15
Rol, Nina
Santos, Emilie
23
Schwartz, Janina
98
Sergeeva, Irina
13
Smeets, Esther
8
Smit, Kirsten
5
Smits, Josien
48
63-1
Stam, Olga
Stienen, Ger
87
Stienen, Susan
50
Szulcek, Robert
78-2
Tiel, Claudia van
41
Trieu, Michelle
43
Truijen, Jasper
59
Turaihi, Ali
LVAD-support Induced Hemodynamic Changes Result in Valve
Interstitial Cell Activation and a Reparative Response in the Aortic Valve
Remodeling of distinct sodium channel pools in the murine failing
heart
Balancing TGFb/BMP in Pulmonary Arterial Hypertension
Absolute thrombus density measurement on non-contrast CT in
patients with acute ischemic stroke has superior Interobserver
agreement than relative ratio
High-Resolution 3D Imaging of Vessels in the Ischemic Rat Hind
Limb for Collateral Quantification
A regulatory domain controls the conserved Nppa-Nppb gene
cluster during heart development and stress
T cell regulator Cbl-b aggravates atherosclerosis via CD8+ T cell
induced macrophage death
Plasma from human volunteers subjected to remote ischemic
preconditioning protects human endothelial cells from hypoxiainduced cell damage.
Endothelial Cells from different vascular beds of Pulmonary
Arterial Hypertension patients show a delayed response to supraphysiological shear stress
Intraleaflet Hemorrhage in Cardiac Valves
Right ventricular hypertrophy and failure are associated with
mitochondrial complex I dysfunction and altered NADH kinetics
Absolute Discharge Levels or Relative Reduction in NT-proBNP
during admission for acute decompensated heart failure as a
target to reduce 6-Month Mortality?
Supra-physiological shear stress unveils novel dysfunction in
microvascular and circulating endothelial cells from patients with
Pulmonary Arterial Hypertension
CD40-Filamin A interactions are required for translocation of CD40 to
lipid rafts in endothelial cells and for endothelial cell activation
Administration of an Angiopoietin-1 mimetic attenuates
microcirculatory perfusion disturbances and vascular leakage
during hemorrhagic shock
Cerebral Autoregulation and the Cerebrovascular Response to
Head-of-Bed Positioning in Acute Ischemic Stroke
The Role of IRS2 in PVAT in the Regulation of Local Muscular
Perfusion and Glucose Uptake
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List of Abstracts in alphabetical order of the presenting author. See page 7 for location of posters.
NR
presenting author
Title
35-2
Uiterwijk, M.
30
In-situ tissue engineering of vascular grafts: Investigate neo-tissue
formation and degradation velocity of bisurea polymeric vascular grafts
Valerio, L
75
Veerman, Christiaan
52
Verkaik, Melissa
The impact of family history on cardiovascular disease is higher
among hypertensive individuals of African and Surinamese
background
Electrophysiological characteristics of iPSC-derived
cardiomyocytes from Brugada syndrome patients without SCN5A
mutations
51
Wanga, Shaynah
94-1
Weerd, Henk van
93
Wiersma, M.
3
Wijk, A.E. van der
97
Wijnker, Paul
88
91
Winter, Josine de
Woudstra, Linde
55
Wust, Rob
102
Zhang, Deli
54
Zonneveld, TP
80
Zweerink, A
FGF23 IN PART EXPLAINS IMPAIRED ENDOTHELIAL FUNCTION AND
MYOCARDIAL PERFUSION OBSERVED IN EXPERIMENTAL CHRONIC
KIDNEY DISEASE
Calcification associates with aortic distensibility in Marfan
Syndrome mice.
A large permissive regulatory domain controls Tbx3 expression in
the cardiac conduction system
The orphan drug 4-phenyl butyrate protects against Atrial
Fibrillation by blocking ER stress-induced autophagy
Modulation of cAMP Signaling Prevents TNFα-induced Endothelial
Barrier Disruption in an In Vitro Model of the Blood-Retinal
Barrier
Expression of missense and truncating MYBPC3 mutations does
not prevent hypercontractility of Mybpc3-targeted knock-out
mouse engineered heart
KBTBD13: A NOVEL GENE IMPLICATED IN CARDIOMYOPATHY
Lymphocytic myocarditis can lead to inflammation in the
coronary arteries which may induce an acute myocardial
infarction
Rapid uptake and slow restitution of mitochondrial calcium in
adult rat cardiomyocytes
PARP1 activation causes NAD+ depletion and contractile
dysfunction in experimental and human Atrial Fibrillation
The ThRombolysis in UnconTrolled Hypertension (TRUTH) study:
an observational study on treatment strategy of elevated blood
pressure in stroke patients eligible for IVT
Correcting QRS duration for Left Ventricular dimension improves
current patient selection for Cardiac Resynchronization Therapy
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Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A1
Identification of endothelial progenitor cells (EPC) in in relation to angiogenic response of
vascular malformations and in angiomas
Ozren Radic, Andrea Dorresteijn, Onno de Boer, Lorine Meijer-Jorna, Hans Niessen*, Allard van
der Wal
Departments of Pathology, VUMC* and AMC, Amsterdam
Objectives:
Immature infantile hemangiomas (IH) are treated successfully with Propranolol, likely interacting
with lesional endothelial progenitor cells through the CXCR4-SDF-1alpha pathway. The
occurrence of endothelial progenitor cells in vascular malformations is unknown. In this study,
we evaluated the presence and immunophenotype of endothelial progenitor cells in vascular
malformations relation to the angiogenic response of symptomatic congenital arteriovenous
malfortmation (AVM).
Methods:
Paraffin-sections from IH and AVM lesions . of skin (N = 22). were immune(double) stained with
the following antibodies: GLUT-1(angioma marker), Ki67 (cell proliferation marker), von
Willebrand Factor and CD34 (endothelial cells) and. CXCR4, SDF-1α and CD133 as endothelial
progenitor cell markers (EPC). Staining patterns were quantified with the use of image analysis
software, and differences between IH and AVM analysed statistically
Results:
CD133 /CD34 proved to be a good marker for EPC, which were found in IH and in AVM, in the
vasoproliferative (high Ki67 index) and in mature vascular areas (low ki67 index) of both types of
lesions. There were no significant differences in EPC densitiy between IH and AVM (P=0,638).
CxCR4 / SDF-1alpha was widely expressed throughout the endothelium both lesions in all cases,
which was confirmed n double stains. of IH and AVM
Conclusion:
SimIlar to proliferative IH, CD133+ CxCR4+ SDF-1alpha + EPC can be identified also in
vasoproliferative areas of AVM. The results suggest that the drug propanol could be effective in
the treatment of AVM with a vasoproliferative component.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A2
The potential use of LA size and LA/RA ratio in discriminating IPAH from PH-HFpEF
A.E. Huis in ’t Veld¹, M.L. Handoko², O.A. Spruijt¹, H.J. Bogaard¹, A. Vonk Noordegraaf ¹
¹Department of Pulmonology, VU University Medical Center, Amsterdam, The Netherlands. ²Department of
Cardiology, VU University Medical Center, Amsterdam, The Netherlands.
Introduction:
Discriminating pulmonary hypertension (PH) due to heart failure with preserved ejection fraction
(HFpEF-PH) from idiopathic pulmonary arterial hypertension (IPAH) remains an important
diagnostic challenge.
Methods:
The aim of this retrospective, proof-of-concept analysis was to determine whether left atrial (LA)
size, right atrial (RA) size and LA/RA ratio throughout the cardiac cycle, assessed by cardiac
magnetic resonance (CMR), can potentially help to differentiate between IPAH and HFpEF. In 20
consecutive treatment-naive patients, referred to the VUMC for evaluation of new suspected
PAH (10 IPAH and 10 PH-HFpEF), LA and RA area was assessed by manually tracing endocardial
contours in multiple slices representing the cardiac cycle, in a 4-chamber view.
Results:
The mean PCWP was 8 ±3 in the PAH group and 18 ± 3 in the PH-HFpEF group (mean; ± SD). LA
size was 1579mm² ± 551 in the PAH group versus 2786 mm² ± 655 in the HFpEF group
(p=0,0001)( fig 1.). When indexed for right atrial size (LA/RA ratio, fig. 2), an LA/RA ratio of >1 was
observed in all but one PH-HFpEF patient, over the entire cardiac cycle.
Conclusion:
CMR-derived LA/RA ratio> 1 may be a useful noninvasive indicator of HFpEF in patients evaluated
for PAH. The finding that LA size is larger than RA size throughout the cardiac cycle, holds
promise for the use of non-ECG gated CT-derived LA/RA ratio as a potential tool to discriminate
HFpEF-PH from PAH.
Figure 1
Figure 2
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A3
Modulation of cAMP Signaling Prevents TNFα-induced Endothelial Barrier Disruption in an In
Vitro Model of the Blood-Retinal Barrier
Anne-Eva van der Wijk, Cornelis J.F. van Noorden, Ingeborg Klaassen, Reinier O. Schlingemann
Ocular Angiogenesis Group, Department of Ophthalmology and Department of Cell Biology and Histology, Academic
Medical Center.
Introduction:
Breakdown of the BRB is an important feature in the pathology of diabetic macular edema
(DME), but cellular mechanisms underlying BRB dysfunction are poorly understood. In recent
years, the involvement of inflammatory cytokines in BRB breakdown has gained attention. In the
present study, we investigated the effects of TNFα and cAMP on the BRB in vitro.
Methods:
For this study we employed an in vitro BRB model, consisting of primary bovine retinal
endothelial cells (BRECs) grown on Transwell filters. Barrier integrity was assessed by
measurements of trans-endothelial electrical resistance (TEER) and permeability to molecular
tracers. mRNA levels and protein expression of BRB-specific components were investigated by
qPCR and immunostaining.
Results:
TNFα disrupted the BRB in vitro at the molecular level. Stimulation of BRECs with TNFα (10
ng/ml) caused a significant decrease in expression of the tight junction components claudin-5
and ZO-1. The changes in tight junctions were reflected by increased permeability to a small
molecular tracer after TNFα stimulation. Interestingly, TNFα significantly decreased intracellular
cAMP levels. When BRECs were preincubated with a membrane permeable cAMP analog, the
effects of TNFα on tight junctions and permeability were mitigated. In addition, basal TEER levels
were higher and permeability was lower in BRECs treated with cAMP.
Conclusions:
Together these results demonstrate that cAMP has a stabilizing effect on the basal barrier
properties of retinal endothelial cells, and importantly, that pre-incubation with cAMP can
prevent TNFα-induced barrier breakdown in our in vitro BRB model. In conclusion, this study
shows that cAMP is essential in the maintenance of the BRB.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A4
H3K27 methylation in macrophages and atherosclerosis
Neele, A.E., van der Velden, S., Gijbels, M.J.J., Hoeksema, M.A., Boshuizen, M.C.S. Van den
Bossche, J., de Winther, M.P.J.
Department of Medical Biochemistry, Academic Medical Center, Amsterdam.
Introduction:
Monocytes and macrophages are key immune cells in the development of atherosclerosis and
clinical outcome. Epigenetic pathways govern many aspects of monocyte and macrophage
differentiation and activation. This is regulated by a large number of histone modifying enzymes
(HMEs) that regulate activating- (e.g. H3K4me3, H3K27Ac) or repressive- (e.g. H3K27Me3)
histone marks. The H3K27 demethylase Jumonji-domain 3 (Jmjd3) is induced upon LPS activation
in macrophages and was shown to be involved in both M1/M2 polarization. The role of its
counteracting partner, the methyltransferase Enhancer of the zeste homolog 2 (Ezh2) however
remains unknown.
Methods:
To study the function of Ezh2 we generated a myeloid specific Ezh2 deficient mice (LysMCre fl/fl).
To validate the role of Ezh2 in atherosclerosis we performed a bone marrow transplantation to
LDLr-/- mice and compared WT (Cre-) to Ezh2 deficient (Cre+) transplanted mice after 9 weeks of
high fat diet.
Results:
Using bone marrow derived macrophages we found that Ezh2 is down regulated upon LPS
activation and oxidized LDL induced foam cell formation. Moreover, human unstable plaques
show an up regulation of Ezh2 compared to stable advanced plaques. To study Ezh2 function we
generated a myeloid specific Ezh2 deleted mice and performed an atherosclerosis experiment.
We observe a 20% reduction in plaque size of the Ezh2 deficient transplanted mice. Plaque
composition is similar between groups, only neutrophil counts are lower indicating that the
plaques of Ezh2 deleted mice are not only smaller but also less inflammatory. We also observe
reduced foam cells formation in peritoneal macrophages of these mice. Currently, we are
investigating the mechanisms by which macrophage Ezh2 regulates macrophage activation and
atherosclerotic plaque development.
Conclusion:
Macrophage Ezh2 deficiency reduces foam cell formation and atherosclerotic lesion
development.
Supported by the GENIUS project (CVON2011-19).
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A5
Endothelial Cells from different vascular beds of Pulmonary Arterial Hypertension patients
show a delayed response to supra-physiological shear stress
J. Smits, R. Szulcek, D. Tasev, N.Rol, A. Vonk- Noordegraaf, P. Koolwijk, HJ. Bogaard.
Department of Physiology and Department of Pulmonology, Institute for cardiovascular research (ICaR-VU), VU
University Medical Centre, Amsterdam.
Rational:
Pulmonary arterial hypertension (PAH) is a deadly lung disease in which progressive increase of
resistance in the vasculature leads to altered pulmonary hemodynamics and increased shear
stress. Previous research done in our laboratory showed that pulmonary micro-vascular
endothelial cells (MVEC) from patients with PAH show a dysfunctional alignment response to
pathological high shear stress (HSS). ECFC are circulating EC easily obtainable from peripheral
blood and play a central role in vascular repair. As lung biopsies are extremely scarce, we aim to
test whether blood-derived Endothelial Colony Forming Cells (ECFC) mimic the EC dysfunction
seen in PAH.
Methods:
ECFC were isolated from whole blood of patients with PAH and healthy controls. ECFC were
subjected to HSS (21-25 dyn/cm²) and low shear stress (LSS, 5 dyn/cm²) for 120 hrs.
Morphological response to flow was analyzed and proliferation rates were determined over a
period of 8 days.
Results:
ECFC were isolated from 28 patients with PAH from which 19 out of 28 isolations were
successful. Age (p=0.04) correlated with the amount of outgrowth. However, patients without
ECFC outgrowth were generally older (42.8±12.3 vs. 53.4±15.9 yrs). Subsequently, ECFC of
healthy controls and PAH patients were subjected to pulsatile shear stress (n=8).Under LSS both
control and PAH ECFC remained in a non-shear adapted state. After 72 hrs of HSS inter-donor
heterogeneity occurred, showing 4 out of 8 PAH donors unable to align to HSS. The remaining 4
donors aligned normally.
Conclusion:
ECFC present with a dysfunctional reaction to flow comparable to lung biopsy derived MVEC, and
can therefore be used to further study the involved mechanisms. However, inter-donor
heterogeneity occurs,
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A6
Angiogenesis in the retina: studying tip cell regulation
Bahar Yetkin-Arik, Ingeborg Klaassen, Cornelis J.F. Van Noorden, Reinier O. Schlingemann
Ocular Angiogenesis Group, Department of Ophthalmology and Department of Cell Biology and Histology, Academic
Medical Center.
Introduction:
Tip cells are the initiator of the angiogenic process and comprise a distincte
ubpopulation of endothelial cells with an unique molecular signature and thereby forming an
attractive target for angiogenic therapy. Therefore, in the present study we explored the
regulation of genes that are involved in tip cell function.
Methods:
In previous research we have shown that CD34 can be used as a tool to select tip cells in vitro. To
investigate the gene regulation in tip cells, a microarray analysis was performed in CD34 positive
and CD34 negative cells. The genes that where upregulated in tip cells were validated and
confirmed by quantitative polymerase chain reaction (qPCR) analysis in Human Umbilical Vein
Endothelial Cells (HUVECs) and human Microvascular Endothelial Cells (hMVECs). The effect of
these genes on CD34 fraction and sprouting were determined by flow cytometric analysis and the
spheroid-based angiogenesis model, respectively.
Results:
Inhibition of a small subset of 6 genes showed no effect on the number of tip cells, both in
HUVECs and HMVECs, whereas silencing of 4 of these genes showed a significant effect on the
number of sprouts and sprout length in HUVECs.
Conclusion:
Gene silencing of selective genes identified with our novel in vitro tip cell model did not have an
effect on the number of tip cells, but showed a significant effect on vascular sprouting. This
suggests that these genes play an important role in angiogenesis and we will explore the role as a
possible therapeutic target of these genes in our future research.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A7
CLEARANCE OF THE BRAIN BY INTERSTITIAL DRAINAGE INTO THE VENTRICLES
Beatrice Bedussi1*, Monique GJTB van Lier1, Jonas W. Bartstra1, Judith de Vos1, Maria Siebes1, Ed
VanBavel1 and Erik NTP Bakker1.
1
Dept. of Biomedical Engineering and Physics, Academic Medical Center, Amsterdam. *[email protected]
Objectives:
Alzheimer’s disease (AD) and cerebral amyloid angiopathy (CAA), are associated with failure of
amyloid-β removal. The aim of this study is to visualize the removal of tracers from the rodent
brain, focusing on ventricular, subarachnoid and perivascular space (PVS) as possible transport
pathways.
Methods:
We performed confocal fluorescence imaging of horizontal mouse brain sections after 30 min of
controlled infusion of a mix of FITC-dextran (MW 500kD) and Texas Red-dextran (MW 3kD) into
the cisterna magna (CM, 5μL) or striatum (2μL). We obtained 3D reconstructions of the mouse
brain and vasculature, using an imaging cryomicrotome.
Results:
After infusion into the striatum, dyes spread in the parenchyma and found their way to the
closest lateral ventricle. Then, they reached the ventricular system, cisterns and subarachnoid
space (SAS). Following CM infusion, dyes dispersed throughout the SAS, the cisterns and along
the PVS. The high molecular weight dye remained confined to the SAS and PVS of cortical vessels,
whereas the small dye also crossed the pia mater into the adjacent parenchyma. However,
neither dye could be detected in the ventricular system. Despite earlier reports , there was no
association of dyes with capillaries or veins in the parenchyma in either group.
Conclusion:
These data reveal a flow of interstitial fluid from the parenchyma to the ventricular system, from
where dyes reach the SAS via the cerebrospinal fluid (CSF). Dyes disperse from the SAS along PVS,
and partly reenter the parenchyma. Failure in this transport pathway could hinder the
physiological drainage of amyloid-β and play a role in AD and CAA.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A8
Plasma from human volunteers subjected to remote ischemic preconditioning protects human
endothelial cells from hypoxia-induced cell damage.
Nina C. Weber1, Isabelle Riedemann1, 2, Kirsten F Smit, Karina Zitta2, Djai van de Vondervoort1,
Coert J. Zuurbier1, Benedikt Preckel1, Markus W. Hollmann1, Martin Albrecht2
1
Department of Anaesthesiology, Laboratory of Experimental Intensive Care and Anaesthesiology (L.E.I.C.A.),
2
Academic Medical Centre (AMC), University of Amsterdam. Department of Anaesthesiology and Intensive Care
Medicine, University Hospital Schleswig-Holstein, Campus Kiel, Kiel, Germany
Introduction:
Short repeated cycles of peripheral ischemia/reperfusion (I/R) can protect distant organs from
subsequent prolonged I/R injury; a phenomenon known as remote ischemic preconditioning
(RIPC). A RIPC-mediated release of humoral factors might play a key role in this protection and
vascular endothelial cells are potential targets for these secreted factors. In the present study,
RIPC-plasma obtained from healthy male volunteers was tested for its ability to protect human
umbilical endothelial cells (HUVEC) from hypoxia-induced cell damage.
Methods:
10 healthy male volunteers were subjected to a RIPC-protocol consisting of 4 x 5 min
inflation/deflation of a blood pressure cuff located at the upper arm. Plasma was collected
before (T0; control), directly after (T1) and 1 h after (T2) the RIPC procedure. HUVEC were
subjected to 24 h hypoxia damage and simultaneously incubated with 5 % of the respective RIPCplasma. Cell damage was evaluated by lactate dehydrogenase (LDH)-measurements. Western
blot experiments of hypoxia inducible factor 1 alpha (HIF1alpha), phosphorylated signal
transducer and activator of transcription 5 (STAT5), protein kinase B (AKT) and extracellular
signal-related kinase 1/2 (ERK-1/2) were performed.
Results:
Hypoxia-induced cell damage was significantly reduced by plasma T1 (p=0.02 vs T0). The
protective effect of plasma T1 was accompanied by an augmentation of the intracellular
HIF1alpha (p=0.01 vs T0) and increased phosphorylation of ERK-1/2 (p=0.03 vs T0).
Phosphorylation of AKT and STAT5 remained unchanged.
Conclusion:
RIPC plasma protects endothelial cells from hypoxia-induced cell damage and humoral mediators
as well as intracellular HIF1alpha may be involved.
Published in: Basic Res Cardiol. 2015 Mar;110(2):17. doi: 10.1007/s00395-015-0474-9.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A9
Characterization of TGF-β/BMP-axis in rat models of PH
C.M. Happé1,2, N. Rol1,2, D. Bos1,2, C.E.E. van der Bruggen2, F.S. de Man2, H.J. Bogaard2.
1
2
VU University medical center, Amsterdam, Department of Physiology and Pulmonology .
Introduction:
Mutations in the bone morphogenetic protein recptor type 2 (BMPR2) comprise a large portion
of familial Pulmonary Arterial Hypertension (PAH) cases. The transforming growth factor-β (TGFβ)/BMP-axis in PAH has been of interest, which is hypothesized to favor TGF-β signaling due to
defective BMPR2 signaling, consequently leading to pro-proliferative signaling within the lung. In
addition, it has been proposed that the BMPRII mutations might affect cardiac adaptation. To
date none of the available animal models have been fully characterized with regard to the TGFβ/BMP pathway. This study assessed the lung and heart TGF-β/BMP-axis in multiple rat animal
models of pulmonary hypertension to ensure translational capability.
Methods:
Heart and lung TGF-β/BMP-axis was assessed by qPCR, western blot and immunofluorescence in
the the monocrotaline (MCT), Sugen-hypoxia (SuHx), Sugen-Pneumonectomy (SuPnx) and
Pulmonary artery banding model (PAB) and compared to control and PAH patient tissues.
Circulating ligands, TGF-β receptor (TGFβR) type 1 and 2 and BMPR2, and canonical downstream
signaling (Smad2/3, Smad1/5/8, and transcription factors) were investigated.
Results:
BMPR2 was down-regulated at both transcription and protein levels in the lung of all PH animal
models (p<0.05). Transcription of pulmonary TGFβR1 and -2 were increased in the SuPNx-model,
compared to control (P<0.001). In both SuHx and SuPnx models an increase in protein Smad2/3
expression was observed by immunofluorescence implying overactivation of TGF-β signaling.
Cardiac TGFβR1 was decreased in PAB model, compared to control (P<0.05), while TGFβR2 was
decreased in both the MCT and PAB model.
Conclusion:
Early indications reveal differences between several pulmonary hypertension animal models,
with regard to the TGF-β/BMP pathway. Additional analysis is needed to fully characterize the
regulation of TGF-β and BMP in the rat models.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A10
Dissecting cardiac and vascular contributions to pressure wave travel and blood flow
distribution
Tessa Kappers1,2,3,4, Frank C. Bennis1,2,3,4, Björn J.P. van der Ster2,3,4, Wim J. Stok3,4,
Berend E. Westerhof4,5, Johannes J. van Lieshout2,3,4,6
1
2
3
University of Twente, Department of Technical Medicine. Department of Internal Medicine, Department of
4
Anatomy, Embryology and Physiology, Academic Medical Center, Amsterdam. Laboratory for Clinical Cardiovascular
5
Physiology, Center for Heart Failure Research, Academic Medical Center, Amsterdam. Edwards Lifesciences BMEYE,
6
Amsterdam. MRC/Arthritis Research UK Centre for Musculoskeletal Ageing Research, School of Life Sciences,
University of Nottingham Medical School, Queen's Medical Centre, United Kingdom.
Introduction:
The pressure wave generated by the pumping heart is reflected by the arterial system. The
traditional concept is that arterial pressure augmentation by reflection as quantified by the
Augmentation Index (AIx) originates exclusively from the arterial vasculature. However, we
showed that during standing AIx decreased while TPR increased,1 an opposite response,
suggesting other underlying causes. We investigate the influence of heart action.
Methods:
The impact of reduced preload to the heart was determined in healthy volunteers with passive
head-up tilt (HUT) and subatmospheric pressure applied to the lower body (LBNP). Both result in
progressive central blood volume depletion and sympathetic activation, however, with and
without hydrostatic effects, respectively. Continuous plethysmography derived finger arterial
pressure was measured and with a transfer function modeled to brachial artery pressure. Carotid
and femoral artery pulse wave form and arterial pulse wave velocity (PWV) were determined by
applanation tonometry. At least 40 consecutive tonometry determined waves during baseline
and between the third and fifth minute of, respectively, HUT and LBNP are analyzed in each
subject. Waveform characteristics are compared with vascular (TPR, PWV) and cardiac (heart
rate, stroke volume, and contractility) parameters.
Results and preliminary conclusion:
Analysis of a cardiac vs. a vascular contribution to the pressure waveform is in progress.
Sympathetic activation by LBNP reduced arterial pressure augmentation whereas TPR increased.
Reduced preload may have changed the forward wave shape such that the summation of the
reflected wave is less apparent.
1) van den Bogaard B et al. Arterial wave reflection decreases gradually from supine to upright. Blood Press.
2011;20:370-5.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A11
Microvasculature Segmentation in Cardiac Cryomicrotome Images
Elco Oost, Jeroen PHM van den Wijngaard, Maria Siebes
Department of Biomedical Engineering & Physics, Academic Medical Center, University of Amsterdam.
References:
[1] Spaan JA, et al. (2005). Visualization of intramural coronary vasculature by an imaging cryomicrotome suggests
compartmentalisation of myocardial perfusion areas. Med Biol Eng Comput 43, 431-435.
[2] Lesage D, et al. (2009). A review of 3D vessel lumen techniques: Models, features and extraction schemes. Med Imag Anal 13,
819-845.
[3] Sato Y, et al. (1998). Three-dimensional multi-scale line filter for segmentation and visualization of curvilinear structures in
medical images. Med Imag Anal 2, 143-168.
Introduction:
Three-dimensional cryomicrotome imaging facilitates the analysis of the (micro)vasculature of a
dissected organ [1]. Organ vessels are flushed and subsequently filled with a fluorescent material.
After freezing the sample, it is cut into thin slices in the cryomicrotome. After each slice, a highresolution episcopic image of the fluorescent material is acquired. From the resulting stack of 2D
images a high resolution (up to 5x5x5 µm) 3D representation of the vasculature is reconstructed.
Subsequent vessel segmentation facilitates analysis and quantification of neovascularization and
vascular remodeling. Three groups of automatic vessel segmentation techniques are found in
literature: vascular model based algorithms, (multi-scale) vascular feature based segmentation,
and vessel extraction schemes [2].
Methods:
A hybrid vessel segmentation algorithm is proposed, exploiting the benefits of feature based
vessel segmentation and region growing (a vessel extraction scheme), while avoiding their
drawbacks. By calculating vesselness features (Sato et al. [3]) at multiple scales, a vesselness
hyperspace is created. Subsequently, the region growing algorithm propagates through this
multi-scale vesselness representation, being allowed to visit only the voxels with a high
vesselness probability.
Results:
Because of the incorporation of knowledge about vesselness, the region grower is able to
segment the vasculature, without leakage into the image background, being robust to image
artifacts, and in reasonable time. Furthermore, there are no apparent performance differences in
segmenting smaller or larger vessels.
Conclusions:
A knowledge based automatic vessel segmentation algorithm is presented. The method shows
accurate and robust segmentation results in cryomicrotome images, enabling the 3D analysis of
vascular networks.
Acknowledgements: This research was supported by the Center for Translational Molecular Medicine (EMINENCE).
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A12
Bone Morphogenetic Protein Receptor Type 2 Mutation in Pulmonary Arterial Hypertension, A
view on the Right Ventricle
Cathelijne E.E. Van Der Bruggen1, CM Happé1,2, P Trip1, OA Spruijt1, N Rol1,2, AC Houweling, MD,
PhD3, B Girerd, PhD4,5,6, P Dorfmuller, MD, PhD4,5,6, O Mercier, MD, PhD7, M Humbert, MD,
PhD4,5,6, ML Handoko, MD, PhD8, J Van Der Velden, PhD2, A Vonk Noordegraaf, MD, PhD1, HJ
Bogaard, MD, PhD1, MJ Goumans, PhD9, FS De Man, PhD1,
1
Department of Pulmonary Medicine, Institute for Cardiovascular Research, VU University Medical Center,
2
Amsterdam. Department of Physiology, Institute for Cardiovascular Research, VU University Medical Center,
3
4
Amsterdam. Department of Clinical Genetics, VU University Medical Center, Amsterdam. Univ. Paris-Sud, Le
5
Kremlin-Bicêtre, France. AP-HP, Service de Pneumologie, Centre de Référence de l’Hypertension Pulmonaire Sévère,
6
DHU Thorax Innovation, Hôpital Bicêtre, Le Kremlin-Bicêtre. INSERM999, LabEx LERMIT, Centre Chirurgical Marie
7
Lannelongue, Le Plessis Robinson, France. Department of Thoracic and Vascular Surgery and Heart-Lung
8
Transplantation, Hôpital Marie-Lannelongue, Le Plessis Robinson, Paris-Sud University, France. Department of
9
Cardiology, Institute for Cardiovascular Research, VU University Medical Center, Amsterdam. Department of
Molecular Cell Biology, Laboratory of Experimental Cardiology, Leiden University Medical Center, Leiden.
Background:
The effect of a mutation in the bone morphogenetic protein receptor 2 gene (BMPR2) gene on
right ventricular (RV) pressure overload in patients with pulmonary arterial hypertension (PAH) is
unknown. Therefore, we investigated RV function in PAH-patients with and without BMPR2
mutation by combining in vivo measurements with molecular and histological analysis of human
RV and left ventricular (LV) tissue.
Methods & Results:
In total, 97 idiopathic or familial PH patients were genetically screened for the presence of a
BMPR2 mutation: 28 patients had a BMPR2 mutation, 69 patients did not have a BMPR2
mutation. In vivo measurements were assessed using right heart catheterization (RHC) and
cardiac magnetic resonance imaging. Despite a similar mean pulmonary artery pressure
(noncarriers 54±15 vs. mutation carriers 55±9 mmHg) and pulmonary vascular resistance (755
(483-1043) vs. 931 (624-1311) dynes*s/cm⁵ ), mutation carriers presented with a more severely
compromised RV function (RV ejection fraction: 37.6±12.8 vs. 29.0±9 %: p<0.05, cardiac index
2.7±0.9 vs. 2.2±0.4 l/min/m²). Differences continued to exist after treatment. To investigate the
role of TGF-β and BMPRII signaling, human RV and LV tissue were studied in controls (n=6),
mutation carriers (n=5) and non-carriers (n=11). However, TGF-β and BMPRII signaling, as well as
hypertrophy, fibrosis, capillary density and inflammation were similar between mutation carriers
and non-carriers.
Conclusion:
Despite a similar afterload, RV function is more severely affected in mutation carriers compared
to non-carriers. However, these differences cannot be explained by a differential TGF-β , BMPRII
signaling or cardiac adaptation.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A13
T cell regulator Cbl-b aggravates atherosclerosis via CD8+ T cell induced macrophage death
Esther Smeets*, Tom Seijkens*, Myrthe den Toom*, Svenja Meiler*,Esther Lutgens*,§
*
Department of Medical Biochemistry, Experimental Vascular Biology, Academic Medical Center, University of
§
Amsterdam, Amsterdam. Institute for Cardiovascular Prevention (IPEK), Ludwig Maximilians University, Munich,
Germany
Introduction:
T-cells play a major role in atherosclerosis. An important negative regulator for peripheral T cell
activation that is also expressed in macrophages is Cbl-b. In human atherosclerosis, CBL-B
expression is decreased in atherosclerotic plaques compared to normal arteries.
Methods
Here we studied the effect of Cbl-b deficiency on atherogenesis in 20 wks old Cbl-b-/-Apoe-/- and
Apoe-/- mice that exhibited early atherosclerotic lesions in the aortic arch, and advanced lesions
in the aortic root.
Results:
In early stages of atherosclerosis, Cbl-b deficient mice showed an 82% increase in plaque size due
to an increase in plaque macrophage content. Cbl-b deficient macrophages showed a strong
migratory response towards MCP-1, exhibited an increase in oxLDL uptake, and had proinflammatory, M1-like characteristics.
In advanced stages of atherosclerosis, where Cbl-b deficient mice also exhibited a 40% increase in
plaque area, plaque phenotype had changed more dramatically. Plaques contained less
macrophages, larger necrotic cores, and an abundance of CD8+ T-cells. In Cbl-b-/-Apoe-/- mice,
CD8+ T-cells were less susceptible to apoptosis and insensitive to Treg suppression. Their
abundance aggravated macrophage apoptosis, resulting in enhanced necrotic core formation and
thus plaque progression. These data emphasize the importance of CD8+ T cells in the propagation
of atherosclerosis.
Conclusion:
In conclusion, we show that Cbl-b is an important regulator of both innate and adaptive immune
reactions in atherosclerosis by mediating macrophage recruitment and activation, CD8+ T-cell
activation, and CD8+ T-cell induced macrophage death.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A14
Galectin-2 polarizes macrophages into an non-arteriogenic phenotype
M. Hollander1, C. Yildrim2, A. Van Der Laan3, DYS. Vogel4, JM. Baggen2, T. Van Der Pouw-Kraan2,
AJG. Horrevoets2, N. Van Royen1
1
2
Department of Cardiology, VU University Medical Center, Amsterdam, Department of Molecular Cell Biology and
3
Immunology, VU University Medical Center, Amsterdam, Department of Cardiology, Academisch Medicsch
Centrum, Amsterdam,
Background: The carbohydrate-binding lectinGalectin-2 (Gal-2)is expressed by monocytic cells
and is known to mediate immune responses and associated with impaired collateral artery
growth (arteriogenesis) in patients with coronary artery disease. Furthermore, systemic
treatment with Gal-2 significantly decreases perfusion restoration after femoral artery ligation in
mice and lowers the quantity of perivascular macrophages, suggesting Gal-2 alters arteriogenesis
by modulating macrophage behavior. Interestingly, macrophages are known to polarize (M1/M2
phenotype), changing their arteriogenic capabilities.
Purpose: Determining the effect of galectin-2 on macrophage polarization in mice.
Methods: 20 mice were allocated into two equal groups and treated twice daily with
intraperitoneal injections of either 10 mg galectin-2 (R&D Systems) or a placebo. Two days after
start of treatment, the left femoral artery was permanently ligated. One week after operation,
left adductor muscles were collected and stained immunohistochemically for smooth muscle
actin (, F4/80 (to identify macrophages), CD40 (M1-macrophages) and Mannose receptor (MR;
M2-macrophages). The ratio between perivascular CD40+ to F4/80+ or MR+ to F4/80+ cells was
calculated to quantify macrophage skewing to either the M1 phenotype (CD40+:F4/80+) or the
pro-arteriogenic M2 phenotype (MR+:F4/80+). In vitro isolated and cultured human monocytes
were incubated with IFN-γ or IL-4 to polarize them into a resp. M1 or M2 phenotype, after which
cells were incubated with Gal-2 for 24 hours. After incubation protein levels of CD40 and MR
were analysed for both subsets by flow cytometry.
Results: In vivo, systemic treatment with galectin-2 significantly increases the perivascular M1macrophage fraction (CD40+:F4/80+)(0.26±0.061 vs 0.029±0.061, p<0.01) and decreases the M2macrophage fraction (MR+:F4/80+)(0.29±0.12 vs 0.75±0.072, p<0.01). In vitro, incubation with
galectin-2 significantly reduces MR expression and increases CD40 expression (p<0.05), indicating
a transformation from a pro-arteriogenic into a phenotype
Conclusion: Galectin-2 polarizes macrophages into a M1-phenotype which is associated with
impaired arteriogenesis.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A15
Absolute thrombus density measurement on non-contrast CT in patients with acute ischemic
stroke has superior Interobserver agreement than relative ratio
Emilie M. M. Santos1,2,3,4, Albert J. Yoo, Ludo F. Beenen, Olvert A. Berkhemer3, Mark D. den
Blanken3,4, Carrie Wismans6, Wiro J. Niessen1,2,7, Charles B. Majoie3, Henk A. Marquering3,4 on
behalf of the MR CLEAN investigators
1
2
Dept. of Radiology, Erasmus MC, Rotterdam, the Netherlands; Dept. of Medical Informatics, Erasmus MC,
3
4
Rotterdam.; Dept. of Radiology, AMC, Amsterdam.; Dept. of Biomedical Engineering and Physics, AMC,
5
6
7
Amsterdam.; Texas Stroke Institute, Plano, Texas, United States; Dept. of Cardiology, AMC, Amsterdam. Faculty
of Applied Sciences, Delft University of Technology, Delft.
Background: Thrombus density is increasingly considered as predictive characteristic for acute
ischemic stroke (AIS) treatment success. However, its value is still under debate since the
published studies are not exclusive. The used thrombus density measurement method varies
widely between studies, which could explain the variations in associations with outcome success.
Aim: This study compares the observer variability of four common thrombus density manual
measurements: absolute HU and relative HU using either one or three measures.
Methods: From the MR CLEAN trial database, 104 consecutive AIS patients with thin-slice ncCT,
three expert observers and a trained observer placed three small standardized regions of interest
(ROI) in the thrombus and in the contralateral vessel. The middle ROI was used as the reference
single measure. Based on these measurements the average absolutes (aHU) and average
relatives (rHU) thrombus densities were determined aHU1, rHU1 and aHU3, rHU3 for single and
triple measure respectively. The interobserver variations were evaluated by the calculation of the
intraclass correlation coefficient (ICC) and performing Bland & Altman analysis.
Results: The results for the Bland & Altman analysis and the ICCs are showed in the table 1.
Conclusions: The interobserver variability for absolute density measurement is superior to
relative ratio. We therefore advise researchers to use relative ratio measurement with care when
studying imaging biomarker in non-contrast CT images.
Table 1 - Bland & Altman analysis and Intraclass Correlation Coefficients
Expert Observer
Trained Observer
Average Difference
Standard Deviation
Maximum Difference
Minimum Difference
Lower Limit of Agreement
Upper Limit of Agreement
aHU3
3.7
5.0
18.6
-12.5
13.7
-6.3
rHU3
-0.01
0.25
0.49
-1.15
0.49
-0.52
aHU1
4.7
8.9
40.6
-21.7
22.5
-13.1
rHU1
0.03
0.35
1.20
-1.23
0.73
-0.66
aHU3
1.3
4.6
14.6
-14.1
10.5
-8.0
rHU3
0.03
0.23
0.54
-0.80
0.49
-0.43
aHU1
1.0
7.1
20.8
-16.4
15.3
-13.2
rHU1
-0.01
0.41
1.01
-1.69
0.82
-0.83
ICC
0.71 *
0.51 *
0.49 *
0.43 *
0.82 *
0.50 *
0.70 *
0.29 †
*p-value < .001, † p-value = .002
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A16
Parasympathetic Nervous System Stimulation by Pyridostigmine Improves Survival and Cardiac
Function in Experimental Pulmonary Arterial Hypertension
Denielli da Silva Goncalves Bos, Xiaoqing Sun, Anton Vonk-Noordegraaf, Frances S. de Man, M.
Louis Handoko.
Departments of Pulmonology, Physiology and Cardiology, VU University Medical Center, Amsterdam
Introduction:
We have previously demonstrated that in pulmonary arterial hypertension (PAH) over-activation
of the sympathetic nervous system accelerates disease progression. However, less is known
about the role of the parasympathetic nervous system, its natural counterpart. Here, we
investigated the potential therapeutic effects of pyridostigmine (PYR), an oral drug that
stimulates the parasympathetic nervous system through acetylcholinesterase inhibition. We
hypothesized that PYR improved right ventricular (RV) contractility and delayed the progression
of right heart failure in experimental PAH.
Methods:
PAH was induced in male Sprague Dawley rats by a single injection of SU5416 (25 mg/kg; s.c.)
followed by 4 weeks of hypoxia (10%; SuHx). In a subgroup of rats pressure-telemetry device was
implanted. At week 6 the development of PAH was confirmed by echocardiography, and PAH-rats
were randomized for vehicle (SuHx-vehicle; n=8) or PYR-treatment (SuHx-PYR; n=12). PYR was
delivered in drinking water (40mg/kg/day). This dosage reduced heart rate by 10% and increased
heart rate variability, without significant effect on systemic blood pressure or animal activity. At
end-of-study (body mass loss >5% or at week 10), echocardiography was repeated, with
additional RV pressure-volume measurements.
Results:
PYR-treatment improved survival (Figure 1A). PYR also increased RV contractility (TAPSE and Ees;
Figure 1B-C), without a significant effect on RV afterload (Ea), resulting in a partial normalization
of RV-arterial coupling (Ees/Ea: 0.86±0.09 vs. 0.37±0.09; p<0.05). In addition, RV stiffness (Eed;
Figure 1D) and RV dilatation (RV end-diastolic diameter 6.1±0.28 vs. 7.1±0.35 mm; p<0,001) were
reduced.
Conclusion:
Parasympathetic nervous system stimulation by PYR-treatment was well-tolerated, and improved
survival and cardiac function in experimental PAH.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A17
A machine-learning based analysis of arterial pressure waveform derived parameters and
thoracic bioimpedance for the determination of central blood volume
Frank C. Bennis*1,2,3,4, Björn J.P. van der Ster*2,3,4, Wim J. Stok3,4, Berend E. Westerhof4,5,
Johannes J. van Lieshout2,3,4,6
1
2
3
University of Twente, Department of Technical Medicine, Department of Internal Medicine, Department of Anatomy,
4
Embryology and Physiology, Academic Medical Center, University of Amsterdam, Amsterdam. Laboratory for Clinical
5
Cardiovascular Physiology, Center for Heart Failure Research, Academic Medical Center, Amsterdam. Edwards Lifesciences
6
BMEYE, Amsterdam. MRC/Arthritis Research UK Centre for Musculoskeletal Ageing Research, School of Life Sciences, University
of Nottingham Medical School, Queen's Medical Centre, United Kingdom. *These authors contributed equally to the study
Introduction:
Blood loss in trauma and surgery jeopardizes tissue perfusion. Current clinical practice of
monitoring arterial pressure (BP) and heart rate (HR) do not recognize features of central
hypovolemia that precede shock since BP is insensitive for detecting ~ 1 liter volume loss [1].
Interpreting a large set of (clinical) parameters at the same time is not straightforward during the
rapid changing interaction of physiological parameters during hypovolemia. Analyzing a large set
of parameters and taking into account complex interactions is a task for machine learning
algorithms.
Methods:
In 28 healthy subjects Lower Body Negative Pressure (LBNP) of -50 mmHg provoked progressive
central hypovolemia until pre-syncope. Continuous BP and thoracic impedance were measured.
Additional parameters from the raw BP curve were extracted and served as inputs to the model
along with beat to beat values for blood pressure, stroke volume (SV), cardiac output, total
peripheral resistance, left ventricular ejection time, pulse pressure, heart rate and thoracic
impedance. All data was synchronized, resampled and inspected for artifacts. A neural network
of 10 hidden and a singular output neuron was trained on 18, validated on 9 and tested on 1
subject using the leave-one-out method. Output of the network was the time remaining till onset
of pre-syncope.
Results:
The average outcome of 50 models performed best with a mean error of 275 seconds, but with
varying results for different subjects. In 27 out of 28 subjects a downward trend was seen when
fitting a straight line, with deviations from the required offset and slope.
Conclusion:
The average outcome of 50 neural networks is able to assist in the prediction of time to presyncope by indicating a downward trend with decreasing central blood volume.
1. Secher, N.H. and J.J. Van Lieshout, Hypovolemic shock, in Clinical Fluid Therapy in the Perioperative Setting, R.G.
Hahn, Editor. 2011, Cambridge University Press. p. 166-176.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A18
Difference of Thrombus Attenuation between CT and CT Angiography Measured in Patients
with Acute Ischemic Stroke: An expression of Thrombus Porosity?
Emilie M.M. Santos†*1,2,3,4, Henk A. Marquering*1,2, Mark D. den Blanken2, Olvert A.
Berkhemer1,7, Merel A. Boers1, Albert J. Yoo5, Ludo F. Beenen1, Kilian Treurniet1, Carrie Wismans1,
Kim van Noort1, Hester F. Lingsma 6, Diederik W.J. Dippel 7, Aad van der Lugt3, Wim H. van
Zwam8, Yvo B.W.E.M. Roos9, Robert J. van Oostenbrugge10, Wiro J. Niessen3,4,11 and Charles B.
Majoie1 on behalf of the MRCLEAN investigators
1
2
Department of Radiology, Academic Medical Center, Amsterdam; Department of Biomedical Engineering and
3
4
Physics, Academic Medical Center, Amsterdam; Department of Radiology, Erasmus Medical Center, Rotterdam;
5
Department of Medical Informatics, Erasmus Medical Center, Rotterdam; Department of Radiology, Texas Stroke
6
7
Institute, Plano, Texas, United States of America; Department of Public Health, Erasmus MC, Rotterdam;
8
Department of Neurology, Erasmus Medical Center, Rotterdam; Department of Radiology, Maastricht University
9
10
MC; Department of Neurology, Academic Medical Center Amsterdam; Department of Neurology, Maastricht
11
University MC and Cardiovascular Research Institute Maastricht (CARIM), Maastricht; Faculty of Applied Sciences,
Delft University of Technology, Delft; * Both authors contributed equally.
Introduction: It has been demonstrated that a thrombus could be porous rather than fully
compact and that porosity is associated with the speed of fibrinolysis such as it may improve
intravenous treatment success in acute ischemic stroke. We hypothesize that CT contrast agent
may penetrate a porous thrombus, ensuing that the porosity could be measured by assessing
thrombus attenuation differences between ncCT and CTA imaging. We aim to characterize
thrombus attenuation changes between ncCT and CTA images as a potential imaging biomarker
of thrombus porosity.
Methods: From the MR CLEAN trial database, 125 consecutive acute ischemic stroke patients
with good quality admission thin-slice ncCT and CTA imaging were included. The ncCT and CTA
images were co-registered, and independent observers placed 3 small standardized regions of
interest in each thrombus such that the attenuation on ncCT and CTA was simultaneously
measured. CTA-based thrombus density measurement larger than 2 times the standard deviation
of the corresponding ncCT measurement was classified as porous.
Results: Thirty out of 125 thrombi (26.4%) were classified as porous with a paired average
increase of +9.8 (+/- 14.3) HU on CTA. The mean thrombus attenuation was 50.1(+/-7.4) for ncCT
and was 60.7(+/-13.4) for CTA images. Six patients (4.8%) had an increase in thrombus density of
more than 40 HU.
Conclusion: CTA-based measurements can demonstrate quantifiable increases in attenuation
value compared to ncCT assessments. This contrast penetration in CTA
likely reflects thrombus porosity, and could be investigated as a potential imaging biomarker for
treatment response to fibrinolysis.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A19
Effect of helium pre- or postconditioning on signal transduction kinases in patients undergoing
coronary artery bypass graft surgery
Kirsten F. Smit, Daniel Brevoord, Bas A. de Mol, Markus W. Hollmann; Nina C. Weber, Benedikt
Preckel
Department of Anesthesiology, Laboratory of Experimental Intensive Care and Anesthesiology (L.E.I.C.A.), Academic
Medical Centre (AMC), Meibergdreef 9, 1100 DD Amsterdam.
Introduction:
The non-anesthetic noble gas helium induces pre- and postconditioning in animals and humans.
Volatile anesthetics induce cardioprotection in humans undergoing coronary artery bypass graft
(CABG) surgery. We hypothesized that helium induces pre- and postconditioning in CABGpatients, affecting signaling molecules protein kinase C-epsilon (PKC-ε), p38 mitogen activated
protein kinase (p38 MAPK), extracellular signal-regulated kinase 1/2 (ERK-1/2) and heat shock
protein 27 (HSP-27) within cardiac tissue, and reducing postoperative troponin levels.
Methods:
After ethical approval and informed consent, 125 elective patients undergoing CABG surgery
were included and randomised in this prospective single-center study. Helium preconditioning (3
x 5 min of 70% helium and 30% oxygen) was applied just before aortic cross clamping;
postconditioning (15 min of helium) was applied before release of the aortic cross clamp.
Signaling molecules were measured in right atrial appendix specimens. Troponin-T was measured
at 4, 12, 24, and 48 hours postoperatively.
Results:
Baseline characteristics of all groups were similar. Helium preconditioning did not significantly
alter the ratio of activated p38 MAPK or ERK 1/2. Ischemia significantly increased HSP27 in all
groups except the helium-preconditioning. PKC-ε levels were unaffected. Postoperative troponinT levels showed no significant differences after helium preconditioning, postconditioning, preand postconditioning, or sevoflurane preconditioning compared to controls.
Conclusions:
No effect was observed of helium preconditioning, postconditioning or the combination of preand postconditioning on activation of p38 MAPK, ERK 1/2 or levels of HSP27 and PKC-ε in the
human heart. Helium pre- and postconditioning did not affect postoperative Troponin release in
patients undergoing CABG surgery.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A20
A role for PLVAP in the regulation of endothelial permeability and angiogenesis
Joanna Wisniewska-Kruk, Ilse M.C. Vogels, Cornelis J.F. Van Noorden, Reinier O. Schlingemann,
Ingeborg Klaassen
Ocular Angiogenesis Group, Department of Ophthalmology and Department of Cell Biology and Histology, Academic
Medical Center.
Introduction:
Plasmalemma vesicle associated protein (PLVAP) is an endothelial cell-specific structural
component of caveolae and fenestrae that is highly upregulated in angiogenic environments such
as in tumors and proliferative diabetic retinopathy.
Methods and Results:
Here we show that targeting PLVAP with small interfering RNA (siRNA) reduces physiological and
pathological angiogenesis in the retina in vivo. Furthermore, knockdown of PLVAP expression
diminished sprouting and migration of endothelial cells in vitro and decreased numbers of
endothelial tip cells, which form the leading edge of angiogenic sprouts. This reduced angiogenic
capacity of endothelial cells was associated with decreased levels of VEGFR2 and NRP2, but not
NRP1. In contrast, PLVAP overexpression resulted in high VEGFR2 levels, increased tip cell
numbers, and augmented endothelial cell sprouting.
Conclusion:
These findings indicate that PLVAP is an essential endothelial cell-specific regulatory cofactor for
VEGFR2 signaling in angiogenesis.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A21
The role of ubiquitination in regulation of endothelial barrier function
Igor Kovacevic1, Antje Schaefer1, Eloise Anthony1 and Peter Hordijk1,2
1
Dept. Molecular Cell Biology, Sanquin Research and Landsteiner Laboratory, Academic Medical Center,
2
Swammerdam Institute for Life Sciences, University of Amsterdam, Dept. Physiology, VU University Medical Center,
Amsterdam.
Introduction:
Ubiquitination is well studied in the context of TNF signaling and the consequent inflammatory
response in endothelial cells. However, the function of ubiquitination in the control of of
endothelial barrier function by TNF or other receptor agonists has not been not extensively
studied .
Methods:
Based on published and our preliminary data we have selected a subset of ubiquitin E3 ligases,
deubiquitinases and adaptor proteins for a targeted shRNA screen in primary human endothelial
cells. To identify candidate regulatory proteins, we analysed cytoskeletal morphology and
formation of adherens junctions using confocal immunofluorescence microscopy in combination
with real-time quantification of transendothelial electrical resistance.
Results and conclusions:
At the molecular level, we focus on ubiquitination-dependent protein traffic and degradation of
small Rho GTPases, key regulators of endothelial permeabilty and cell-cell contacts. In a
subsequent set of experiments we have tested the impact of the loss of identified candidates on
the interaction of endothelial cells with immune cells and with the extracellular matrix. By
applying a targeted knockdown screen in primary human endothelial cells we identified several
components of the ubiquitination/deubiquitination machinery that are responsible for the
maintenance of endothelial integrity and that control the transendothelial migration of immune
cells. Data obtained from the screen will be presented at the meeting.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A22
The tyrosine kinase Arg/Abl2 mediates endothelial barrier disruption by impairing integrin
α5β1-mediated cell adhesion
Jurjan Aman1; Erik Valent1; Joana M. Azevedo1; E. Eringa1; Jan van Bezu1; Ramaswamy Krishnan2;
Victor W.M. van Hinsbergh1; Geerten P. van Nieuw Amerongen1.
1
2
Department of Physiology, Institute for Cardiovascular Research, VU University Medical Center, Amsterdam, Center
for Vascular Biology Research, Beth Israel Deaconess Medical Center, Boston, Massachusetts, United States of
America.
Introduction:
Endothelial barrier disruption and vascular leakage contribute to organ dysfunction during
inflammation. We recently identified the kinase Abl-related gene (ARG/Abl2) as mediator of
endothelial barrier disruption, but the mechanisms by which Arg mediates endothelial barrier
disruption remain largely unknown. As Arg forms a potential target for treatment of vascular
leak, we investigated the role of Arg in endothelial barrier regulation in vitro and in vivo.
Methods & Results:
Here, we report that Arg is dispensable for barrier function in the resting endothelium, but is a
key mediator of inflammation-induced endothelial barrier disruption in vitro and in vivo. While
showing a normal inflammatory response, Arg-/- mice are protected against pulmonary vascular
leak upon exposure to intratracheal lipopolysaccharide. We show that Arg/Abl2 is activated in
the endothelium (phosphorylation of the Arg/Abl2 downstream target CrkL) during inflammation,
both in cultured endothelial cells and in lung tissue of critically ill patients. Arg is involved in
internalization and spatial distribution of α5β1 integrin, as Arg-depleted endothelial cells show
enhanced expression of α5β1 integrin which is located predominantly at the cell periphery, close
to cell-cell contacts. In parallel, Arg/Abl2 attenuates Rap1 activation upon cell-cell junction
disruption. The α5β1 redistribution and the enhanced Rap1 activation are associated with reduced
force development at the cell-matrix interface and reduced cell retraction, thus limiting
intercellular gap formation.
Conclusion:
This study reveals a novel Arg-driven mechanism of endothelial barrier disruption, in which Argmediated breakdown of peripheral cell matrix interactions allows for cell retraction and subsequent gap formation and cell detachment.
Acknowledgements:
This work was funded by the Netherlands Heart Foundation (grants #2003T3201 and #2014T064 (JA) and #2003T032
(GPvNA)).
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A23
High-Resolution 3D Imaging of Vessels in the Ischemic Rat Hind Limb for Collateral
Quantification
Janina C.V. Schwarz, Monique G.J.T.B. van Lier, Elco Oost, Erik N.T.P. Bakker, Jos A.E. Spaan, and
Maria Siebes
Department of Biomedical Engineering and Physics, Academic Medical Center
Introduction:
Three-dimensional quantification of vascular architecture is essential for validation of molecular
imaging biomarkers for neovascularization. Fluorescent markers can be quantified in 3D using an
imaging cryomicrotome.
Methods:
Neovascularization was stimulated by femoral artery ligation. After sacrifice, the vasculature was
flushed and filled with fluorescent cast material. In 27 healthy legs, the casting procedure was
optimized for penetration into small vessels. Eighteen ischemic rats were filled according to the
optimized protocol, embedded in black gel and frozen. Registered 3D reflectance and
fluorescence image stacks (24 µm isotropic voxel resolution) were obtained by alternate slicing
and episcopic imaging with an automated imaging cryomicrotome. Subsurface fluorescence was
corrected with a system-specific point-spread-function. Diameters of the gracilis collaterals were
estimated by a semi-automated technique based on vesselness filtering.
Results:
A mixture of Batson’s with methylmethacrylate (ratio 7:3), infused at physiological pressure, was
optimal for filling arterioles and collateral vessels. Initial clamping of the healthy iliac artery
resolved unbalanced filling of healthy and ischemic side. Median diameter of the collateral
arteries increased from 110 µm in healthy legs to 179 µm 28 days post-ligation.
Conclusions:
The proposed method provides a powerful basis for 3D quantitative analysis of collateral
adaptation in the ischemic rat hind limb, together with visualization of perfusion and coregistered fluorescent biomarkers.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A24
Cerebral perfusion and white matter hyperintensities in hypertension older people
Jan Willem van Dalen, Henri J.M.M. Mutsaerts, Aart J. Nederveen, Hugo Vrenken, Martijn D.
Steenwijk, Matthan W.A. Caan, Charles B.L.M. Majoie, Willem A. van Gool, Edo Richard.
Department of neurology, Academic medical Center, Amsterdam.
Introduction:
This study aims to assess whether white matter hyperintensities (WMH) of presumed vascular
origin in elderly with hypertension are associated with reduced perfusion not only in the WMH,
but also in the surrounding normal appearing white matter (NAWM) and grey matter (GM).
Methods:
We studied 185 participants of the Prevention of Dementia by Intensive Vascular care (preDIVA)
trial, with systolic hypertension, aged between 72 and 80 years old. WMH volume and cerebral
blood flow (CBF) were derived from 3D FLAIR and Arterial Spin Labelling (ASL) MRI respectively.
We compared WMH CBF, NAWM CBF and GM CBF across quartiles of WMH volume and assessed
the continuous relation between these CBF estimates and WMH volume using linear regression.
Results:
From the lowest two quartiles of WMH volume upward, with each quartile increase in WMH
volume mean WMH CBF markedly declined. There was a negative association between WMH
volume and WMH CBF (beta=-0.248, p=0.001). There was no difference in NAWM or GM CBF
across quartiles of WMH volume nor a relation between WMH volume and NAWM CBF (beta=0.065, p=0.643) or GM CBF (beta=-.035, p=0.382).
Conclusion:
WMH in elderly with hypertension are associated with lower perfusion in WMH, but not in the
surrounding NAWM and GM. This suggests that WMH in these patients relate to local
microvascular alterations rather than a general cerebral perfusion deficit.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A25
Resveratrol Inhibits Aortic Root Dilatation in the Fbn1C1039G/+ Marfan Mouse Model
Stijntje Hibender*1, Romy Franken*2,3, Inge van der Made4, Edith E Schermer1, E Lutgens1,5, Yigal
M Pinto4, Maarten Groenink2,3,6, Barbara JM Mulder2,3, Aeilko H Zwinderman7, Carlie JM de
Vries1, Vivian de Waard1
* both authors contributed equally
1 Department of Medical Biochemistry, Academic Medical Center Amsterdam,
2 Department of Cardiology, Academic Medical Center Amsterdam,
3 Interuniversity Cardiology Institute of The Netherlands
4 Department of Experimental Cardiology, Academic Medical Center, Amsterdam
5 Institute for Cardiovascular Prevention (IPEK), Ludwig Maximilians University,
Munich, Germany
6 Department of Radiology, Academic Medical Center Amsterdam.
7 Department of Clinical Epidemiology and Biostatistics, Academic Medical Center
Amsterdam.
Introduction:
Marfan syndrome (MFS) is a connective tissue disorder
caused by mutations in the fibrillin-1 gene (FBN1), resulting in aortic aneurysm formation and
dissections. Currently, β-blockers and losartan are used, however, more treatment possibilities
are required. Since spontaneous aneurysm formation occurred in smooth muscle cell-specific
sirtuin-1 (SIRT1) deficient mice, SIRT1 is considered a potential target. SIRT1 is an NAD+dependent deacetylase and prevents senescence. Resveratrol induces SIRT1 activation and
reduces senescence, which prompted us to investigate its effectiveness in the Fbn1C1039G/+
MFS mouse model.
Results:
Senescence was indeed observed in the ascending aorta of MFS mice and correlated with the
aortic root dilatation rate. Resveratrol inhibited aortic dilatation more efficient than losartan. It
did not affect activation of SMAD2 and ERK1/2, downstream pathways of the angiotensin-II
receptor 1 (AT1R), which is blocked by losartan. The aortic root of resveratrol-treated mice
showed increased nuclear SIRT1, a decrease in medial area, and more healed elastin fibres.
Expression of mRNA in aortic tissue from MFS mice treated with resveratrol revealed increased
microRNA-21a and decreased microRNA-29b, resulting in upregulation of marker of proliferation
Ki-67 (Mki67) and pro-survival gene B-cell CLL/lymphoma 2 (Bcl-2).
Conclusion:
Resveratrol inhibits aortic root dilatation in MFS mice via a different mechanism of action than is
considered for losartan, by promoting elastin fibre healing and cell survival. A switch in the
detrimental aneurysm microRNA pattern is proposed to be the underlying mechanism.
Therefore, resveratrol may hold promise as a therapeutic for MFS patients.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A26
Establishment of viable ex vivo human and calf aortic tissues as basis for research on the
pathophysiology of aortic aneurysms.
Jorn P. Meekel, Erik N. van Tol, Menno E. Groeneveld, René J.P. Musters, Behrouz Z. Doulabi,
Dimitra Micha, Gerard Pals, Arjan W.J. Hoksbergen, Willem Wisselink, Jan D. Blankensteijn, Kak K.
Yeung.
Institute for Cardiovascular Research (ICaR-VU), VU University medical center, Amsterdam.
Background and aim of the study:
Abdominal aortic aneurysms (AAA) are a common health problem, however their
pathophysiology remains still unclear. Thus far, fixated cells or isolated cell cultures have been
used for research on AAA. However, complex organ function as in aorta is based on cell-to-cell
interactions, cell receptors and communication with surrounding tissue, all of which will vanish in
preceding stated techniques. Contrary to isolated or fixated cells, we suggest that slice
preparations of aortic tissue maintain anatomical, functional and microstructural characteristics.
Aim of the study is to develop an ex vivo method to thinly slice aorta and iliac artery tissue, to
maintain viability of sliced tissue and quantify aforementioned viability.
Methods:
Adult human aorta or iliac artery tissue was obtained during aneurysm surgery (n = 7) and aorta
of calf was obtained from an abattoir (n = 3). After ice-cold transport, agar-embedded cubes of
tissue were cut into 150-μm-thick slices with a vibratome (Leica VT1200 S, Leica Biosystems,
Nussloch, Germany). Slices contained media, adventitia and conceivably intima and were
conserved in cell proliferation stimulating media at 37° Celsius until fourteen days after retrieval.
Viability was quantified using LIVE/DEAD® Viability/Cytotoxicity Kit (Life Technologies, Carlsbad,
CA, United States). Furthermore, immunohistochemical analysis of several molecules including αSMA, smoothelin, CD45, rhodamine-labeled-wheat-germ-agglutinin and dapi was performed on
the tissue samples.
Results:
We showed that about 50% of the cells in the aorta and iliac artery lived more than ten days after
harvesting and cut with the vibratome. Day three showed a drop of cells alive, but subsequently
day five and day seven showed peaks of living cells; over ten per cent increase of viable cells
compared to day zero. While dead cells were located mostly on the surface of the tissue, living
cells could only be found in the central of vascular tissue slices. Staining showed that mostly
smooth muscle cells stayed alive during staining.
Discussion and conclusion:
Live aortic or iliac vessel cells in tissue slices can be preserved and can proliferate ex vivo over ten
days. This study provides a new solid basis for experimental pharmacological and provocation
studies based on living aortic or iliac vessel cells. One of our future studies involves studying the
effect of TGF-β on the SMAD regulation in these tissues.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A27
HDAC inhibition improves atherogenic macrophage function and affects cellular metabolism
Jan Van den Bossche1, Annette E. Neele1, Marten A. Hoeksema1, Jeroen Baardman1, Menno P.J.
de Winther1
1
Department of Medical Biochemistry, Experimental Vascular Biology, Academic Medical Center, Amsterdam.
Introduction:
While macrophages play a well-established role in the pathogenesis of atherosclerosis, the
pathways that regulate their atherogenic functions remain ill-defined. Chromatin remodeling by
epigenetic enzymes governs immune responses and specific metabolic pathways are being
recognized as critical hallmarks of macrophage subsets. Yet, it remains unclear to what extent
chromatin modifying enzymes (CME) and changes in metabolism control atherogenic
macrophage functions.
Hypothesis and methods:
We hypothesize that the plaque micromilieu, through epigenetic and metabolic changes, dictates
macrophage activation and polarization, thereby determining the outcome of atherosclerosis.
Therefore, we aim to study whether the inhibition of CME can be applied to modulate cellular
metabolism and atherogenic macrophage activities as a way to improve atherosclerosis outcome.
Results:
We found that broad-spectrum inhibition of histone deacetylases (HDAC) and histone
methyltransferases (HMT) has both pro- and anti-inflammatory effects. The inhibition of HDACs
decreased macrophage apoptosis and increased histone acetylation and gene expression of the
cholesterol efflux regulators ABCA1 and ABCG1.
To assess whether the pro- and anti-inflammatory effects of CME-inhibitors are associated with
metabolic changes, we performed metabolic flux analysis to determine the extra-cellular
acidification rate (ECAR; as a measure of glycolysis) and the oxygen consumption rate (OCR; as a
measure of oxidative phosphorylation [OXPHOS]). While HATi, HDMi and HMTi had no
noteworthy effects on the cellular metabolism in macrophages, HDAC inhibition resulted in
remarkable alterations in cellular metabolism in M1, but not in naïve and M2 macrophages.
Confirming previous studies, we found that classical M1 macrophage activation by IFN-gamma
plus LPS induces glycolysis and impairs OXPHOS. The HDACi strongly induced ECAR and prevented
the IFN-gamma/LPS-induced drop in OCR which is normally observed during classical
macrophage activation. Hence, HDACi-treated M1 macrophages have a reduced inflammatory
response and display a metabolic shift towards both high glycolysis and OXPHOS.
Conclusion:
Based on our data, we propose the inhibition of specific HDAC enzymes in macrophages as a
novel potential target to improve their metabolism and atherogenic functions.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A28
LVAD-support Induced Hemodynamic Changes Result in Valve Interstitial Cell Activation and a
Reparative Response in the Aortic Valve
Jan Willem van Rijswijk1, Aryan Vink2, Jerson R. Martina1, Roel A. de Weger2, Faiz Z. Ramjankhan1,
Nicolaas de Jonge3, Jolanda Kluin1.
1
2
Dept of Cardiothoracic Surgery, University Medical Center Utrecht, Utrecht. Dept of Pathology, University Medical
3
Center Utrecht, Utrecht. Dept of Cardiology, University Medical Center Utrecht, Utrecht.
Introduction:
Left ventricular assist device (LVAD)-support and subsequent explantation of the heart during
heart transplantation provide a unique human model to study the effect of altered flow dynamics
on an otherwise normal aortic valve. To date, no data exists on the effect of LVAD-support on
aortic valvular interstitial cell (VIC) phenotype and microenvironment. The aim of this study was
to characterize the response of the aortic valve leaflet and its embedded VICs to LVAD-induced
hemodynamic changes.
METHODS: Aortic valves of 21 continuous flow-LVAD-patients were examined after heart
transplantation or autopsy using (immuno)histochemistry.
Results:
Significant commissural fusion was observed in 15 out of 21 (71%) LVAD aortic valves. Cell
density and -proliferation were markedly increased in the ventricularis layer after LVAD support,
coinciding with a localized increase in myofibroblast-like cells, which positively correlated to the
duration of LVAD-support. CD68 positive macrophages were increased in all valve layers in the
LVAD group, which especially in the ventricularis layer appeared to be of M2 type (CD163
positive). M1 macrophages (iNOS positive) were mostly observed in sclerotic lesions that did not
have an association with LVAD support.
Conclusions:
LVAD support induces increased numbers of macrophages of the M2 subtype and smooth muscle
like cells in the aortic valve. The presence of M2 macrophages points to a reparative response
rather than a pro-inflammatory one. In addition our results suggest phenotypic adaptation of
VICs to a more contractile phenotype as a result of LVAD-induced changes in hemodynamic
forces. Our results show that the human aortic valve is a dynamic part of the circulation that is
able to adapt to a new hemodynamic situation.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A29
EVENING INTAKE OF ASPIRIN IS ASSOCIATED WITH A MORE STABLE 24 HOUR PLATELET
INHIBITION COMPARED TO MORNING INTAKE: A STUDY IN CHRONIC ASPIRIN USERS
Jeske Joanna Katarina van Diemen¹*, Drs. Wessel Willem Fuijkschot¹*, Drs. Tim Jon Wessels¹, Dr.
Gerrit Veen², Prof. Dr. Yvo Michiel Smulders¹, Dr. Abel Thijs¹
¹ Internal medicine, VU university medical center, Amsterdam ; ² Cardiology, VU university medical center,
Amsterdam * These two authors contributed equally to this work
Background: Daily generation of novel platelets may compromise aspirin’s platelet inhibitory
action, especially near the end of the regular 24-hour dosing interval. A contributor to this
attenuation could be the endogenous circadian rhythm.
Methods: The primary objective of this study was to assess platelet activity 12 and 24 hours after
different times of aspirin intake (c.q. 8.00 AM and 8.00 PM). A randomized open label cross-over
study was conducted, comprising outpatients with stable cardiovascular disease taking aspirin
once daily. We measured platelet aggregation with the Platelet Function Analyzer (PFA)-200® and
light transmission aggregometry (LTA).
Results: The attenuation of aspirin’s inhibitory action was most apparent in the 8.00 AM
regimen. The PFA-CT was 78 seconds faster at 24 hours than at 12 hours after intake in the 8.00
AM regimen (IQR: 166,8 - 301 vs. 132,8 – 301; p= 0,006) and 0 seconds faster at 24 hours than at
12 hours after intake in the 8.00 PM regimen (IQR: 198,8 - 837,0 vs. 169,8 – 301; p= 0,653). The
ADP 1,0 µmol/L maximum amplitude was 5,40% higher at 24 hours than at 12 hours after intake
in the 8.00 AM regimen (95% CI: -0,03 - -10,8; p= 0,040) and was 0,75% higher 24 hours than at
12 hours after intake in the 8.00 PM regimen (95% CI: -4,83 – 3,33; p=0,705).
Conclusion: The platelet inhibitory effect of aspirin decreases after 24 hours, particularly after
intake in the morning. These results suggest that patients might benefit from evening intake or
twice daily intake regimens.
Figure 1. PFA-200 ClosureTime 12 versus 24 hours after intake.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A30
The impact of family history on cardiovascular disease is higher among hypertensive individuals
of African and Surinamese background
L. Valerio (1), S.J. Pinto (2), K. Zwinderman (2), R. Peters (3)
(1)Department of Public Health, Academic Medical Centre, Amsterdam (2)Department of Clinical Epidemiology and
Biostatistics, Academic Medical Centre, Amsterdam (3)Department of Cardiology, AMC, Amsterdam.
Introduction: Hypertension is an important risk factor for cardiovascular disease (CVD), but the
overall baseline CVD risk for subjects with hypertension is low: individual risk varies greatly
among them. Also, hypertension-related CVD varies sharply across ethnicities. We hypothesized
that a positive family history of CVD, possibly reflecting predisposition to damage by
hypertension, independently identifies high-risk subpopulations of hypertensive subjects, and
that its impact on CVD differs across ethnic backgrounds.
Methods: We cross-sectionally investigated the impact of family history on CVD in the
hypertensive subjects (N=7476) of the HELIUS cohort in Amsterdam, including participants of
native Dutch, South-Asian Surinamese, African Surinamese, Ghanaians, Turks, and Moroccan
backgrounds. We defined positive family history of CVD as a reported vascular event in a firstdegree relative before age 60; and CVD as past diagnosis or symptoms consistent with coronary
heart disease or cerebrovascular disease before age 60. Analyses were adjusted for age, gender,
smoking status, education level, waist circumference, diabetes, and dyslipidemia.
Results: Family history of CVD ranged from 10.3% among Ghanaians to 50% among South-Asian
Surinamese, and yielded the highest independent risk for CVD among hypertensive subjects with
South-Asian Surinamese (Odds Ratio [95% CI]: 2.35 [1.79-3.10]), Ghanaian (2.20 [1.48-3.22]) and
African Surinamese background (2.01 [1.53-2.64]), p<0.005. No independent effect on risk was
found in native Dutch and Moroccan backgrounds (Figure 1).
Discussion. Family history of CVD is a clinically significant discriminant for CVD among
hypertensive subjects in patients of African and Surinamese origin. Future studies should clarify
whether genetic predispositions explain these interethnic differences.
Association between
family history of CVD and preval
Ethnicity
Dutch (N=1063)
S.Asian Surinamese (N=1276)
Afr. Surinamese (N=1775)
Ghanaians (N=1501)
Turks (N=1080)
Moroccans (N=781)
1
2
Adjusted OR
3
Figure 1 – Association between positive family history of CVD and prevalent CVD under age 60 in subjects with
hypertension, stratified by ethnic background. A31
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A31
Nlrp3 plays no role in acute cardiac infarction due to low cardiac expression
Willeke M.C. Jong1,2, Jaklien C. Leemans4, Nina C. Weber1,2, Nicole P. Juffermans1,3, Marcus J.
Schultz1,3, Markus W. Hollmann1,2, Coert J. Zuurbier1,2
1
2
Laboratory of Experimental Intensive Care and Anesthesiology (LEICA), Department of Anesthesiology,
4
Department of Intensive Care, Department of Pathology, Academic Medical Center, Amsterdam.
3
Introduction:
Myocardial ischemia-reperfusion (I/R) and ischemic preconditioning (IPC) are associated with
activation of inflammatory pathways that contribute both to injury and protection. Here we
examine to which extent deletion of Nlrp3 (NOD-like receptor) contributes to acute cardiac
infarction, inflammation and STAT3 signalling under I/R and IPC.
Methods:
Ischemia of 30 min and reperfusion of 3 h by LAD occlusion were compared between wild-type
(WT) and Nlrp3 / mice in a closed-chest model.
Results:
Nlrp3 deletion did not affect infarct size after I/R, and did not alter 4x5min IPC-induced
cardioprotection. Cardiac levels of TNFα were reduced in Nlrp3 / mice following I/R; IL 1β and IL
6 levels were unchanged. Nlrp3 deletion was associated with reduced cardiac STAT3
phosphorylation following I/R and IPC+I/R. Inflammation was largely attenuated in the closedchest I/R model, as compared to commonly used open-chest models. The lower inflammatory
response in the closed-chest model was associated with no detectable cardiac NLRP3 expression,
whereas cardiac NLRP3 was readily detectable in the open-chest model.
Conclusion:
The innate immune NLRP3 receptor is necessary for cardiac STAT3 phosphorylation but plays no
role in acute infarction after I/R or IPC in a closed-chest model.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A32
Variability in IV Adenosine-Induced Coronary Microvascular Resistance and Systemic Pressure
Compromises FFR Determination despite Stable Stenosis Hemodynamics
Lorena Casadonte1, Koen M. Marques2, Jos A.E. Spaan1, Maria Siebes1
1
2
Department of Biomedical Engineering and Physics, Academic Medical Center, Amsterdam. Department of
Cardiology, VU University Medical Center, Amsterdam.
Introduction:
Intracoronary (IC) and intravenous (IV) adenosine are used to induce hyperemia for functional
stenosis severity assessment. Our aim was to investigate the effect of IC and IV adenosine
administration on variability in coronary and stenosis hemodynamics.
Methods:
In 12 diseased coronary vessels (25-56% DS), aortic pressure (Pa), distal pressure (Pd) and flow
velocity (v) were simultaneously measured at baseline and hyperemia induced by IC (40 μg bolus)
and IV adenosine administration (140 μg/kg/min). At baseline and throughout the hyperemic
response we obtained cycle-averaged pressure and velocity signals to derive microvascular
resistance (MR = Pd/v) and FFR= Pd/Pa. From fitted stenosis pressure gradient-velocity (ΔP-v)
relations, we determined ΔP at v= 30 cm/s (dPv30).
Results:
Maximal v was similar for IC and IV adenosine (50±4 vs. 54±5cm/s) with no difference in MR
(1.73±0.10 vs. 1.74±0.13 mmHg/cm/s). However, MR during IV hyperemia rose between 9 and
135% in the post-peak period, with concomitant fluctuations in v (13 to 61% decline) and
pressures (up to 34% decline). Minimal FFR coincided with peak v for IC adenosine, but occurred
21±8 s after peak v for IV adenosine, when Pa had decreased by 13% and HR increased by 25% vs.
baseline. IV and IC derived stenosis ΔP-v relations followed a stable common quadratic
relationship, with no difference in dPv30.
Conclusion:
IV adenosine caused persistent systemic and coronary hemodynamic variability. IC adenosine
yielded comparable hyperemia with minor effects on systemic hemodynamics. Consistent ΔP-v
relations and dPv30 provide reliable means for stenosis evaluation free from maximal
vasodilation.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A33
Increased native T1-values at the interventricular insertion regions of precapillary pulmonary
hypertension patients
Spruijt OA1, Bogaard HJ1, Visser L1; Hofman MBM1, Vonk-Noordegraaf A1, Marcus JT2
1
2
Department of Pulmonary Medicine, Department of Physics and Medical Technology,
VU University Medical Center, PO Box 7057, 1007MB, Amsterdam.
Background:
Due to pressure overload of the right ventricle (RV) in precapillary pulmonary hypertension (PH),
the interventricular insertion regions showed Late Gadolinium Enhancement (LGE), representing
fibrosis. Our aim was to characterize the interventricular insertion regions in precapillary PH
patients using native T1-mapping.
Methods:
70 precapillary PH patients (mean Pulmonary Arterial Pressure = 47±13mmHg) were included.
Native T1-mapping was acquired on a Siemens 1.5 T Avanto MRI scanner. A modified Look-Locker
inversion-recovery sequence was used on a mid-ventricular short axis plane. Three, three, and
five non-segmented images were acquired at end-diastole to sample the recovery of longitudinal
magnetization after the inversion pulse. Native T1-values were assessed using regions of interest
at the interventricular insertions, septum, RV free wall and LV free wall.
Results:
Native T1-values at the interventricular insertion regions were higher compared to the RV free
wall, LV free wall, and septum (p<0.001). Native T1-values at the insertion regions were related
to right atrial pressure (pearson r=0.310; p=0.01), RV end-diastolic volume (pearson r=0.376;
p=0.001), RV ejection fraction (pearson r=-0.282; p=0.018) and NT pro-BNP (pearson r=0.392;
p=0.001).
Conclusion:
Native T1-values at the interventricular insertion regions are increased in precapillary PH and are
related to disease severity. This finding is in line with previous PH studies using LGE.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A 34
MACROPHAGE LIPID LOADING LEADS TO TYPE I INTERFERON HYPORESPONSIVENESS
Marieke C.S. Boshuizen, Marten A. Hoeksema, Annette E. Neele, Saskia van der Velden, Jan Van
den Bossche, Menno P.J. de Winther
Department of Medical Biochemistry, Academic Medical Center, University of Amsterdam, Amsterdam.
Introduction:
Macrophage-derived foam cells are critical components of atherosclerotic lesions and the ways in
which their inflammatory response influences atherogenesis is of great interest. Previously we
demonstrated that interferon-beta (IFNβ) promotes atherogenesis. But how IFNβ influences
foam cell inflammatory responses is still understood. IPA analysis indicated a downregulated
interferon signaling upon monocyte and macrophage lipid loading. Hence we wanted to assess
whether macrophage lipid loading also results in a decreased interferon response.
Methods:
We loaded human primary macrophages and bone marrow-derived macrophages overnight with
acLDL followed by 6h IFNβ treatment. For in vivo validation, LDLR-/- mice were put on normal
chow (NC) or a high cholesterol diet (HCD) for 10 weeks. Peritoneal macrophages (PEMs) were
collected 4 days after intraperitoneal thioglycollate administration, combined with IFNβ, IFNγ or
PBS administration 24 and 8h before sacrifice.
Results and Conclusion:
Both in vitro and diet-induced lipid loading impaired the induction of IFNβ target genes like CCL5
and CXCL10. In addition, ex vivo culturing of PEMs from IFNβ-treated animals on HCD versus NC
showed a decreased inflammatory activity. We observed this hyporesponsiveness to be specific
for IFNβ, as these effects were not observed following IFNγ stimulations. Decreased IFNβ-induced
activation of STAT2 may be the underlying mechanism of this hypercholesterolemia-induced
hyporesponsive state. Currently we assess whether this also affects the IFNβ-induced antiviral
response. Altogether, we observed that macrophage lipid loading results in hyporesponsiveness
to IFNβ. Future research will clarify whether this hyporesponsiveness also affects IFNβ-mediated
antiviral activity, which might have implications for obesity related disorders.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A35-1 (first session)
Early specification and progressive restriction of the cardiac conduction system lineage during
heart development
Rajiv Mohan, Mathilda Mommersteeg, Vincent Wakker, Arie Verkerk, Vincent Christoffels
Department of Anatomy, Embryology & Physiology, Academic Medical Center, Amsterdam.
Introduction:The adult cardiac conduction system (CCS), comprising the sinus node,
atrioventricular node and junction, the atrioventricular bundle and bundle branches, is composed
of specialized cardiomyocytes that orchestrate the rhythm of the heart. In the adult heart, they
are the only cells with a pacemaker phenotype, capable to generate spontaneous action
potentials, a feature that is absent in the working myocardium (WM) of the chambers. It is
unclear from which progenitors the adult CCS components originate during embryogenesis and
precisely when the CCS lineage becomes specified. The adult CCS components specifically express
T-box transcription factor Tbx3, which is expressed early in heart development and is required for
the correct development and functioning of the CCS. We hypothesize that 1) the Tbx3+
cardiomyocytes in the embryo function as a CCS framework and 2) they are the progenitors of
the adult CCS.
Methods & Results: To address these issues, transgenic mouse lines have been generated
expressing either fluorescent protein Venus or tamoxifen-inducible Cre under control of the Tbx3
locus. Expression and electrophysiological analyses of the Venus-positive cardiomyocytes
revealed that Tbx3 expression specifically marks cardiomyocytes with a pacemaker phenotype
throughout development from as early as embryonic day 10.5, briefly after the initiation of
chamber differentiation. To investigate the origin of the mature CCS, Tbx3+ cells were pulse
labeled at subsequent stages of development using the tamoxifen-inducible Cre system, and the
fate of their progeny was assessed in the late fetal heart when the CSS components have been
well formed. Interestingly, as early as embryonic day 7.5-8.5, just after gastrulation, Tbx3+ cells
contribute to all components of the CCS except for the sinus node. One day later, the heart still a
tubular structure, the Tbx3+ population contributes to all CCS components. The embryonic Tbx3+
population also contributed to WM cardiomyocytes in the chambers. Both the contribution and
the distance from the CCS to the labeled WM cells progressively decreased with labeling at later
stages of embryonic development.
Conclusions:We conclude that 1) the Tbx3+ cardiomyocytes in the early embryonic heart
function as the embryonic CCS, 2) the mature CCS components originate from the Tbx3+
cardiomyocytes in the early developing heart, and 3) Tbx3+ cardiomyocytes initially are bipotent,
giving rise to CCS and WM cardiomyocytes, but become progressively restricted to the CCS
lineage.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A35-2 (second session)
In-situ tissue engineering of vascular grafts
Investigate neo-tissue formation and degradation velocity of bisurea polymeric vascular grafts
Funded by CVON and the Dutch heart foundation.
M. Uiterwijk, J. Kluin (AMC), H.M. Janssen, S.H.M. Söntjens (SyMO-Chem), P.Y.W Dankers, C.V.C
Bouten (TU/e)
Introduction:
Tissue engineering can meet the increasing demand of patients who need cardiovascular
substitutes. In the laboratory (in vitro) or in the body (in-situ) neo-tissue can be regenerated on a
mold (scaffold). These scaffolds are made of polymeric biocompatible material that degrades in
time. Meanwhile neo-tissue grows on the scaffold by recruited cells. The balance between the
formation of neo-tissue and degradation of the polymer is crucial to maintain a stable construct.
We will perform an animal in-situ study to investigate neo-vascular tissue formation and polymer
degradation in order to compare three different supramolecular bisurea based polymeric
vascular grafts.
Methods:
The abdominal aorta will be will be replaced by an electrospun vascular graft in Swifter rats
(n=60). Three groups, based on polymeric subtype will be formed with follow up time of 2 weeks
and 1 month. All grafts are shielded with Gore-Tex to prevent transmural and transanastomotic
ingrowth, resulting in tissue formation caused by circulating cells only. At finish of follow up
time, immunohistochemical and biomechanical assays of the explanted grafts will be performed.
Degradation velocity is calculated on molecular weights of the polymers, measured by gel
permeation chromatography (GPC) before and after removal of residual tissue. Based on the first
short-time results, 3 new follow up times will be set, in a time range of 1 and 18 months.
Study start January 2016
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A36
Automatic Aortic Root Landmarks Detection in CTA images for Transcatheter Aortic Valve
Implantation Planning
Mustafa Elattar1, Esther Wiegerinck2, Floortje van Kesteren2,3, Lucile Dubois4, Nils Planken3, Ed
Vanbavel1, Jan Baan2,and Henk Marquering1,3
1
2
Biomedical Engineering and Physics, Academic Medical Center, University of Amsterdam; Heartcenter, Academic
3
Medical Center, University of Amsterdam; Radiology, Academic Medical Center, University of Amsterdam;
4
Biomedical Engineering, Polytech Lyon, Université Claude Bernard Lyon.
Introduction:
Transcatheter aortic valve implantation is currently a well-established minimal invasive
treatment option for patients with severe aortic valve stenosis. CT Angiography is used for the
pre-operative planning and sizing of the prosthesis. To reduce the inconsistency in sizing due to
interobserver variability, we introduce and evaluate an automatic aortic root landmarks
detection method to determine the sizing parameters.
Methods:
The automated sizing approach detects the two coronary ostia and three valvular hinge points on
a segmented aortic root surface. Using these aortic root landmarks, the automated method
determines annulus radius, annulus orientation, and distance from annulus plane to right and left
coronary ostia. Validation is performed by the comparison with manual measurements of two
observers for 20 CTA image datasets of patients eligible for TAVI.
Results:
Detection of landmarks showed high accuracy where the mean distance between the
automatically detected and reference landmarks was 2.5 ± 1.4 mm, comparable to the
interobserver variation of 2.3 ± 1.3 mm. The mean annulus to coronary ostium distance was 16.7
± 2.9, and 17.1 ± 2.8 mm for the automated and the reference manual measurements,
respectively, with a mean paired difference of 1.6 ± 1.5 mm and interobserver mean paired
difference of 0.96 ± 0.63 mm.
Conclusions:
Automated detection of aortic root landmarks enables automated sizing with good agreement
with manual measurements, which suggests applicability of the presented method in current
clinical practice.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A37
CD34 marks angiogenic tip cells in human vascular endothelial cell cultures
Marchien G. Dallinga1, Martin J.Siemerink1, Ilse M. C. Vogels1, Ingeborg Klaassen1, Arjen W.
Griffioen2, Pieter Koolwijk3, Cornelis J. F. van Noorden1, Reinier O. Schlingemann1
1
Ocular Angiogenesis Group, Department of Ophthalmology and Department of Cell Biology and Histology,
2
Academic Medical Center. Angiogenesis Laboratory, Department of Medical Oncology, VU Medical Center,
3
Department of Physiology, VU Medical Center.
Introduction:
The growth of new blood vessels from the pre-existing vascular network, called angiogenesis, can
be harmful in many diseases ranging from age-related macular degeneration in the eye to tumor
angiogenesis that promotes growth and metastases. Angiogenic research has had a boost since
the discovery that specialized endothelial cells take on a differentiated phenotype, called tip cell,
that enables them to lead the growing sprout. Targeting the tip cell phenotype for therapy would
specifically target angiogenic blood vessels and leave the quiescent network unaffected. In
cultures of endothelial cells, there is a subpopulation of cells that expresses the sialomucin CD34.
We hypothesized that these CD34+ cells are the in vitro representatives of tip cells.
Methods and Results:
Using fluorescently activated cell sorting (FACS) to separate CD34+ cells from CD34- cells, we were
able to characterize the CD34+ subpopulation in human umbilical vein endothelial cells (HUVECs),
human microvascular endothelial cells (hMVECs) and a dermal microvascular cell line (HMEC-1).
We found that, similar to tip cells in vivo, CD34+ cells proliferate less than CD34- cells, extend
filopodia, show increased mRNA expression of genes known to be expressed in tip cells in vivo
and that more CD34+ arise in the cultures upon stimulation with vascular endothelial growth
factor (VEGF). Furthermore, it seems that the CD34+ cells are necessary for in vitro sprouting in
response to VEGF stimulation.
Conclusion:
In summary, we found that cultured endothelial cells contain a subpopulation of CD34+ cells that
have hallmarks of tip cells. Therefore, CD34 as marker for tip cells in vitro allows us to study the
behaviour and regulation of tip cells in more detail, possibly resulting in novel therapeutic
possibilities to target tip cells to inhibit unwanted angiogenesis.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A38
Endocardial Prevalence of Coronary Collaterals Becomes Transmurally More Uniform in a
Human Heart with Left Ventricular Hypertrophy
Monique GJTB van Lier1, Elco R Oost1, Allard C van der Wal2, Jos AE Spaan1, Maria Siebes1 and
Jeroen PHM van den Wijngaard1.
1
2
Departments of Biomedical Engineering & Physics and Pathology, Academic Medical Center, Amsterdam.
Introduction:
Myocardial tissue ischemia may be limited by a well-developed coronary collateral circulation.
The aim of the study was to identify and quantify the coronary collaterals in a healthy human
heart and a hypertrophied heart with prior infarction.
Methods:
We obtained one heart with left ventricular hypertrophy (LVH), and an old inferior posterolateral
infarction (64 years, 595g) and one healthy heart without prior coronary artery disease (84 years,
330g). The coronary arteries were filled with fluorescent replica material. Hearts were frozen,
alternately cut and block-face imaged using an imaging cryomicrotome (voxel size 32 μm).
Collateral transmural distribution, connectivity and diameter were determined.
Results:
In the healthy heart, collaterals were mainly in the LV free wall (endo: 62%, mid: 7%, epi: 2%),
whereas a more uniform distribution over the LV free wall (endo: 10%, mid: 3%, epi: 4%) and
septum (endo: 56%, mid: 19%, epi: 3%) was found in the LVH heart. More intraCC (within major
branches) than interCC (between major branches) were detected in the healthy (12.93/g vs.
0.37/g) and LVH (1.39/g vs. 0.16/g) heart. InterCC were most prevalent between LCX-LAD (99%)
in the healthy and between LCX-LAD (68%) and LAD-RCA (28%) in the LVH heart. Most intraCC
were within the LAD and LCX territories in both hearts. The median CC diameter was larger in the
LVH compared to the heathy heart for intraCC (91.4 vs. 101.1 µm, p<0.001).
Conclusions:
Human hearts have abundant intra- and interCC collaterals. Transmural distribution was more
homogeneous in LVH, with fewer but larger collaterals.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A39
Use of antiplatelet agents is associated with intraplaque hemorrhage on carotid MRI
The Plaque At RISK (PARISK) study
Madieke I. Liem, Floris H.B.M. Schreuder , Anouk C. van Dijk, , Alexandra A.J. de Rotte, Martine
T.B. Truijman, Mat J.A.P. Daemen, Anton F.W. van der Steen, Jeroen Hendrikse, Aart J.
Nederveen, Aad van der Lugt, M. Eline Kooi, Paul J. Nederkoorn.
Academic Medical Center, Amsterdam.
Background and Purpose:
Intraplaque hemorrhage (IPH), visualised with MRI, has shown to be associated with the risk of
stroke in patients with carotid artery stenosis. The mechanisms of IPH development are partly
understood. In this study we investigated the association between clinical patient characteristics
and carotid IPH on high-resolution MRI.
Methods:
Patients participate in the Plaque At RISK (PARISK) study. This prospective, multicenter cohort
study included patients with recent amaurosis fugax, hemispheric TIA, or non-disabling stroke in
the internal carotid artery territory and an ipsilateral carotid stenosis of <70%, who were not
scheduled for carotid revascularisation procedure. One hundred patients, recruited between
2010 and 2012, underwent a 3-Tesla high-resolution carotid MRI. We documented clinical patient
characteristics and performed multivariable logistic regression analysis to investigate their
association with IPH.
Results:
IPH was observed in 45 patients (45%) in one or both carotid arteries. Male gender and use of
antiplatelet agents before the index event were associated with IPH in univariable analysis. In a
multivariable analysis, only prior use of antiplatelet agents was significantly associated with IPH
(OR 2.71, 95%CI: 1.12-6.61). Cardiovascular risk factors and a history of symptomatic arterial
diseases were not associated with IPH.
Conclusion:
In this cohort of 100 patients with recently symptomatic carotid stenosis, previous use of
antiplatelet agents is associated with carotid IPH on MRI. Antiplatelet therapy may increase the
risk of IPH, but our findings need to be confirmed in larger patient cohorts. The implications for
risk stratification remain to be determined.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A40
Secondary prevention for cardiovascular disease is challenging and differs between ethnic
groups: the HELIUS study
M. Minneboo1, S. Lachman1, M.B. Snijder2, K. Stronks2, R.J.G. Peters1
1
2
Department of Cardiology, AMC, Amsterdam. Department of Public Health, AMC, Amsterdam.
Introduction: Prevention of cardiovascular morbidity and mortality by improving lifestyle-related
risk factors is challenging. Data on ethnic differences for secondary prevention are lacking. We
evaluated lifestyle related risk factors among 6 ethnic groups living in Amsterdam in the
Netherlands.
Methods: The HEalthy LIfe in an Urban Setting (HELIUS) study included participants aged 18-70
years, originating from 6 different ethnicities: South-Asian Surinamese, African Surinamese,
Ghanaian, Turkish, Moroccan and Dutch origin living in a urban community in the Netherlands.
Prevalent cardiovascular disease (CVD) was defined as self-reported coronary heart disease,
stroke and/or PCI or coronary bypass surgery (CABG). Healthy lifestyle was defined as nonsmoking, BMI<25kg/m2 and physically active (>30 min/5x per week of moderate intensity by
SQUASH questionnaire).
Results: Out of 15951 participants, 833 patients (5.2%) reported a history of CVD [coronary heart
disease (n=277), stroke (n=278), PCI or CABG (n=473)]. Among those with a history of CVD, mean
age was 53 years and 44% were female. Non-smoking was predominant among Ghanaian (91%)
and Moroccan (88%). Prevalence of a BMI<25kg/m2 was overall low, Dutch had the highest
percentage (38%). Dutch reported to be the most physically active (73%).
Conclusion: A majority of the patients with CVD does not achieve guideline standards for lifestyle
related secondary prevention, with a low prevalence of BMI<25kg/m2 and physical activity.
Between ethnic groups, there are significant differences in lifestyle related risk factors. Since the
prevalence of lifestyle related risk factors was high across all ethnic groups, and without specific
determinants of the differences, increased efforts in non-ethnic specific secondary prevention
programs may be appropriate.
Table 1. Lifestyle related risk factors among those with self-reported CVD, stratified by ethnicity
Non smoking
BMI<25kg/m
2
Physically active
Dutch
South-Asian
Surinamese
African
Surinamese
Ghanaian
Turkish
Moroccan
n= 97
n=227
n=154
n=102
n=167
n=85
p value
59 (61%)
137 (60%)
85 (55%)
93 (91%)
111 (67%)
75 (88%)
<0.001
37 (38%)
68 (30%)
40 (26%)
20 (20%)
18 (10%)
18 (21%)
<0.001
71 (73%)
122 (54%)
94 (61%)
53 (52%)
79 (47%)
30 (35%)
<0.001
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A41
Administration of an Angiopoietin-1 mimetic attenuates microcirculatory perfusion
disturbances and vascular leakage during hemorrhagic shock
Michelle Trieu, 1,2, Matijs van Meurs,4,5, Anoek L.I. van Leeuwen 1,2, Tom Grandjean1,2, Nick J.
Koning 1, Paul van Slyke6, Dan J. Dumont 7, Leo M.G. Geeraedts Jr. 3, Christa Boer1, Charissa E. van
den Brom1,2
1
2
3
Departments of Anesthesiology , Physiology and Trauma Surgery , VUmc, Amsterdam. Departments of Pathology
4
5
6
and Medical Biology and Critical Care , UMCG, Groningen. Vasomune Therapeutics , Toronto, Canada. Department
7
of Medical Biophysics , University of Toronto, Toronto, Canada
Background:
Microcirculatory dysfunction following hemorrhagic shock (HS) is associated with multiple organ
failure and might be prevented by modulation of the Angiopoietin(Ang)/Tie2 system. This study
investigated effects of an Ang-1 mimetic on microcirculatory perfusion disturbances and vascular
leakage after HS.
Methods:
Anesthetized and mechanically ventilated male Wistar rats underwent sham surgery or HS
receiving placebo or Ang-1 mimetic. HS was induced by blood withdrawal to achieve a mean
arterial pressure (MAP) of 30mmHg for one hour, followed by resuscitation with crystalloids and
blood until baseline MAP was reached. Microcirculatory perfusion was measured using intravital
microscopy on the cremaster muscle. Vascular leakage was assessed by Evans Blue Dye (EBD)
extravasation.
Results:
One hour after HS, deterioration of microcirculatory perfusion was reflected by a 42% reduction
of the proportion of perfused vessels (PPV) and 91% increase of the proportion of stopped
vessels (PSV). In the placebo group, microcirculatory perfusion remained impaired despite fluid
resuscitation, whereas in the Ang-1 mimetic group fluid resuscitation restored microcirculatory
perfusion with a 30% increase in PPV and 22% decrease in PSV. Vascular leakage was increased in
liver (25±6 vs. 9±5EBD/g dry organ weight), lung (17±9 vs.11±2μg/g) and kidney (19±9 vs.
6±2μg/g) in shock group compared to controls, which was attenuated by Ang-1 mimetic.
Moreover, significant less blood was required to restore baseline MAP with Ang-1 mimetic
compared to placebo (1.1±1.1 vs. 2.9±0.9ml).
Conclusions:
Ang-1 mimetic administration protects against microcirculatory perfusion disturbances and
reduces the required amount of fluids after hemorrhagic shock possibly due to less vascular
leakage.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A42
Continuous vital signs monitoring using the VitalConnect MD Healthpatch
M.C. Hermans1, M.S. van Mourik1, J. Baan jr1, H. Hermens2, and M.M. Vis1
1: AMC Heart Center, University of Amsterdam 2: MIRA, institute of Technical Medicine and Biomedical technology,
University of Twente.
Introduction: Mobile health technology and wearable sensor technology is emerging to enable
continuous monitoring of multiple medical signals simultaneously. To explore the usability of
VitalConnect MD Healthpatch as a potential tool for remote patient monitoring, we verified
signal data quality of the corresponding monolead ECG, heart rate (HR) and respiratory
frequency (RF).
Methods: Five healthy subjects were monitored by the Healthpatch during various activities
including rest, metronome breathing, cycling and daily life activities. The ECG morphology, HR
and RF were validated using 6-lead reference ECG and respiratory flow (Dipha, Inbiolab,
Groningen). Visibility of P-waves and artifacts was scored by three investigators independently.
Results: The ECG showed good correspondence with the reference ECG during most tasks and
involved a mean maximal correlation of 0.9 in rest. P-waves were visible in 72-100% of the time,
but could not be distinguished for all recommended patch locations. Artifacts were particularly
seen during arm movement and sensor disturbances. The HR and RF parameters were associated
with a mean Pearson’s coefficient of 0.94 and 0.64 respectively, and were able to represent the
trends in the data with a minimal delay.
Conclusions: The Healthpatch provides accurate monolead ECG during most activities if placed in
specific chest position similar to accepted golden standards. The HR and RF parameters reflect
global changes. The Healthpatch might particularly be suitable for observation of trends in the
ECG, HR and RF. Further studies investigating the applicability in telemonitoring applications are
warranted.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A43
Cerebral Autoregulation and the Cerebrovascular Response to Head-of-Bed Positioning in
Acute Ischemic Stroke
Jasper Truijen1,2, MD; Line Sylvest Rasmussen3, MD; Yu-Sok Kim1,2, MD, PhD; Jan Stam4, MD, PhD;
Wim Stok2, MSc; Frank C. Pott3, MD, DMSc; Johannes J. van Lieshout1,2,5* MD, PhD.
1
2
Departments of Internal Medicine and Anatomy, Embryology and Physiology, Laboratory for Cardiovascular
Physiology, AMC Center for Heart Failure Research, Academic Medical Center, University of Amsterdam, The
3
Netherlands, Bispebjerg Hospital Research Unit for Anesthesia and Intensive Care, University of Copenhagen,
4
Denmark, Stroke Unit, Department of Neurology, Academic Medical Center, University of Amsterdam, The
5
Netherlands, MRC/ARUK Centre for Musculoskeletal Ageing Research, School of Life Sciences, University of
Nottingham Medical School, Queen's Medical Centre, United Kingdom
Introduction:
In patients with acute ischemic stroke horizontal head-of-bed positioning has the potential to
enhance cerebral perfusion, however with currently unexplained inter-individual heterogeneity.
It remains uncertain to what extent cerebrovascular autoregulation (CA) performance in patients
with ischemic stroke relates to the cerebrovascular response to horizontal head-of-bed
positioning.
Methods:
In patients with acute ischemic stroke and in reference subjects, bilateral cerebral CBF velocity by
transcranial Doppler ultrasound and near-infrared spectroscopy determined total hemoglobin
tissue concentration ([tHb]) as a measure for cerebral blood volume during head-of-bed lowering
from 30° to 0° were related to CA performance expressed as the phase difference (Φ) of the
arterial pressure-to-CBF velocity transfer function.
Results:
In both groups CBF velocity did not significantly increase following head-of-bed lowering whereas
parameters of cerebrovascular resistance and [tHb] increased in both patients and reference
subjects (+0.4±2.6 μM vs. +2.1±2.0 μM). In patients’ hemispheres with lower (< median Φ) CA
performance, [tHb] increased upon head-of-bed lowering vs. a decrease in hemispheres with
higher (> median Φ) CA performance (+1.0±1.3 μM vs. -0.5±1.0 μM; p<0.05). The [tHb] response
to head-of-bed lowering was inversely related to CA performance (r=−0.68; p<0.001) in the
patients, but not in the reference subjects.
Conclusions:
In patients with acute ischemic stroke vs. reference subjects cerebral autoregulation
performance impinges on the cerebrovascular response to changes in head-of-bed position.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A44
Derangements of microcirculatory perfusion after cardiopulmonary bypass continue in the
early postoperative period
N.A.M. Dekker1,2, D. Veerhoek3, N.J. Koning1, C.E. van den Brom1,2, A.B.A. Vonk4, C. Boer1
1
2
3
4
Depts. of Anesthesiology, Physiology, Perfusion and Cardio-thoracic surgery, Institute for Cardiovascular
Research, VU University Medical Center, Amsterdam..
Introduction:
Cardiopulmonary bypass (CPB) during coronary artery bypass grafting surgery (CABG)
deteriorates microcirculatory perfusion and contributes to postoperative organ failure. However,
the role of endothelial glycocalyx integrity and if microcirculatory perfusion postoperatively
restores is unknown. Therefore, this study investigated the course of microcirculatory perfusion
and glycocalyx integrity up to three days following CPB.
Methods:
This prospective observational study included 30 patients undergoing CABG with CPB.
Microcirculatory perfusion and perfused boundary region (PBR) were assessed using sublingual
sidestream dark-field imaging after anesthesia induction, initiation of CPB, infusion of cell
salvaged red blood cells (RBC), and 72 hours following surgery. Microvascular perfusion was
calculated by the perfused vessel density (PVD). PBR, an indirect parameter for glycocalyx
thickness, was calculated as the difference between RBC column width and the outer edge of the
RBC-perfused lumen.
Results:
Initiation of CPB was associated with increased PBR (2.2±0.3 μm vs. 1.9±0.3 μm; P=0.003) and
decreased PVD (11.4±3.2 mm/mm2 vs. 14.4±3.9 mm/mm2; P=0.001) compared to baseline
measurements. PBR was inversely correlated with PVD (r=-0.88; P=0.004), hemoglobin (r=-0.80,
P=0.03), hematocrit (r=-0.88, P=0.009) and capillary RBC filling (r=-0.98, P<0.001). Despite
retransfusion of cell salvaged blood at the end of CPB, PBR and PVD did not restore three days
after surgery.
Conclusions:
CPB-induced microcirculatory perfusion disturbances continue throughout the first postoperative
days. This is paralleled by increased perfused boundary region, suggesting impaired glycocalyx
integrity as a possible pathophysiological mechanism. Further studies should reveal whether
targeting these microvascular alterations could prevent the development of postoperative organ
failure after cardiac surgery.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A45
Optical Clearing Of Brain Tissue For Three-Dimensional Ultramicroscopic Imaging Of The
Cerebral Vasculature
Nadia Lachkar1, Judith de Vos1, Erik NTP Bakker1, Ed VanBavel1
1
Department of Biomedical Engineering and Physics, Academic Medical Center University of Amsterdam,
Meibergdreef 5 Amsterdam. [email protected],
Objectives: Whole brain imaging without time-consuming tissue sectioning, imaging and 3D
reconstruction can be performed by light-sheet laser-scanning ultramicroscopy. For this 3D
imaging technique, optical clearing of biological tissue is required. Several techniques have
become available to make mouse brains optically transparent. Here we report on clearing rat
brain tissue to recover the 3D architecture of the vascular bed.
Methods: Endothelial cells were labeled in vivo by intravenous injection of lectin (DyLight 488
Labeled tomato). The cerebral vasculature was visualized by the ultramicroscope (MCBI, VUmc).
We tested two methods for optical clearing of whole rat brains; passive CLARITY [1], which
involves tissue clearing by the transformation of lipids to a hydrogel network, and 3DISCO [2],
which uses organic solvents to match the refractory indexes of the different brain tissue layers.
Results: Clearing of adult rat brains using 3DISCO resulted in optically transparent tissue. The
entire protocol took 8 days and consisted of sequential 24-hour-incubation steps with an
increased concentration of tetrahydrafuran (i.e. 50-100%) followed by 2 days in dibenzylether. As
a result, we obtained a hard glass-like tissue sample. Although the ultramicroscope imaging of
the brain contained background noise, the images showed the vasculature with penetrating
arteries.
Passive clearing of adult rat brains with CLARITY took 3 months, which included hydrogel
embedding, lipid removal and glycerol incubation. From this protocol we retrieved an expanded
brain as a gel-like structure, which was optically clear. The cerebral arteries were clearly imaged
by the ultramicroscope. However, smaller arteries could not be distinguished from the brain
tissue due to background noise.
Conclusion: Rat brains could be cleared by 3DISCO and CLARITY and subsequently imaged by
ultramicroscopy. The obtained images contained background noise, likely due to the scattering of
light in the large brain tissue sample. Based on these preliminary data, in optical clearing of rat
brain samples for the purpose of imaging the vascular architecture, 3DISCO might be superior to
the CLARITY protocol.
References:
1. Chung, K. and K. Deisseroth, CLARITY for mapping the nervous system. Nat Methods, 2013. 10(6): p. 508-13.
2. Erturk, A., et al., Three-dimensional imaging of solvent-cleared organs using 3DISCO. Nature Protocols, 2012. 7(11): p. 1983-1995.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A46
Balancing TGFb/BMP in Pulmonary Arterial Hypertension
Nina Rol1, Robert Szulcek1, Marie José Goumans4, Chris Dickhoff3, Anton Vonk-Noordegraaf1,
Harm Jan Bogaard1, Geerten van Nieuw Amerongen2
1
2
3
Department of Pulmonary Medicine; Department of Physiology; Department of Surgery, VUmc, Amsterdam;
Department of Molecular Cell Biology, LUMC, Leiden.
4
Introduction:
Hemodynamic alterations contribute to disease progression in Pulmonary Arterial Hypertension
(PAH), a deadly lung disease associated with mutations in the Transforming Growth Factor beta
(TGFb) –Superfamily. Mutations of the bone morphogenetic protein receptor 2 (BMPR2) and
increased expression of TGFb support the idea for a central role of TGFb/BMP imbalance in PAH.
Therefore we investigated whether shear stress induces dysfunctional TGFb/BMP responses in
pulmonary microvascular endothelial cells (MVEC) from PAH patients.
Methods:
Control MVECs from cancer lobectomies were compared to PAH MVECs isolated from autopsy
and transplantation biopsies. Confluent cells were stimulated with TGFb1 or BMP9 or subjected
to shear stress, and downstream phosphorylation of Smad2/3 and Smad1/5 was quantified.
Reversal of shear stress induced responses was tested by addition of TGFb neutralizing antibody
1D11 and the selective inhibitor of the BMP type I receptor kinases LDN-193189.
Results:
Both control and PAH MVECs showed an increased phosphorylation of Smad2/3 upon TGFb1 and
shear stress stimulation, which was blocked by 1D11. Suprisingly, BMPR2 mediated responses
appeared to be intact and led to increased phosphorylation of Smad1/5 when exposed to BMP9
and shear stress, which was inhibited in presence of LDN-193189.
Conclusions:
Under the in vitro conditions used, both TGFb and BMP mediated signaling is intact in PAH
MVECs. Upon exposure to shear stress or ligand stimulation, control and PAH MVECs showed no
differences in downstream signaling of the TGF-β/BMP pathway. The effective inhibition of Smad
phosphorylation during shear stress shows the importance of TGFb ligands and BMPRI with
regards to shear adaptation in MVECs.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A47
A randomized trial of intra-arterial treatment for acute ischemic stroke – MR CLEAN
Olvert A. Berkhemer*, Puck S.S. Fransen*, Debbie Beume*, Lucie A. van den Berg, Hester F.
Lingsma, Wim H. van Zwam#, Yvo B.W.E.M. Roos#, Aad van der Lugt#, Robert J. van
Oostenbrugge#, Charles B.L.M. Majoie# and Diederik W.J. Dippel#, on behalf of the MR CLEAN
investigators
#
Academic Medical Center, Amsterdam. *, Authors contributed equally to the study.
Background:
In patients with acute ischemic stroke caused by a proximal intracranial arterial occlusion,
intraarterial treatment (IAT) is highly effective for emergent revascularization. However, proof of
a beneficial effect on functional outcome is lacking.
Methods:
Patients with a proximal arterial occlusion in the anterior cerebral circulation demonstrated on
vessel imaging, treatable by IAT within 6 hours after symptom onset, were randomized to IAT
plus usual care, or usual care alone. Primary outcome was the modified Rankin Scale (mRS) at 90
days; mRS is a categorical scale measuring functional outcome, with range from 0 (no symptoms)
to 6 (dead). The treatment effect was estimated with ordinal logistic regression as a common
odds ratio, adjusted for prespecified prognostic factors (acOR). The acOR is a measure of the
likelihood that treatment leads to better mRS scores (shift analysis).
Results:
We included 500 patients enrolled from 16 medical centers in the Netherlands. Mean age was 65
years (range 23 to 96), and 445 patients (89%) were treated with intravenous alteplase before
randomization. Retrievable stents were used in 190 of 233 patients (82%) allocated to IAT. The
acOR was 1.67 (95% CI: 1.21-2.30). There was an absolute difference of 13.5% (95% CI: 5.9% to
21.2%) in functional independence (mRS 0-2) in favor of the intervention (32.6% vs. 19.1%).
There were no differences in mortality or occurrence of symptomatic intracranial hemorrhage.
Conclusions:
Intraarterial treatment in patients with acute ischemic stroke caused by a proximal intracranial
occlusion of the anterior circulation is effective and safe within 6 hours after stroke onset.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A48
Intraleaflet Hemorrhage in Cardiac Valves
Olga CG Stam1, Mat JAP Daemen1, Allard C van der Wal1
1
Department of Pathology, Academic Medical Center – University of Amsterdam, Amsterdam, the Netherlands
Introduction: Similar to hemorrhage in atherosclerotic plaques, hemorrhage in cardiac valves
could potentially contribute to the increase in cardiac valve mass. Such a concept of occurrence
of intraleaflet hemorrhage (ILH) has received only little attention. In this study the prevalence
and the potential source of ILH is investigated, in a series of surgically resected symptomatic
cardiac valves with different etiologic backgrounds.
Methods: Pathology specimen of 100 consecutive patients who underwent cardiac valve surgery
for symptomatic aortic and/or mitral valve disease, respectively 85 and 20 valves, were selected.
The valves were histologically and immunohistochemically examined for the presence and age of
ILH and the association of ILH with neo-angiogenesis, calcification and/or microvascular leakage.
ILH was categorized in (1) ‘recent’ if only extravascular localization of intact erythrocytes was
found, (2) ‘old’ when the tissue contained erythrocyte fragments and/or iron deposits, and (3)
‘ongoing’ when both features of ‘recent’ and ‘old’ ILH were found.
Results: ILH was found in 73 aortic valves (86%), containing all cases with post-inflammatory
(rheumatic) and infectious changes and 59 degenerative altered valves. Most aortic valves, 48
out of 73, were labeled as ongoing ILH. Recent ILH was found in 14 cases and 11 cases were
categorized as old ILH. Hemorrhage was found between calcium deposits in 69 valves.
Microvessels were found in almost 95% of the aortic valves. 52 out of 73 valves showed
microvascular leakage in the anti-vWF immunostain. A total of 38 valves showed microvascular
leakage in proximity of the hemorrhage (52%).
In 15 out of 20 mitral valves (75%) ILH was seen, containing 78% of the post-inflammatory
(rheumatic) and 80% of the infectious cases and 67% of the degenerative cases. Ongoing ILH was
scored in 67% of the mitral valves. Recent ILH was found in 2 cases and 3 cases fit the criteria for
old ILH. In 7 valves hemorrhage was found between calcium deposits. In all mitral valves
microvessels were found, of which 93% showed microvascular leakage. Eight valves showed
microvascular leakage in proximity of the hemorrhage (53%).
Conclusion: The prevalence of ILH in diseased cardiac valves is very high, both in aortic and mitral
valves and irrespective of the type of underlying. In approximately half of the cases ILH was
associated with microvascular leakage. These findings suggest that, similar to the situation in
atherosclerotic plaques were intraplaque bleeding provokes plaque growth; ILH may contribute
to the increase in cardiac valve mass and therefore the degree of cardiac valve dysfunction.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A49
Colchicine aggravates Coxsackievirus B3 infection in mice
Linde Woudstra1,4,* ; Bernard J Smilde1,* ; Gene Fong Hing1 ; Diana Wouters 5 ; Sacha Zeerleder5,6 ;
Jean-Luc Murk7 ; Marieke van Ham5; Lynda J. Juffermans2,4 ; Albert C. van Rossum2,4 ; Hans W.M.
Niessen1,2,3 Paul A.J. Krijnen1,2,# ; Reindert W. Emmens1,3,4,#
1
2
3
Department of Pathology, Cardiology, Cardiac Surgery, VU University Medical Center, Amsterdam, the
4
5
Netherlands. ICaR-VU, VU University Medical Center, Amsterdam. Department of Immumopathology, Sanquin
6
7
Research, Amsterdam. Department of Hematology, Academic Medical Center, Amsterdam. Department of
Virology, Utrecht Medical Center, Utrecht.
#
*Both authors contributed equally to this study, Both authors contributed equally to this study
Introduction:
Clinical trials have found the anti-inflammatory drug colchicine to be safe and effective in
treating pericarditis. Pericarditis is often caused by a viral infection and often coincides with
myocarditis. No anti-inflammatory treatment option exists for patients with acute viral
myocarditis however. Here, we investigated colchicine treatment in a mouse model of acute
coxsackievirus B3(CVB3)-induced myocarditis.
Methods:
Four groups of male C3H/HeJ mice (Harlan) were used. Included was an uninfected control group
(n=8), a group only infected with CVB3 (1x105 virions, n=10), a group only treated with colchicine
(2 mg/kg, n=5) and a group that received both CVB3 and colchicine (n=10). Both the heart and
the pancreas were analysed for tissue damage (histology), lymphocytes, macrophages and
neutrophils (immunohistochemistry) and viral RNA/protein levels (qPCR /
immunohistochemistry).
Results:
The experiment was terminated 3 days post-infection, as the group that received both CVB3 and
colchicine became terminally ill. Colchicine treatment of CVB3-infected mice resulted in near
complete destruction of the pancreatic acini, while the Langerhans islets were largely unaffected.
In both the heart and the pancreas, colchicine treatment of CVB3-infected mice resulted in
significantly increased number of neutrophils (p<0.01), but a significantly decreased number of
macrophages (p<0.01). Lymphocyte numbers were unaffected. Lastly, both pancreas and heart
had a significantly higher viral presence as result of colchicine (p<0.001).
Conclusions:
Colchicine treatment causes severe illness in CVB3-infected mice, likely caused by increased viral
infiltration and inflammatory damage in the pancreas. This suggests that colchicine may not be
suitable to treat patients with acute viral myocarditis.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A50
Supra-physiological shear stress unveils novel dysfunction in microvascular and circulating
endothelial cells from patients with Pulmonary Arterial Hypertension
Robert Szulcek1,2, J Smits1,2, N Rol1,2, C Happe1,2, A Vonk-Noordegraaf1, P Koolwijk2, and HJ
Bogaard1
Department of 1Pulmonology and 2Department of Physiology, Institute for Cardiovascular Research (ICaR-VU), VU University Medical Center,
Amsterdam, The Netherlands
Introduction:
Endothelial cell (EC) dysfunction and altered pulmonary hemodynamics are hallmarks of
Pulmonary Arterial Hypertension (PAH), a deadly lung disease characterized by severe vascular
remodeling and progressive narrowing of small pulmonary arteries. However, the exact
contributions of high fluid shear stress (HSS) to the cellular and vascular alterations in PAH are
unclear. We hypothesized that HSS is the pivotal source for EC injury and promoter of defective
vascular repair required for the remodeling in PAH.
Methods:
Microvascular EC (MVEC, n=7), Pulmonary Artery EC (PAEC, n=3) and Endothelial Colony Forming
Cells (ECFC, n=7) isolated from PAH patients with different etiologies were subjected to
physiological (LSS, 2.5 dyn/cm2) and supra-physiological (HSS, 16 dyn/cm2) rates of pulsatile shear
stress and compared to healthy controls.
Results:
PAH MVEC presented significantly impaired barrier integrity (8653±315 vs. 11249±209 ohms,
p=0.007), significantly decreased wound-healing capability (15.3±3.6 vs. 24.0±2.9 µm/h, p=0.003)
and significantly delayed morphological shear-adaptation (39.6±4.0 vs. 23.7±4.2% not aligned EC
at 72 h HSS, p=0.025). The delayed shear-response increased susceptibility to shear-induced
injury, evidenced by cell loss at sites characterized by non-uniform flow profiles. PAEC isolated
from the same PAH lung showed no impairments and a functional shear-response. PAH ECFC on
the contrary expressed a similar shear-sensing defect to PAH MVEC, which became manifest
under HSS, but not LSS. In addition ECFC continued proliferating under HSS reaching cell densities
almost twice as high as the healthy controls (40,000 vs. 72,000 cells/cm2, p=0.004). Protein
cleavage of the shear-sensor PECAM-1 was found responsible for the defective shearresponsiveness of PAH MVEC, whereas the reason for the hyper-proliferative phenotype of PAH
ECFC remains elusive.
Conclusion:
Our data point towards the conclusion that HSS contributes to the vascular remodeling in PAH by
increasing vulnerability of PAH MVEC to shear-induced injury and promoting exuberant
outgrowth of ECFC from the site of injury. Thereby treatment strategies to normalize pulmonary
blood flow and targeting ECFC proliferation could prevent and/or reverse vascular remodeling in
PAH.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A51
Calcification associates with aortic distensibility in Marfan Syndrome mice.
Shaynah A. Wanga1,2,3, Stijntje Hibender 1, Romy Franken2,3, Luigi van Riel1, R Yanto Ridwan4, J
Nicole van Vliet4, Jeroen Essers4, Inge van der Made5, Yigal M Pinto3,5, Maarten Groenink2,3,
Aeilko H Zwinderman6, Barbara JM Mulder2,3, Carlie JM de Vries1, Vivian de Waard1,2.
1) Dept. Medical Biochemistry, AMC, Amsterdam 2) Interuniversity Cardiology Institute of the Netherlands 3) Dept.
Cardiology, AMC, Amsterdam 4) Dept. Cell Biology and Genetics, Erasmus MC, Rotterdam. 5) Dept. Experimental
Cardiology, AMC, Amsterdam. 6) Dept. Clinical Epidemiology and Biostatistics and Bioinformatics, AMC, Amsterdam
Aim: We aim to study calcification in mouse and human Marfan aortic tissues and its potential
participation in aortic dilatation.
Methods and Results: Calcification is observed in the aortic sinus and aorta ascendens of Marfan
patients, as determined by alizarin red staining. Alcian blue staining revealed increased
accumulation of glucosaminoglycans(GAGs) in Marfan patients compared to control. In human
Marfan aorta, alcian blue staining is also used as marker for medial necrosis, since these GAGs
tend to accumulate at sites of smooth muscle cell loss. Mostly GAGs are known to be present in
bone, where they are visualized by alcian blue. Whether these GAGs contribute to aortic
calcification/stiffness is still unclear. In a Fibrillin-1 mutation mouse model of Marfan
(FBN1C1039G/+), alcian blue staining revealed osteoblast-like cells surrounded by GAGs in the
medial layer of the aortic sinus. In these mice, the percentage of mice with osteoblast-like cells
increased with age, unlike in wild type mice. Ex vivo imaging of the Marfan aorta revealed
increased incorporation of the fluorescent OsteoSense probe, specifically in the aortic sinus and
ascending aorta. In the ascending aorta no osteoblast-like cells were observed, however, there is
a strong positive correlation between Osteosense signal and aorta ascendens diameter. In
addition, Osteosense signal negatively correlated with aorta ascendens distensibility.
Conclusions: Calcification is present in the aortic sinus and aorta ascendens of Marfan patients
and Marfan mice. The degree of calcification negatively correlates with the aorta ascendens
distensibility, suggesting that not only elastic lamina degradation (common in Marfan) may be
responsible for increased aortic stiffness in Marfan, but also increased calcification processes.
Figure legend. osteosense was injected intra peritoneally and visualized ex vivo using odyssee. Most of the
osteosense is localised in the vascular wall of the ascending aorta. There is minimal signal in the descending aorta (A)
Significantly more osteosense is incorporated in the Marfan aortas compared to wildtype (B).
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A52
FGF23 IN PART EXPLAINS IMPAIRED ENDOTHELIAL FUNCTION AND MYOCARDIAL PERFUSION
OBSERVED IN EXPERIMENTAL CHRONIC KIDNEY DISEASE
M Verkaik1,2, PM ter Wee1, MG Vervloet1, EC Eringa2
1
2
Department of Nephrology, and Department of Physiology, VU University Medical Center, Amsterdam
On behalf of the NIGRAM consortium
Introductions and Aims:
Cardiovascular causes account for approximately 50% of mortality in patients with chronic kidney
disease (CKD). FGF23, a phosphate-lowering protein and elevated in CKD, is associated with
cardiovascular mortality and endothelial dysfunction. We hypothesized that CKD impairs vascular
function and myocardial perfusion and that this can be attributed to FGF23.
Methods:
Eight weeks old male wild type C57Bl/6J mice were subjected to partial nephrectomy (5/6Nx) or
sham-surgery, and after 6 weeks mice were subjected to myocardial contrast echocardiography
(MCE) to test myocardial perfusion and a pressure myograph setup to test ex vivo vascular
function. To examine whether FGF23 contributes to the cardiovascular phenotype observed in
CKD, a second group received either PBS or FGF23 i.p. injections for 7 consecutive days twice
daily.
Results:
Plasma FGF23 significantly increased after Nx surgery (1.7-fold p<0.01), as well as fractional
excretion of phosphate (FEP) (4-fold p<0.001). 5/6Nx blunted ex vivo vasodilator responses to
acetylcholine (p<0.05), whereas responses to sodium nitroprusside (SNP) or endothelin were
normal. Long-term in vivo FGF23 injections completely mimicked this vascular defect, but shortterm ex vivo FGF23 administration did not change vascular reactivity. 5/6Nx mice showed
reduced myocardial blood volume during acetylcholine infusion (-17%, p=0.023) and during SNP
infusion (-13%, p=0.014).
Conclusions:
Renal failure compromises myocardial perfusion. In addition, chronic kidney disease impairs
endothelium-dependent vasodilation, which is mimicked by FGF23 injections in healthy mice.
FGF23 thus might explain endothelial dysfunction in CKD.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A53
THROMBIN-ACTIVATABLE FIBRINOLYSIS INHIBITOR (TAFI) PEPTIDES SELECTIVELY MODULATE
THROMBOMODULIN-DEPENDENT TAFI ACTIVATION
T. Plug1; P.F. Marx1; J.C.M. Meijers1,2
1
Department of Experimental Vascular Medicine, Academic Medical Center, Amsterdam.
Department of Plasma Proteins, Sanquin Research, Amsterdam.
2
Background:
Activated thrombin-activatable fibrinolysis inhibitor (TAFI) protects a blood clot from premature
lysis. Elevated levels of TAFI have been associated with coronary heart disease. TAFI is activated
by thrombin or the thrombin-thrombomodulin complex. Specifically targeting TAFI activation can
be a powerful strategy for the treatment of both thrombosis and bleeding. The aim of this study
was to generate TAFI peptides that modulate TAFI activation.
Methods:
34 peptides, based on the amino acid sequence of TAFI, were synthesized. The enzyme kinetics
were determined to quantify the effects of these peptides on TAFI activation by thrombin or the
thrombin-thrombomodulin complex.
Results:
Peptides 2 (Arg12-Glu28) and 34 (Cys383-Val401) inhibited thrombomodulin-dependent TAFI
activation by thrombin in the µM range. However, no inhibition was observed in the absence of
thrombomodulin. Arg12 of TAFI is known to play a significant role in thrombomodulin-dependent
TAFI activation but it is not known whether Arg12 binds to thrombomodulin directly or whether
Arg12-cleavage leads to TAFI binding to thrombomodulin. Substitution of Arg12 in peptide 2 did
not alter the inhibitory effects and thus, is in favor with the Arg12-cleavage hypothesis.
Furthermore, substitution of Trp395 to alanine in peptide 34 led to an almost complete loss of its
inhibitory effect, suggesting an important role for Trp395 in thrombomodulin-dependent TAFI
activation.
Conclusions:
TAFI-derived peptides reveal that Thr13-Glu28 and Cys383-Val401 in TAFI are involved in
thrombomodulin-dependent TAFI activation. TAFI activation in the absence of thrombomodulin
was not affected, therefore these peptides can be used for the design of potent, selective TAFI
activation inhibitors.
Keywords:
Thrombin-activatable fibrinolysis inhibitor (TAFI), Thrombin, Thrombomodulin, Fibrinolysis
inhibitors
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A54
The ThRombolysis in UnconTrolled Hypertension (TRUTH) study: an observational study on
treatment strategy of elevated blood pressure in stroke patients eligible for IVT
T.P. Zonneveld, A. Algra, D.W.J. Dippel, L.J. Kappelle, R.J. van Oostenbrugge, Y.B.W.E.M. Roos,
M.J. Wermer, H.B. van der Worp, P.J. Nederkoorn, N.D. Kruyt.
Introduction:
Intravenous thrombolysis (IVT) with recombinant tissue plasminogen activator is an effective
treatment in acute ischemic stroke. However, IVT is contraindicated when blood pressure (BP) is
above 185/110 mm Hg, because of an increased risk on symptomatic intracranial hemorrhage
(sICH). In Dutch clinical practice, two distinct strategies are used in this situation. The active
strategy comprises lowering BP with antihypertensive agents below these thresholds to allow
start of IVT. In the conservative strategy, IVT is administered only when BP drops spontaneously
below protocolled thresholds. Our main hypothesis is that the active strategy leads to a better
functional outcome three months after acute ischemic stroke.
Methods:
The TRUTH is a prospective, observational, cluster-based, parallel group, follow-up study, in
which participating centers continue their current local treatment guidelines. Outcomes of
patients admitted to centers with an active will be compared to those admitted to centers with a
conservative strategy.
We based our power analysis on an ordinal analysis of the modified Rankin Scale with the
“proportional odds” model, using the treatment effect found in a recent retrospective analysis.
Corrections for expected imbalance in group size and clustering effects resulted in a required
sample size of 1235 patients.
Discussion:
The TRUTH is the first large prospective study specifically studying IVT-candidates with elevated
BP, and has the potential to change clinical practice and optimize acute stroke care in these
patients.
Study progress:
As of September 1st 2015, 36 Dutch centers have agreed to participate. The study started in 30
centers, resulting 61 included patients.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A55
Rapid uptake and slow restitution of mitochondrial calcium in adult rat cardiomyocytes
Rob C.I. Wüst1, Michiel Helmes1,2, Jeroen W. Kole1, Jody L Martin3, René JP Musters1 and Ger JM
Stienen1
1
Department of Physiology, Institute for Cardiovascular Research, VU University Medical Centre, Amsterdam. 2IonOptix
3
LLC, Milton, MA, USA. Cell and Molecular Physiology, Loyola University, Chicago, USA
Introduction: Calcium plays a role in regulating cardiac contractile activity and mitochondrial
energy production and thus plays a pivotal role in matching energy supply and demand in cardiac
muscle. However, information on the uptake and release of Ca2+ by mitochondria in intact
cardiac cells operating at different workloads is scarce. We determined the magnitude and speed
of uptake and release of Ca2+ by mitochondria in intact cardiac cells and evaluated their
dependencies on stimulation frequency and external calcium concentration in intact cardiac
myocytes.
Methods: Using adenoviral infection, a ratiometric mitochondrially-targeted FRET-based calcium
indicator (4mtD3cpv, MitoCam) was expressed and the free mitochondrial calcium concentration
([Ca2+]mito) was measured in cultured adult rat cardiomyocytes at different stimulation
frequencies (0.1-4 Hz) and external calcium concentrations (1.8-3.6 mM) at 37oC (Figure).
Results: The increases in [Ca2+]mito during electrical stimulation at 0.1 Hz were rapid (t10-90%=48±2
ms), while the decreases in [Ca2+]mito occurred more slowly (t50%=1.24±0.07 s). This asymmetry
resulted in a gradual rise in [Ca2+]mito at elevated stimulation frequencies. [Ca2+]mito at rest, the
magnitude of Ca2+-uptake and the speed of Ca2+-release increased when the external calcium
concentration was increased. Inhibition of the mitochondrial sodium-calcium exchanger by
CGP37157, resulted in a rise in diastolic [Ca2+]mito and -paradoxically- in an acceleration of Ca2+release.
Conclusions: The rapid increase in [Ca2+]mito allows for a fast adjustment of mitochondrial energy
production to changes in demand in cardiac muscle. The effects of CGP37157 on mitochondrial
Ca2+-release suggest that other extrusion processes act in parallel with the mitochondrial sodiumcalcium exchanger.
2+
2+
Figure: MitoCam targeting and temporal relations between sarcomere shortening, free [Ca ]cytosol and [Ca ]mito. (A)
MitoCam expression in adult cardiomyocytes (scale bar: 100 µm). (B) Colocalization of MitoCam (green) and
2+
MitoTracker Red (red) and merge (left). (C) Sarcomere shortening, cytosolic calcium (Fura-4) and [Ca ]mito in
electrically stimulated (at t=1s) cardiac myocytes at 0.1 Hz.
A
C
S a rc o m e re
C y to s o lic
le n g th
[C a
2 +
]
B
M ito c h o n d r ia l
[C a
2 +
]
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A56
Genome-wide profiling of nuclear receptor Nur77 by ChIP-seq reveals interplay with NF-kB and
YY1 in regulation of inflammatory signaling
Duco S Koenis, Anouk AJ Hamers, Claudia M van Tiel, Wilbert Zwart, Carlie JM de Vries
1. Department of Medical Biochemistry, Academic Medical Center, University of Amsterdam, Meibergdreef 15,
1105AZ Amsterdam. 2. Division of Molecular Pathology, Netherlands Cancer Institute, Amsterdam.
Introduction:
Nur77 belongs to the NR4A family of nuclear receptors. It has an anti-inflammatory function in
macrophages and is critical for bone marrow differentiation and survival of Ly6Clo (“patrolling”)
monocytes.
Methods:
We performed ChIP-sequencing on RAW264.7 mouse macrophages with inducible
overexpression of Nur77 to gain insight into the genome-wide binding profile of this nuclear
receptor.
Results:
We identified 3350 Nur77-binding sites, of which 10.3% are located within 3 kb upstream of a
transcriptional start site. Motif analysis on the underlying genomic DNA sequences identified the
classical Nur77 response element (NBRE; 5’-AAAGGTCA-3’) as the most highly enriched motif
across the dataset (p-value = 10-2295), thereby providing support for the validity of our approach.
Motif analysis on our dataset revealed significant enrichment of response elements for the
transcription factor Yin-Yang 1 (YY1), which has recently been identified as an NF-kB target gene.
YY1 in-turn regulates the expression of the genes Cx3cr1 (Fractalkine receptor), Cxcl12 (Stromal
cell-derived factor 1), and miR29 (microRNA miR-29), which play an important role in
monocyte/macrophage migration and macrophage activation. We show that Nur77 and YY1
interact with each other and mediate opposing effects on the expression of these three genes.
Conclusions:
ChIP-sequencing for the nuclear receptor Nur77 in macrophages allowed us to identify an
interaction between Nur77 and the transcription factor YY1 regulating the expression of the
Cxcl12 gene.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A 57
Sudden cardiac arrest and rare genetic variants in the community
Annalisa Milano1*, Marieke T. Blom1*, Elisabeth M. Lodder1*, Daniel A. van Hoeijen1, Julien Barc1,
Tamara T. Koopmann1, Abdennasser Bardai1, Leander Beekmanc1, Peter Lichtner2, Maarten P.
van den Berg3, Arthur A.M. Wilde1,4, Connie R. Bezzina1, Hanno L. Tan1
1) Department of Clinical and Experimental Cardiology, Heart Center, Amsterdam Medical Center, University of
Amsterdam, Amsterdam, the Netherlands; 2) Institut für Humangenetik, Helmholtz Zentrum, Technische Universität
München, Munich, Germany; 3) Department of Cardiology, University Medical Center Groningen, University of
Groningen, Groningen, the Netherlands; 4) Princess Al-Jawhara Al-Brahim Centre of Excellence in Research of
Hereditary Disorders, Jeddah, Kingdom of Saudi Arabia
* These authors contributed equally to this work
Introduction:
Sudden cardiac arrest (SCA) ranks among the most common causes of death worldwide. Because
SCA is most often lethal, yet mostly occurs in individuals without previously known cardiac
disease, the identification of patients at risk for SCA could save many lives. In unselected SCA
victims from the community, common genetic variants (which are not disease-causing per se, but
may increase susceptibility to VF) are associated with increased SCA risk. However, whether rare
genetic variants contribute to SCA risk in the community is largely unexplored.
Methods:
We here investigated the involvement of rare genetic variants in SCA risk at the population level,
by studying the prevalence of six founder genetic variants present in the Dutch population (PLNp.Arg14del, MYBPC3-p.Trp792fsX17, MYBPC3-p.Arg943X, MYBPC3-p.Pro955fsX95, PKP2p.Arg79X, and the Chr7q36 idiopathic ventricular fibrillation risk-haplotype) in a cohort of 1440
unselected Dutch SCA victims included in the AmsteRdam REsuscitation Study (ARREST).
Results:
The six studied founder mutations were found to be more prevalent (1.1%) in the ARREST SCA
cohort compared to an ethnically and geographically matched set of controls (0.4%, n=1379,
p<0.05) or a set of Dutch individuals drawn from the Genome of the Netherlands (GoNL) study
(0%, n= 500 p<0.02).
Conclusion:
This finding provides proof-of-concept for the notion that rare genetic variants contribute to
some extent to SCA risk in the community.
Acknowledgements:
We acknowledge the support from the “Netherlands CardioVascular Research Initiative”: the Dutch Heart
Foundation, Dutch Federation of University Medical Centres, the Netherlands Organisation for Health Research and
Development and the Royal Netherlands Academy of Sciences (PREDICT project).
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A58
Impaired insulin-mediated myocardial microvascular recruitment and cardiac function in early
obesity are associated with decreased activity of AMPK
Chantal. A. Boly1,2, Christa Boer1, Rob F.P. van den Akker1,2, Etto C. Eringa2, Charissa E. van den
Brom1,2
1
2
Departments of Anesthesiology and Physiology, Institute for Cardiovascular Research, VU University Medical Center
Amsterdam, the Netherlands, [email protected]
Background and aims: Obesity-induced microvascular dysfunction has been suggested to
challenge myocardial perfusion and function, but the exact mechanism remains to be
established. Here we hypothesized that mild obesity impairs basal and insulin-mediated
myocardial perfusion and function. As AMP-activated protein kinase (AMPK) determines eNOS
activity and substrate metabolism, which are involved in the regulation of perfusion and function
of the myocardium, we assessed whether obesity reduced AMPK activity in the myocardium.
Materials & Methods: Male C57Bl6 mice were exposed to a western (n=19) or control diet
(n=17). After 4 weeks, M-mode echocardiography was used to determine myocardial function,
and myocardial contrast-echocardiography (MCE) for myocardial perfusion which was defined as
the product of microvascular filling velocity and microvascular blood volume. The protocol was
performed at baseline and during hyperinsulinemic-euglycemic clamp-induced hyperinsulinemia.
After sacrifice, phosphorylation status of ACC1, ACC2 and AMPKα were determined by western
blotting in the left ventricle.
Results: Western diet feeding induced obesity (27.0±0.4 versus 23.8±0.3 grams in control mice,
p<0.0001) without insulin resistance, hyperglycemia or hypertension. At baseline, myocardial
perfusion was 43% lower in obese mice compared to control mice (0.004 ± 0.001 versus 0.007 ±
0.001 A.U., p=0.03).Cardiac function, determined as fractional shortening, was impaired in obese
mice (43±2% versus 56±3% in controls, p=0.0008). There was a significant correlation between
baseline myocardial microvascular blood volume and contractile function (Pearson’s r=0.5,
p=0.048). Hyperinsulinemia increased microvascular blood volume and therefore myocardial
perfusion (to 0.01 ± 0.001 A.U., p=0.001) in healthy mice,which was blunted in obese mice
(p=0.04 for interaction). Obesity strongly reduced AMPK activity, as determined by
phosphorylation of its substrates ACC1 and ACC2 (46% and 49%, p=0.01 and 0.02). This was not
caused by decreased phosphorylation of AMPKα, which was increased 2.6-fold in myocardium of
obese mice.
Conclusion: Mild obesity impairs insulin-mediated microvascular recruitment and cardiomyocyte
contractility in murine myocardium. These myocardial microvascular defects precede and are
independent from whole-body insulin resistance and are associated with decreased myocardial
AMPK activity.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A59
The Role of IRS2 in PVAT in the Regulation of Local Muscular Perfusion and Glucose Uptake
Ali Turaihi, Femke Hoevenaars, Zeineb Gam, Yvo Smulders, Victor van Hinsbergh, Ed Eringa
ICaR-VU, VU University Medical Center, Amsterdam.
Introduction:
Perivascular adipose tissue (PVAT) plays an important role in the regulation of local muscle
perfusion. Insulin receptor substrates (IRS) 1 and IRS2 have shown strong yet incompletely
explained relationships to type 2 diabetes and cardiovascular disease in genome-wide association
studies. Moreover, insulin sensitivity of adipose tissue has been shown to control muscle insulin
sensitivity. To what extent does IRS2 signaling play a role in PVAT's function is still largely
unknown. Here, we tested the hypothesis that IRS2 regulates the vasoactive properties of PVAT.
Methods:
We isolated IRS2+/+ and IRS2-/- PVAT from 8 week old congenic strain C57/Bl6SV129 carrying a
genetic mutation. We then isolated muscle resistance arteries from C57/Bl6 (Bl6) mice and
studied insulin-mediated vasoreactivity ex vivo in the pressure myograph in the presence or
absence of IRS2+/+, IRS2-/- or Bl6 PVAT. Basal constriction of the arteries was achieved by KCL
[4.7 mM], The integrity of the endothelial cell layer was tested using 10-7 mM of acetylcholine
(Ach) at the end of the experiments.
Results:
Basal tone among all arteries (n=42) was 55.5% (SD=8.6). Insulin failed to induce diameter change
in arteries without the presence of PVAT (N=9). Insulin [2.0 nM] induced 26% (SD= 25.7; p=0.03)
and 17% (SD=14.5; p=0.016) increase in diameter in the presence of Bl6 PVAT (N=14) and IRS2+/+
PVAT (N=9), respectively, when compared to arteries without PVAT. On the other hand, insulin
induced 7% (SD=14.9) decrease in diameter in the presence of IRS2-/- PVAT (N=10) which was not
significantly different from the arteries without PVAT.
Conclusion:
IRS2 plays a major role in PVAT signaling and when absent, PVAT fails to unveil insulin-mediated
vasodilation in skeletal muscle arterioles.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A60
The Prevention of Dementia by Intensive Vascular Care (preDIVA) trial –preliminary blood
pressure findings
L.S.M. Eurelings1, E. Richard1, E.P. Moll van Charante2, E. van Bussel2, J. van Dalen1, M.
Hoevenaar-Blom1, S.A. Ligthart2, W.A. van Gool1
1
2
Department of Neurology, Academic Medical Center, Amsterdam Department of General Practice, Academic
Medical Center, Amsterdam, PreDIVA Study Group*
Introduction: Cardiovascular risk factors, including hypertension, are associated with an
increased risk of dementia. Whether a multi-component intervention strategy targeting vascular
risk factors can prevent or postpone dementia in older individuals is yet unknown. Here we show
preliminary findings regarding the effect of intensive vascular care (IVC) on blood pressure (BP).
Methods: A cluster-randomized trial (Prevention of Dementia by Intensive Vascular Care
(preDIVA)) was carried out in primary care to evaluate whether nurse-led IVC decreases the risk
of dementia and disability in older individuals (70-78 years, n=3532). The intervention consisted
of tailor-made life-style advice delivered in 4-monthly visits to a practice nurse, and if indicated,
initiation or improvement of medical therapy based on national guidelines. The effect of the
intervention on BP was assessed stratified by baseline hypertension status according to the
World Health Organization classification of BP.
Results: A final follow-up visit was completed in 1933 (54.7%) individuals. During 6-8 years
follow-up, mean BP decreased from 156 to 151 mmHG in the intervention group (nbaseline=1892)
and from 154 to 151 mmHG in the control group (nbaseline=1637) (∆2.5 mmHG, p=0.02). The
effect of the intervention was larger for participants with a higher baseline BP, with the largest
benefit for those with grade III hypertension (∆6.2 mmHG, p=0.05) (figure 1).
Conclusion: Preliminary results regarding BP indicate a small beneficiary effect of the
intervention on BP, with the largest benefit for individuals with grade III hypertension. Main
results regarding primary and secondary outcomes, including dementia and mortality, are
expected shortly.
Figure 1: trend in systolic blood pressure for individuals with baseline hypertension grade III
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A61
Understanding the role of KBTBD13 in striated muscle function and in nemaline myopathy
Vaishali Kakkar1, Josine M. de Winter1, Julia C. Kuszewski1, Diederik Kuster1, Jolanda van der
Velden1, Coen A.C. Ottenheijm1.
1
Institute of Cardiovascular Research (ICaR-VU), Dept. of Physiology, VU University Medical Center, Amsterdam
Introduction:
Nemaline myopathy (NM) is a clinically and genetically heterogeneous disease that is
characterized by muscle weakness and the presence of rod-like structures (i.e. nemaline bodies)
in muscle fibers. It is among the most common non-dystrophic congenital myopathies. A recent
gene implicated in NM is KBTBD13 - a member of the kelch protein family (disease referred as
NM-KBTBD13). Only little is known regarding the functions of KBTBD13, which might include a
though it is shown to work as an adaptor to Cullin-3 ligase, thus implicating its role in the
ubiquitin-proteasome degradation system (UPS).
Methods and results:
In the current study, we used a C2C12 cellular model, which forms myotubes on differentiation,
to study the functioning of KBTBD13 and its role in NM. We used a mycDDK-KBTBD13 plasmid to
transfect the cells and various biochemical assays as read out. We observed that mRNA levels of
KBTBD13 go up with differentiation of C2C12 cells. To elucidate the cellular localization of
KBTBD13, we performed immunocytochemistry. Our preliminary results show a cytosolic
localization in undifferentiated cells whereas a typical perinuclear staining of KBTBD13 occurs
upon differentiation. This might indicate that KBTBD13 localizes with the Golgi apparatus or the
endoplasmatic reticulum. Furthermore, we treated the cells with MG132, a proteasomal
inhibitor, in the presence or absence of KBTBD13. The expression of KBTBD13 lead to a slight
increase in the ubiquitination pool, indicating a role in the stabilization of ubiquitination of
targeted substrates. The direct interacting partners of KBTBD13 are still unknown, and therefore
in future work we aim to perform proteomics analyses on C2C12 cells that overexpress KBTBD13
We are also generating disease associated mutations in KBTBD13 (Arg248Ser, Lys390Asn,
Arg408Cys) to more closely study the disease phenotype associated with NEM6.
Conclusion:
A better understanding of the molecular mechanism underlying the function of KBTBD13 will
enable the identification of targets in NM-KBTBD13.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A62
Immune cell infiltration in obese brown adipose tissue
Susan van den Berg1, Andrea van Dam2, Tom Seijkens1, Gerdien van den Heuvel1, Linda Beckers1,
Myrthe den Toom1, Mariette Boon2, Patrick Rensen2, Menno de Winther1, Esther Lutgens1,3
1
Department of Medical Biochemistry, Subdivision of Experimental Vascular Biology, Academic Medical Centre,
2
University of Amsterdam . Department of Endocrinology, Leiden University Medical Center
3
Institute for Cardiovascular Prevention (IPEK), Ludwig Maximilian’s University, Munich, Germany
Introduction:
Obesity is associated with a variety of medical conditions such as type 2 diabetes and
cardiovascular diseases and is therefore responsible for high morbidity and mortality rates. A
current novel strategy to decrease obesity-associated morbidity and mortality is by increasing
energy expenditure by brown adipose tissue (BAT), which could counterbalance the excessive
energy stores in obesity. In contrast to the established contribution of the different immune cells
in white adipose tissue, the contribution of the immune system on generation, activity and
function of BAT is largely unknown.
Methods:
To assess the immune cell composition in BAT in diet-induced obesity, we studied C57Bl/6 mice
on a 45% high-fat diet (HFD) in a time course ranging from 1 day up to 18 weeks and analyzed
which, when and how immune cells infiltrate BAT. Results The absolute number of CD45+, F4/80+
and Ly6G+ immune cells in BAT did not increase upon diet feeding. However, the ratio between
pro-inflammatory (M1) and anti-inflammatory (M2) macrophage subsets increased already
within 1 week of HFD. Concordantly, Ccl2, Cxcl1 and Il-6 mRNA increased in BAT already after 3
days of HFD. In vitro experiments using a BAT cell line (T37i) revealed that stimulation with the
cytokines TNF and IFNγ decreased Ucp1 and Pgc1α mRNA expression, suggesting decreased
activity of the brown adipocytes. Furthermore, mRNA of Ccl2 and Ccl5 increased in T37i brown
adipocytes upon cytokine stimulation. Conclusion This study identified new targets in BAT to
modulate the activity of the tissue via the immune system.
This research was funded by the Rembrandt Institute of Cardiovascular Sciences.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A63-1 (first session)
Right ventricular hypertrophy and failure are associated with mitochondrial complex I
dysfunction and altered NADH kinetics
Rob C.I. Wüst1, Heder J de Vries1, Hans WM Niessen2 and Ger JM Stienen1
1
2
Department of Physiology and Pathology , Institute for Cardiovascular Research, VU University Medical Center,
Amsterdam.
Background:
In cardiac hypertrophy (CH) and heart failure (HF), alterations occur in mitochondrial enzyme
content and activities but the implications of these changes for mitochondrial function are
incompletely understood.
Methods:
Right ventricular CH or HF was induced in rats by a single injection of monocrotaline (40 and 60
mg.kg-1, respectively) and results were compared to control (CON). NADH and FAD
autofluorescence were recorded in thin intact cardiac trabeculae during transitions in stimulation
frequency, to assess mitochondrial complex I and complex II function, respectively. Oxygen
consumption, mitochondrial morphology and protein content were assessed by respirometry,
electron microscopy and western blotting.
Results:
NADH autofluorescence upon an increase in stimulation frequency showed a rapid decline
followed by a slow recovery. FAD autofluorescence followed a similar time course, but in
opposite direction. The amplitude of the early rapid change in NADH autofluorescence was
severely depressed in CH and HF compared to CON. The rapid changes in FAD autofluorescence
in CH and HF were reduced to a lesser extent. Complex I-coupled respiration showed a four-fold
reduction both in CH and HF; complex II-coupled respiration was depressed two-fold in HF.
Western blot analyses revealed a modest (~15%) reduction in complex I protein content in CH
and HF. Mitochondrial density was similar in all groups, but mitochondria were smaller and more
solitary in CH and HF compared to CON.
Conclusion:
These results indicate that the alterations in mitochondrial function observed in CH and HF can
be mainly attributed to complex I dysfunction.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A63-2
Atherosclerotic plaque area increases after orthopedic surgery in Apo E -/- mice
W.W. Fuijkschot; R. van der Linden; P.A.J. Krijnen; L.F.H. Theyse; Y.M. Smulders; H.W.M. Niessen
ICaR-VU, VU-University Medical center, Amsterdam
Introduction:
Observational studies show a peak incidence of cardiovascular events after major surgery. For
example, the risk for acute myocardial infarction is increased 25-fold early after total hip
replacement. The acuteness of this increased risk suggests plaque complications, which are
closely correlated with intra-plaque inflammation. We hypothesized that systemic inflammation
caused by major orthopedic surgery leads to plaque complications.
Methods:
ApoE -/- mice (n=48) were fed a western diet for 10 weeks after which the operation group
underwent midshaft femur osteotomy followed by realignment with a intramedullary K-wire, to
mimic major orthopedic surgery. Mice in both operation and control group were sacrificed 5 or
15 days post-surgery resp. post-saline injection. Paraffin embedded slides of the aortic root were
stained with Von Kossa, EVG, Toluidine blue (mast cells), MAC-3 (macrophages), Ly-6g
(granulocytes) and CD45 (lymphocytes).
Results:
Plaque area in operated animals versus controls had increased by 5,5 % compared after 5 days
(n.s.), and by 34% after 15 days (p=0.02). This plaque enhancement was especially related to an
increase (180 %) in the necrotic core, which was 99.0 x 103 μm2 [71.2-155] in controls, versus 257
x 103 μm2 [151-315] post-surgery (p=0.013). No significant differences were found in the density
of inflammatory cells.
Conclusions:
Atherosclerotic plaque area is enhanced after major orthopedic surgery in Apo E -/-mice, mainly
due to enlargement of the necrotic core, pointing towards increased plaque vulnerability.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A64
STORM superresolution microscopy to measure Z-disk and A-band width in human diaphragm
muscle
S.J.P Bogaards1, J. Girard1,2, J. de Winter1, M.W. Lawlor3, G.J.M. Stienen1,2, E.J. Peterman2, E.D.
van Deel1, C.A.C Ottenheijm1
1
Department of Physiology, Institute for Cardiovascular Research, VU University Medical Center Amsterdam,
2
3
Amsterdam , VU, Department of Physics and Astronomy, VU University Amsterdam, Amsterdam., Division of
Pediatric Pathology, Department of Pathology and Laboratory Medicine, Medical College of Wisconsin, Milwaukee,
Wisconsin, USA.
Introduction:
To date, no light microscopy techniques are available to image sarcomeres with nanoscale
precision. Therefore, we developed STochastic Optical Reconstruction Microscopy (STORM)
(spatial resolution ~20 nm) for imaging sarcomeres. To validate the technique, we compared
STORM data with electron microscopy (EM) images.
Subsequently we used STORM to study structural changes in diaphragm sarcomeres of
mechanically ventilated patients. Diaphragm weakness in ventilated patients augments ventilator
dependency, morbidity and long-term functional limitations. Animal studies indicate shortening
of diaphragm myofibrils after ventilation. We hypothesize that diaphragmatic weakness is partly
explained by structural changes in the sarcomere as a result of disuse during ventilation.
Methods:
Diaphragm muscle biopsies were obtained from mechanically ventilated patients (ICU; n=3) and
control subjects (CON; n=4). The tissue was processed for EM and single myofibrils, isolated for
STORM, were stained for myosin (type I and II), α-actinin and nebulin and imaged using STORM.
Z-disk and A-band widths were determined from both EM and STORM images.
Results:
Z-disk and A-band widths measured with STORM were in line with, though slightly larger then,
EM measured widths, which might be due to tissue shrinkage during EM processing.
Additionally, Z-disk width tended to be increased in ICU patients (STORM: 93±4 vs 106±3; EM:
92±3 vs 99±5 nm, CON vs ICU). In contrast, A-band width tended to be smaller in ICU than in
CON (STORM: 1624±10 vs 1553±53; EM: 1436±22 vs 1277±15 nm, CON vs ICU).
Conclusion:
STORM is a feasible light microscopy technique to localize individual sarcomeric proteins with
nanoscale precision.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A65
HEY2, a novel susceptibility gene for Brugada Syndrome, controls depolarization and
repolarization gradients in the RVOT and across the ventricular wall
S.Podliesna1, C.C. Veerman1, A.O.Verkerk1, R.Wolswinkel1, L.Beekman1, J.Barc1, M.Gessler2,
V.M.Christoffels1, A.A.Wilde1, C.A.Remme1, C.R. Bezzina1
1
2
Academic Medical Centre, Amsterdam., Theodor-Boveri-Institute, Biocenter, University of Wurzburg, Wurzburg,
Germany
Introduction:
In a recent GWAS, we identified a transcriptional repressor HEY2 as a novel susceptibility gene
for Brugada syndrome. Here, we investigated the effects of HEY2 on regional and transmural
depolarization and repolarization patterns.
Methods:
Male adult Hey2+/- mice and wild-type (WT) littermates were studied. Action potentials and
membrane currents were measured in cardiomyocytes isolated from RVOT and subepicardial
(EPI) and subendocardial (ENDO) regions of the right ventricle using the patch clamp technique.
Real-time RT-PCR was performed on EPI and ENDO tissues from the left ventricle.
Results:
In WT cardiomyocytes, action potential duration (APD90) and upstroke velocity (Vmax) were
significantly lower in WT-RVOT and WT-EPI compared to WT-ENDO. The voltage-clamp
experiments demonstrated higher transient outward K+ current (Ito) densities in WT-RVOT and
WT-EPI compared to WT-ENDO, while steady state K+ currents were similar. Kcnd2 mRNA
expression was significantly higher in WT-EPI compared to WT-ENDO. These regional and
transmural differences were abrogated in Hey2+/- hearts due to an increase in APD90 and Vmax
specifically in EPI and RVOT. Ito density and Kcnd2 mRNA expression was significantly decreased
in Hey2+/--EPI but was unaffected in Hey2+/--ENDO.
Conclusions:
HEY2 regulates APD90 and upstroke velocity in the RVOT and controls the transmural
depolarization and repolarization gradients across the ventricular wall.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A66
Commonalities and differences between dilated and peripartum cardiomyopathy; titin, fibrosis
and myomesin
Ilse AE Bollen1, Floor Bouwman1, Romy Fessl1, Elisabeth Ehler2, Diederik WD Kuster1, Jolanda van
der Velden1
1
2
Department of Physiology, VU University Medical Center, Amsterdam., The Randall Centre of Cell and Molecular
Biophysics King’s College London
Introduction:
Dilated cardiomyopathy (DCM) and peripartum cardiomyopathy (PPCM) share aspects of clinical
presentation such as ventricle dilation and reduced systolic function. However, because of
differences in underlying cause and disease progression, we hypothesized that the
pathomechanisms of the two cardiomyopathies are different. Therefore, we investigated titin
isoform composition, fibrosis and induction of fetal myomesin expression in end-stage PPCM and
DCM patients compared with non-failing donors.
Methods:
Left ventricular tissue was obtained via cardiac transplantation surgery of 6 PPCM, 11 DCM
patients and 13 non-failing donors. Titin isoform composition was determined by agarose gel
electrophoresis. Fibrosis and fetal myomesin expression were determined by
immunohistochemical staining.
Results:
Relative expression of the compliant titin isoform (N2BA/N2B ratio) was higher in PPCM and DCM
patients (0.82±0.15 and 0.88±0.11 respectively) compared with controls (0.57±0.06). The size of
N2BA titin was also increased in both cardiomyopathy groups. Increased fibrosis was seen in both
PPCM (4.5%) and DCM (5.5%) patients compared with donors (1.5%). Induction of the fetal
isoform of myomesin was observed only in DCM patients and not in PPCM patients and donors.
Conclusions:
Expression of the compliant titin isoforms and increased fibrosis was seen in both DCM and
PPCM patients. This could mean that titin isoform shift might compensate for the increased
fibrosis. Induction of fetal myomesin was specific for DCM patients, and provides a first indication
that remodelling might be different in the two cardiomyopathies.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A67
An alternative splice variant of myotonic dystrophy protein kinase induces loss of sarcomere
structure and cardiomyopathy
Amin Damanafshan1, Ies Elzenaar1, Benoit Samson-Couterie1, Meriem Bourajjaj3, Ingeborg van
der Made1, Maarten M. van den Hoogenhof1, Abdelaziz Beqqali1, Elisabeth Ehler2, Leon J. De
Windt3, Yigal M. Pinto1, Ralph J. van Oort1
1
2
Department of Experimental Cardiology, Academic Medical Center, Amsterdam. Cardiovascular Division, King’s
3
College, London, United Kingdom. Department of Cardiology, Maastricht University, Maastricht.
Introduction:
The pathology of heart failure is characterized by poorly contracting and dilated ventricles, which
is associated with lengthening of individual cardiomyocytes and loss of sarcomeres. Myocyte
enhancer factor-2 (MEF2) has previously been identified as important trigger for adverse
cardiomyocyte remodeling. Neonatal rat cardiomyocytes overexpressing MEF2 elongated and
lost sarcomeric structure. Here, we identified myotonic dystrophy protein kinase (DMPK) as a
direct MEF2 target gene and mechanistically link it with sarcomere disruption in structural
remodeling of the cardiomyocyte.
Methods and Results:
Microarray analysis revealed DMPK as a MEF2 target gene and siRNA mediated knockdown of
DMPK prevented MEF2-induced cardiomyocyte elongation and loss of sarcomeres. Interestingly,
RT-PCR analysis demonstrated a relative increase of a DMPK isoform (DMPK E) in failing mouse
hearts. Adenoviral overexpression of DMPK E, but not a control kinase dead mutant DMPK E
(K100A), resulted in severe loss of sarcomeres in vitro and transgenic mice overexpressing DMPK
E displayed disruption of sarcomere structure and cardiomyopathy in vivo. Moreover,
quantitative PCR analysis showed a decrease in mRNA levels for several sarcomeric genes after
overexpression of DMPK E. These genes are confirmed targets of the transcription factor serum
response factor (SRF) and DMPK is known to phosphorylate SRF. Therefore, we tested the effect
of DMPK on SRF activity and demonstrated that DMPK E acts as an inhibitor of SRF transcriptional
activity. Additionally, AdDMPK E infected neonatal rat cardiomyocytes showed a translocation of
SRF from the nucleus.
Conclusions: Our data indicate that MEF2 induces loss of sarcomeres mediated by DMPK.
Moreover, increased expression of DMPK E results in a decrease in sarcomeric gene expression,
which involves inhibition of SRF transcriptional activity. Furthermore, increased DMPK E
expression in vivo leads to loss of sarcomere structure and cardiomyopathy. Together, these
results assign a novel function to MEF2 and DMPK in adverse cardiomyocyte remodeling during
heart failure development.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A68
Effect of neurohormonal blockade on postural hemodynamic response in patients with chronic
heart failure
Anne-Sophie G.T. Bronzwaer*1,4, Lysander W.J. Bogert*1,4, Berend E. Westerhof 2,4,5, Mat J.A.P.
Daemen,3, Johannes J. van Lieshout1,2,4,6
*These authors contributed equally to the study
1
2
3
Departments of Internal Medicine, Anatomy, Embryology and Physiology, and Pathology, Academic Medical
4
Center, University of Amsterdam, Amsterdam. Laboratory for Clinical Cardiovascular Physiology, Center for Heart
5
6
Failure Research, Academic Medical Center, Amsterdam. Edwards Lifesciences BMEYE, Amsterdam. MRC/Arthritis
Research UK Centre for Musculoskeletal Ageing Research, School of Life Sciences, University of Nottingham Medical
School, Queen's Medical Centre, United Kingdom.
Introduction:
Sympathetic activation is the hallmark in patients with chronic heart failure (HF). Pharmacological
neurohormonal blockade aims to offset the adverse neurohormonal response, and has reduced
mortality and morbidity. In contrast to the pre-neurohormonal blockade era, HF patients
nowadays regularly suffer from postural hypotension which is considered attributable to medical
treatment. This study addressed the postural hemodynamic adaptation in HF and hypertensive
(HT) patients both treated with neurohormonal blockade.
Methods:
Blood pressure (BP; Finapres), stroke volume (SV; CO-trek), and total peripheral vascular
resistance (TPR) responses to postural stress were measured in 33 HF patients (10 New York
Heart Association (NYHA) I, 15 NYHA II and 8 NYHA III) and10 HT patients without heart failure
treated with neurohormonal blockade (ACE-inhibitors and/or beta-blockers), and 10 healthy agematched subjects (HS).
Results:
The initial BP upon standing decreased more in HF patients compared to HT patients and HS,
without full recovery at 60 sec of standing (systolic BP: -9±15% vs. 4±7% and 2±8% (p<0.001);
diastolic BP: -8±21% vs. 8±6% and 7±14% (p=0.003), respectively). The postural reduction in SV
was smallest in HF patients vs. HS and HT patients whereas in HF the physiological increase in TPR
was absent (-9±17% vs. 3±9% and 19±21% (p<0.001), respectively). The magnitude of the
cardiovascular responses to standing was inversely related to NYHA classification.
Conclusion:
In HF patients treated with neurohormonal blockade, the cardiovascular response to postural
stress is slow and relates to the cardiac condition rather than treatment.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A69
Selective phosphorylation of PKA targets after β-adrenergic receptor stimulation impairs
myofilament function in Mybpc3-targeted HCM mouse model
Aref Najafi1,4, Vasco Sequeira1, Michiel Helmes1, Ilse AE Bollen1, Max Goebel1, Jessica A Regan1,
Lucie Carrier2,3, Diederik WD Kuster1, Jolanda van der Velden1,4
1
2
Department of Physiology, VU University Medical Center, Amsterdam; Institute of Experimental Pharmacology and
3
Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Germany; DZHK
4
(German Centre for Cardiovascular Research), partner site Hamburg/Kiel/Lübeck, Germany; ICIN-Netherlands Heart
Institute, Utrecht.
Aims:
Hypertrophic cardiomyopathy (HCM) has been associated with reduced β-adrenergic (β-AR)
receptor signaling, leading downstream to a low protein kinase A (PKA) mediated
phosphorylation. It remained undefined whether all PKA targets will be affected similarly by
diminished β-AR signaling in HCM. We aimed to investigate the role of β-AR signaling on
regulating myofilament and calcium handling in an HCM mouse model harboring a gene
mutation (G>A transition on the last nucleotide of exon 6) in Mybpc3 encoding cardiac myosinbinding protein C (cMyBP-C).
Methods and Results:
Cardiomyocyte contractile properties as well as phosphorylation state were measured in left
ventricular (LV) permeabilized and intact cardiomyocytes isolated from heterozygous (HET) or
homozygous (KI) Mybpc3-targeted knock-in mice. A significant higher myofilament Ca2+sensitivity and passive tension was detected in KI mice, which were normalized after PKA
treatment. Loaded intact cardiomyocyte force-sarcomere length (SL) relation was impaired in
both HET and KI mice, suggesting a reduced length-dependent activation. Unloaded
cardiomyocyte function revealed an impaired myofilament contractile response to isoprenaline
(ISO) in KI, while the calcium handling response to ISO was maintained. This disparity was
explained by an attenuated increase in cardiac troponin I (cTnI) phosphorylation in KI, while the
increase in phospholamban (PLN) phosphorylation was maintained to wild-type (WT) values.
Conclusions:
These data provide evidence that in the KI HCM mouse model β-AR stimulation leads to
preferential PKA phosphorylation of PLN over cTnI, resulting in an impaired inotropic and
lusitropic response.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A70
Trial protocol: Healthy Ageing Through Internet Counselling in the Elderly – the HATICE
randomized controlled trial for the prevention of cardiovascular disease, cognitive impairment
and dementia
Susan Jongstra1, Cathrien Beishuizen1, Eric Moll van Charante1, Miia Kivipelto2, Sandrine
Andrieu3, Hilkka Soininen4, Carol Brayne5, Yannick Meiller6, Bram van der Groep7, Pim van Gool1,
Edo Richard1,8
1
2
3
Academic Medical Center, Amsterdam; Karolinska Institutet, Stockholm; University of Toulouse, Toulouse;
5
6
7
University of Eastern Finland, Helsinki; University of Cambridge, Cambridge; Novapten, Paris; Vital Health
8
Software, Ede; Radboud university medical centre, Nijmegen
4
Introduction:
Cardiovascular disease(CVD) and dementia share a number of risk factors including hypertension,
hypercholesterolemia, smoking, obesity, diabetes and physical inactivity. The rise of E-health has
led to great opportunities for large-scale delivery of prevention programs encouraging selfmanagement, also for older people.
Aim:
To investigate whether an interactive internet intervention to optimise self-management of
cardiovascular risk factors in older persons can improve the cardiovascular risk profile and reduce
the risk of CVD, cognitive decline and dementia.
Methods:
Design HATICE is a multi-national, multi-centre, investigator initiated, open-label blinded
endpoint, randomised controlled trial with 18-months follow-up. Recruitment of 4250 older
people (>=65 years) with increased risk of CVD will take place in Finland, France and the
Netherlands. Participants randomized to the intervention condition will have access to an
interactive platform, with distant support by a coach.
The primary outcome is a composite score based on the average z-score of the difference
between baseline and 18 months follow-up values of systolic blood pressure, low-densitylipoprotein and BMI. Main secondary outcomes include effect on the individual components of
the primary outcome, effect on number of self-reported risk factors, difference in estimated 10year cardiovascular disease risk based on the Framingham score, incident CVD, mortality, CAIDE
dementia risk-score, cognitive decline, mood and cost-effectiveness.
Conclusion:
The platform supported by a coach and the stimulation of self-management are expected to be
the key factors in changing older peoples lifestyle and therefore improving the cardiovascular risk
profile and reduce the risk of CVD, cognitive decline and dementia.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A71
Exploration of the function of Class I and Class IIa HDACs in experimental Atrial Fibrillation
Deli Zhang1, Xu Hu1, Marit Wiersma1, Femke Hoogstra-Berends1, Qiang Zhuang2, Miguel A.
Esteban2, Gunseli Cubukcuoglu Deniz3,4, Serkan Durdu3,4,5, Ruchan Akar4 Robert H. Henning1,
Bianca J.J.M. Brundel1
1 Department of Clinical Pharmacy & Pharmacology, University Medical Center Groningen, University of Groningen.
2 Key Laboratory of Regenerative Biology, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of
Sciences, Guangzhou, China. 3 Ankara University Biotechnology Institute, Ankara, Turkey 4 Ankara University Stem
Cell Institute, Ankara, Turkey, 5 Department of Cardiovascular Surgery, Ankara University School of Medicine, Turkey
Background: Atrial Fibrillation (AF), is associated with altered gene expression resulting in
functional loss and AF progression. Recent research showed class I and IIa histone deacetylases
(HDACs) to regulate pathological fetal gene expression resulting in hypertrophy and contractile
dysfunction. Whether class I and IIa HDACs are involved in AF progression is unknown. Therefore,
we investigated their role in tachypacing-induced contractile dysfunction and pathological fetal
gene expression in experimental models for AF and in clinical AF.
Methods and Results: Tachypacing (TP) of HL-1 atrial cardiomyocytes resulted in CaT loss.
Overexpression of HDAC class I HDAC1 and HDAC3 did not protect from CaT loss in tachypaced
and control non-paced cardiomyocytes. Whereas overexpression of Class IIa HDACs, HDAC5 and
HDAC7, protected against TP-induced CaT loss, in constrast to HDAC4 and HDAC9, which were
not protective. Notably, cardiomyocytes overexpressing a dominant negative HDAC5 or HDAC7
mutant, which bears a mutation in the myosin enhancer factor 2 (MEF2) binding domain, were
not protected against loss of CaT. Furthermore, TP increased phosphorylation of HDAC5,
promoted its translocation from the nucleus to cytoplasm, and consequently increased MEF2related fetal gene expression (β-MHC, BNP). In line with these experimental findings, patients
with AF showed a significant increase in both phosphorylated HDAC5 levels and fetal gene
expression (β-MHC, BNP).
Conclusion: Overexpression of HDAC5 and HDAC7 protects against tachypacing-induced CaT
reduction in HL-1 cardiomyocytes. Furthermore, tachypacing induces HDAC5 phosphorylation
and its nuclear export, resulting in MEF2 regulated fetal gene expression. These features are also
present in AF patients. As HDAC5 is abundantly expressed in the heart, in contrast to HDAC7,
HDAC5 is a promising therapeutic target in clinical AF to attenuate pathological gene expression,
contractile dysfunction and progression of the disease.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A72
Title: A specified atrioventricular conduction system in the American alligator (Alligator
mississippiensis) heart.
Bjarke Jensen1*, Bastiaan J Boukens2*, Dane A Crossley II3, Ruth Elsey4, David Sedmera5, Vincent
M Christoffels1
1
Department of Anatomy, Embryology, and Physiology, Heart Failure Research Center, Academic Medical Center,
University of Amsterdam, Amsterdam.
2
Department of Biomedical Engineering, Washington University, St. Louis, MO, USA
3
Department of Biological Sciences, University of North Texas, Denton, TX 76210, USA
4
Rockefeller Wildlife Refuge, Louisiana Department of Wildlife and Fisheries, Grand Chenier, LA, 70643, USA
5
Institute of Anatomy, First Medical Faculty, Charles University, Prague, Czech Republic
*these authors contributed equally
Introduction:
Mammals and birds have a specialized atrioventricular conduction pathway through their
ventricular septum enabling rapid activation of the ventricles. The conduction system is seen as
necessary to support the high heart rates required for warm-bloodedness (endothermy).
Mammals and birds evolved independently from a reptilian ancestor, and it remains unresolved
whether a distinct atrioventricular conduction pathway is present in reptiles that possess a full
ventricular septum. Tbx3 and Tbx5 are conserved T-box transcription factors required for
development and function of the mammalian atrioventricular conduction pathway.
Methods and Results:
In situ hybridizations showed Tbx3 in juvenile alligators expressed in a population of myocardium
from the atrioventricular junction to the dorso-cranial part of the ventricular septum. Insulating
connective tissue interrupted the atrioventricular myocardial connection ventrally, but not
dorsally. The Tbx3-positive myocardium was also rich in Tbx5 expression. The septal part of the
Tbx3-positive domain did not express atrial myosin heavy chain, indicating its ventricular identity.
Gap junction protein 5 (Cx40), marker of the ventricular conduction system of mammal and bird
hearts, was expressed throughout the highly trabeculated ventricle. Optical mapping of the
ventricular action potentials revealed early activation of the dorsal ventricular base. Surgical cuts
through the Tbx3-positive domain resulted in complete AV block (N=3). Conversely, ventral cuts
did not interfere with AV conduction (N=3) consistent with the extent of the insulating plane.
Conclusions:
The American alligator has a specialized atrioventricular conduction pathway and, in contrast to
what is commonly thought, the specialized atrioventricular conduction pathway can evolve in the
absence of high heart rates.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A73
Muscle fiber contractile measurements in nemaline myopathy patients affected by ACTA1
mutations
B. Joureau1, J.M. de Winter1, Stefan Conijn1, Michaela Yuen3,4, A.H.Beggs2, N. F. Clarke3,4, G. J. M.
Stienen1,5, C.A.C Ottenheijm1
1
2
Department of Physiology, VU University Medical Center Amsterdam. Division of Genetics and Program in Genomics, The
3
Manton Center for Orphan Disease Research, Children's Hospital Boston, Harvard Medical School, Boston, MA, USA. Institute for
4
Neuroscience and Muscle Research, The Children’s Hospital at Westmead, NSW, 2145 Discipline of Pediatrics and Child Health,
5
University of Sydney, Australia VU University, Dept. of Physics and Astronomy, Amsterdam, The Netherlands.
Background: Nemaline myopathy (NM) is a heterogeneous congenital non-dystrophic myopathy,
characterized by generalized muscle weakness and the presence of nemaline rods in the skeletal
muscle. Mutations in ACTA1 account for ~25% of NM cases and up to 50% of the severe forms.
ACTA1 encodes skeletal muscle α-actin, the main component of the sarcomeric thin filament. We
aimed to investigate the contractile performance of muscle fibers isolated from NM patients with
mutations in ACTA1 (NM-ACTA1).
Methods: Biopsies from controls (CTRL, n=4) and NM-ACTA1 (n=7) patients were compared.
Permeabilized muscle fibers were dissected and mounted between a force transducer and length
motor and activated by exposure to solutions containing saturating [Ca2+] (pCa 4.5). Maximal
tension, cross bridge cycling kinetics (ktr), calcium-sensitivity of force (pCa50, fibers were bathed
in solutions with pCa’s ranging from 4.5 to 9.0), Hill coefficient (nH), and the active stiffness were
determined at a sarcomere length of 2.5 µm. Furthermore, maximal tension was determined at
incremental sarcomere lengths (range 2.0–3.5 µm) to obtain the force-sarcomere length
relationship (F-SL) in NM-ACTA1 and CTRL patients. The tensions at incremental sarcomere
lengths were fit using a 2nd order polynomial, from which the sarcomere length at maximal force
(SLopt) and the sarcomere length at 50% of the maximal force (SL50) was derived.
Results: Contractile data are summarized
in the table below:
Maximal Tension (mN/mm2)
Ktr (s-1)
The Calcium Sensitivity of force generation (pCa50)
Hill coefficient (nH)
Active Stiffness (mN/mm2)
Tension/Stiffness ratio
SLopt (µm)
SL50 (µm)
CTRL
ACTA1
153.37±8.07
9.55±0.7
6.08±0.007
3.43±0.15
74.57±7.01
2.11±0.22
2.54±0.03
3.58±0.02
81.39±17.89*
4.86±0.90*
6.08±0.032
2.71±0.09*
45.97±8.65
1.72±0.17
2.59±0.043
3.47±0.03*
Mean±SEM; * p<0.05 CTRL vs. ACTA1
Conclusion: Single muscle fibers of NM-ACTA1 patients show contractile weakness, and changes
in cross bridge cycling kinetics. These findings suggest that, in NM-ACTA1 patients, muscle
weakness is at least partly caused by impaired functioning of the myofilaments.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A74
Non-coding RNAs in cardiac aging
D. Julia Trembinski1, Denise Berghäuser1, Patrick Hofmann,1 Stefanie Dimmeler1, Reinier A.
Boon1,2
1
Institute for Cardiovascular Regeneration, Goethe University Frankfurt, Frankfurt am Main, Germany
Department of Physiology, VU University Medical Center, Amsterdam.
2
Introduction:
Aging is the major risk factor for developing cardiovascular complications like endothelial
dysfunction, aneurysm formation, acute myocardial infarction and heart failure. The mechanisms
involved in cardiovascular aging are poorly understood, but non-coding RNAs, including
microRNAs, have emerged as key biological regulators.
Methods/Results:
We have described the crucial role for miR-34a in cardiac aging, which regulates cardiomyocyte
apoptosis and telomere length. Current studies in the laboratory focus on determining the role of
long non-coding RNAs (lncRNAs) in aging of the cardiovascular system and how these affect
organ homeostasis and cellular metabolism. We first showed that many lncRNAs (>200 nt) are
expressed in endothelium and that the lncRNA MALAT1 is required for endothelial proliferation
in vitro and in vivo. Several lncRNAs are also regulated during cardiac ageing. We selected 74 ageregulated cardiomyocyte-enriched lncRNAs for an siRNA-based apoptosis screen. This assay
showed that several lncRNAs regulate apoptosis of cardiomyocytes, in particular the lncRNA we
termed lncCM1. Silencing of the nuclear -localized lncCM1 with siRNA or RNase H-inducing LNA
GapmeRs potently induced apoptosis in human and mouse cardiomyocytes. Furthermore,
lncCM1 is strongly downregulated upon acute myocardial infarction in mice.
Conclusion:
Together, these data indicate that lncCM1 induction or gene transfer may ameliorate
cardiomyocyte apoptosis after acute myocardial infarction, especially in the elderly.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A75
Electrophysiological characteristics of iPSC-derived cardiomyocytes from Brugada syndrome
patients without SCN5A mutations
Christiaan C. Veerman1, Isabella Mengarelli1, Kaomei Guan2, Arthur A. M. Wilde1, Arie O. Verkerk3
Connie R. Bezzina1
1: Heart Center, Academic Medical Center, University of Amsterdam, Amsterdam.
2: Department of Cardiology and Pneumology, Heart Centre, Georg-August-University of Göttingen, Göttingen,
Germany
3: Department of Anatomy, Embryology and Physiology, Academic Medical Center, University of Amsterdam,
Amsterdam.
Introduction:
The Brugada syndrome (BrS) is a rare cardiac rhythm disorder associated with sudden cardiac
death and characteristic ST-elevation in the right-precordial ECG leads. In ~20 % of patients, a
loss-of-function mutation in the sodium channel gene SCN5A is identified, while other gene
mutations are found in sporadic cases. In the remaining BrS patients the genetic cause, as well as
the pathophysiological mechanism, remain elusive. We here characterized the
electrophysiological properties of cardiomyocytes derived by induced pluripotent stem cell
technology (iPSC-CMs) from three BrS patients who were negative for coding region mutations in
SCN5A, and compared them with those of iPSC-CMs derived from two control individuals.. We
also compared them to iPSC-CMs from a carrier of the SCN5A-1795insD mutation, which we
previously showed could recapitulate the decreased action potential upstroke velocity caused by
the SCN5A mutation, and could thereby serve as a control for the typical BrS-causing SCN5A
mutation.
Methods and results:
Action potential characteristics in all groups were determined in single iPSC-CMs by the
perforated patch clamp technique, to which in silico IK1 was added through dynamic clamp.
Action potential measurements revealed no abnormalities in iPSC-CMs from the three BrS
patients, as compared to the two controls. In contrast, iPSC-CMs harbouring the 1795insD
mutation displayed reduced action potential upstroke velocity, as compared to all control and
BrS iPSC-CMs. In agreement with unaltered action potential upstroke velocity, sodium current
density was not different in iPSC-CMS from the three BrS patients compared to the controls. A
marked reduction was observed in sodium current density in iPSC-CMs carrying mutation
1795insD.
Conclusion:
hiPSC-CMS from SCN5A-mutation negative BrS patients do not exhibit marked cellular
electrophysiological abnormalities that could explain the disease. This indicates that baseline ion
channel defects, in particular in the cardiac sodium channel, are not a prerequisite for the
pathogenesis of BrS.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A76
Electrical and structural abnormalities in homozygous Scn5a-1798insD mutant embryos
Gerard A. Marchal, Rianne Wolswinkel, Bastiaan J. Boukens, Connie R. Bezzina, Carol Ann Remme
Department of Experimental Cardiology and Department of Anatomy, Embryology and Physiology, AMC Amsterdam,
The Netherlands
Introduction:
Sodium channels are essential for cardiac excitability and ensure proper conduction throughout
the myocardium. Evidence suggests that sodium channels not only regulate electrical properties
of cardiomyocytes, but may also affect cardiac structure and function. We here investigated the
role of the cardiac sodium channel Scn5a in cardiac development in mice carrying the Scn5a1798insD mutation.
Methods and results:
Scn5a-1798insD homozygous mice were generated by crossing heterozygous Scn5a-1798insD
(HET) male and female mice. Scn5a-1798insD homozygous mice were not viable. Scn5a-1798insD
homozygous embryos (HOMO) were observed at the expected Mendelian ratio at embryonic day
(ED) 8.5-9.5, but were found at a low frequency at ED10.5 and were absent from ED11.5
onwards. On visual inspection, HOMO embryos at ED9.5 (n=5) displayed increased AV-delay as
compared to wild-type (WT) in addition to gross structural alterations. At ED10.5, HOMO
embryos (n=7) appeared smaller in size (sometimes necrotic), and displayed both structural and
functional abnormalities. On optical mapping (ED10.5), WT and HET embryos (n=3) showed
organized, fast ventricular activation, whereas HOMO embryos (n=3) displayed chaotic
ventricular activation patterns. At ED9.5, electrical properties appeared similar between WT,
HET, and HOMO embryos.
Conclusions:
Homozygous Scn5a-1798insD embryos display structural abnormalities at early embryonic age,
when cardiac sodium channels may not yet be essential for cardiac electrical function. These
findings indicate that intact sodium channel function is required for normal embryonic
development of the heart.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A77
Novel HSP-inducing Compounds to Restore Cardiomyocyte Function in Experimental Atrial
Fibrillation
D.M.S. van Marion (UMCG, Groningen), E. Lanters (EMC, Rotterdam), N. de Groot (EMC,
Rotterdam), F. Hoogstra-Berends (UMCG, Groningen), R.H. Henning (UMCG, Groningen), B.J.J.M.
Brundel (UMCG Groningen, VUmc, Amsterdam)
Purpose:
Atrial Fibrillation (AF) is the most common clinical tachyarrhythmia associated with significant
morbidity and mortality. AF is a persistent disease, caused by a progressive, often age-related,
derailment of proteostasis resulting in structural remodeling and contractile dysfunction of
cardiomyocytes. It has been widely acknowledged that the progressive nature of the disease
hampers the effective functional conversion to sinus rhythm in patients and explains the limited
efficacy of current drug therapies. Previously, we identified that inducers of heat shock proteins
(HSPs), such as geranylgeranylacetone (GGA) by inducing Heat Shock Proteins (HSPs) expression,
suppresses derailment of proteostasis and remodeling of cardiomyocytes. As a result, GGA
attenuates the AF substrate in cellular, Drosophila melanogaster and animal experimental
models. Clinical application of GGA is however hampered by the high dosage needed, because of
its high LogP value. Therefore, the aim of the current study was to identify novel HSP-inducing
compounds, which protect against AF remodeling.
Methods and results:
We synthesized 83 GGA-derivatives and explored their action (at 10µM) in HL-1 cardiomyocytes
pretreated with a mild heat shock to activate heat shock factor-1 (HSF-1) (430C 10 min, 10 min
recovery 370C). We identified 30 GGA-derivatives that significantly induced HSP70 expression
and other HSF-1 related HSPs, including HSP25, HSP90, HSP40, but not Grp78 (not HSF-1 related).
The magnitude of induction was comparable or higher compared to GGA. Next, HL-1
cardiomyocytes were pretreated with the most potent HSP70-inducing GGA-derivatives (n=13)
for 8 hrs, followed by 8 hrs tachypacing (5 Hz) or normal pacing (1 Hz) and contractile function
was determined by measuring calcium transients (CaT). Tachypacing significantly reduced the
amplitude of CaT, and 6 GGA-derivatives revealed a significant protective effect against CaT loss,
which was improved or comparable to GGA.
Conclusion:
We identified novel GGA derivatives with improved HSP-inducing, cardioprotective and
pharmaco-chemical properties compared to GGA. Ultimately, these HSP-inducing compounds
may prevent expansion of the structural remodeling during AF, resulting in improved outcome of
cardioversion and/or delay in progression towards permanent AF.
Keywords:
Atrial fibrillation, HSP inducers, cardiomyocytes, contractility
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A78-1 (first session)
Homozygous Mutations in GNB5 and SYDE2 in patients with autosomal recessive bradycardia
Elisabeth M Lodder(1), Eline A. Nannenberg(1), Leander Beekman(1), Mischa Klerk(1), Najim
Larouchi(1), Karin Y. van Spaendonck(1), Ilham Ratbi(2), Connie R. Bezzina(1)
1) Amsterdam Medical Center, Amsterdam, the Netherlands; 2) Université Mohammed V Souissi, Rabat, Morocco
Background & Aim:
We recently identified a small family with severe sick sinus syndrome which was inherited in a
autosomal recessive manner: the parents (of Moroccan descent) were unaffected, both the index
patient and his affected sister needed pacemaker implantation as a result of severe sinus node
dysfunction directly after birth and had mild mental retardation. We here aimed to identify the
causal mutation for the phenotype using whole exome sequencing.
Methods and results:
We performed whole exome sequencing in all four family members, and filtered for variants that
followed the recessive mode of inheritance and were not present in any of the known variant
databases (e.g. EVS, 1000G, GoNL, G69 & in house data) with a minor allele frequency of more
than 1%. Two missense variants were identified: GNB5-p.S81L and SYDE2-p.S34F, both were
validated by Sanger sequencing.
GNB5 encodes a G-protein beta subunit (Gbeta5) and is known to bind to and stabilise RGS6
(regulator of G-protein signalling 6). The RGS6/Gbeta5 hybrid stimulates the de-activation of the
IKACh after activation by the muscarinic M2-receptor. Gnb5-/- mice exhibit mild bradycardia and
motor learning deficits. SYDE2 (synapse defective 1, Rho GTPase, homolog 2 (C. elegans)) is a
gene with unknown function; it is thought to encode a GTPase based on homology. As the deactivation of the M2-receptor signalling depends on the hydrolysation of the GTP-bound G-alpha
subunit to the GDP bound state, it is very well possible that SYDE2 is involved in the modulation
of this process.
Conclusions:
In summary, we here provide novel genetic data linking homozygous mutations in GNB5 and
SYDE2 to severe bradycardia in human. Based on the severe phenotype there is a possible
synergistic effect between both mutations, functional studies will be needed to resolve their
independent effects.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A78-2 (second session)
CD40-Filamin A interactions are required for translocation of CD40 to lipid rafts in endothelial
cells and for endothelial cell activation
Claudia M. van Tiel1, Patrick Burger1, Noam Zelcer1, Peter L. Hordijk2, Jaap D. van Buul2, Esther
Lutgens1,3
1
Department of Medical Biochemistry, subdivision Experimental Vascular Biology, Academic Medical Center,
University of Amsterdam, Amsterdam, The Netherlands
2
Department of Molecular Cell Biology, Sanquin Research and Landsteiner Laboratory, Academic Medical Center,
University of Amsterdam, The Netherlands
3
Institute for Cardiovascular Prevention (IPEK), Ludwig Maximilians University, Munich, Germany
Background and Aim:
CD40 is a member of the co-stimulatory tumor necrosis factor receptor superfamily that is not
only constitutively expressed on professional antigen presenting cells like macrophages, dendritic
cells and B-cells, but also on endothelial cells. Interactions between CD40 and its ligand, CD40
ligand (CD40L) are crucial for proper immune cell activation. Activation of CD40 signaling plays a
role in chronic diseases such as rheumatoid arthritis, inflammatory bowel disease and
atherosclerosis. However, since CD40-CD40L interactions modulate immune reactions as well as
thrombosis, blocking CD40(L) can have adverse side-effects. Therefore, in search for new
therapeutic targets, we aimed to identify new CD40-binding partners.
Methods and Results:
We created a cDNA library of murine aortas containing atherosclerotic plaques at various stages
and performed a yeast-two-hybrid with the C-terminal domain of CD40 as bait. We identified
filamin A as a novel CD40 binding partner in atherosclerosis, which was confirmed by coimmunoprecipitation. Accordingly, using confocal microscopy we showed that, upon activation of
CD40, filamin A was recruited to CD40 in endothelial cells. Site directed mutagenesis revealed
that Filamin A binds to the intracellular domain of CD40, near the transmembrane domain, at a
site distinct from the tumor necrosis receptor associated factor (TRAF) binding sites. In
endothelial cells, CD40-signaling is dependent on intact lipid rafts. Interestingly, when we
knocked-down filamin in endothelial cells using siRNAs, the translocation of CD40 to lipid rafts
upon activation of CD40 signaling was inhibited, resulting in repression of CD40-mediated Akt
signaling and subsequent inhibition of VCAM-1 and CCL-2, but not of CD40-mediated JNK
signaling.
Conclusions:
We show that CD40 interacts with filamin A in endothelial cells. This interaction is crucial for
translocation of CD40 to the lipid rafts and CD40-mediated activation of the Akt pathway. The
reduced upregulation of VCAM-1 and CCL-2 makes CD40-filamin interactions an interesting
target for treatment of atherosclerosis.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A79
p47phox-dependent reactive oxygen species stimulate nuclear translocation of the transcription
factor FOXO1 in ischemia-challenged H9C2 rat cardiomyocytes
E.N. ter Horst1,5,6,7, N.E. Hahn1,6, D. Geerts8, R.J.P. Musters1,6, W.J. Paulus2,6, A.M. van der Laan5,
J.J. Piek5, A.C. van Rossum3,6, C. Meischl1,6, H.W.M. Niessen1,4,6, P.A.J. Krijnen1,6
1
2
3
4
Department of Pathology, Physiology, Cardiology and Cardiac Surgery, VU University Medical Centre, Amsterdam.
5
Department of Cardiology, Academic Medical Centre, University of Amsterdam.
6
ICaR-VU, Institute of Cardiovascular Research, VU University Medical Centre, Amsterdam.
7
Interuniversitary Cardiology Institute of the Netherlands, Utrecht.
8
Department of Pediatric Oncology/Hematology, Erasmus University Medical Centre, Rotterdam.
Introduction:
Reactive oxygen species (ROS) control forkhead box O (FOXO) transcription factors by influencing
their nuclear translocation. However, knowledge of the cellular source(s) of ROS involved herein
remains scarce. Recently, we have shown in H9C2 rat cardiomyoblasts that activation of ROSproducing NADPH-oxidases (NOX) at the nuclear pore under ischemia is p47phox-dependent. This
localizes NOX perfectly to affect translocation of nuclear proteins, including transcription factors.
In the current study, we analyzed the involvement of p47phox-dependent ROS production in
FOXO1 nuclear translocation during ischemia in H9C2 cells.
Methods:
The effect of ischemia on H9C2 cells was mimicked by metabolic inhibition. Nuclear translocation
of FOXO1 was determined via quantitative digital imaging fluorescence and Western blot
analysis. Subsequently, the effect of inhibiting p47phox-dependent ROS production via short
hairpin RNA (shRNA) transfection on FOXO1 translocation was analyzed by digital-imaging
microscopy.
Results:
Ischemia induced a significant translocation of FOXO1 into the nucleus of H9C2 cells. Transfection
with p47phox-shRNA successfully inhibited p47phox expression and resulted in a reduced nuclear
ROS generation and an inhibition of FOXO1 nuclear translocation.
Conclusions:
Our data show for the first time that ischemia-induced nuclear import of FOXO1 in H9C2 rat
cardiomyoblasts is critically dependent on p47phox-mediated ROS production.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A80
Correcting QRS duration for Left Ventricular dimension improves current patient selection for
Cardiac Resynchronization Therapy
A Zweerink; L Wu; GJ de Roest; R Nijveldt; CC de Cock; AC van Rossum; CP Allaart.
Department of Cardiology, and Institute for Cardiovascular Research (ICaR-VU), VUmc, Amsterdam.
Introduction: Current patient selection for Cardiac Resynchronization Therapy (CRT) is hampered
by high rates of non-response (~30%). The aim of this study is to improve patient selection by
correcting the selection criterion, QRS duration, for Left Ventricular (LV) dimension.
Methods: Thirty-six CRT candidates (age 65 ± 9 years, 23 men) underwent CMR cine- and
Delayed Contrast Enhanced (DCE)-imaging to provide detailed information on Left Ventricular
(LV) morphology, function and myocardial tissue characterization. Baseline QRS duration was
corrected for Left Ventricular End Diastolic Volume (LVEDV). All patients underwent invasive
pressure-volume loop assessment at baseline and during biventricular pacing to determine acute
pump function (SW) improvement during CRT.
Results: At baseline LVEDV showed large variation (288 ± 83) and was positively related to QRS
duration (R=0.39; P=0.019). During biventricular pacing, SW improved by +37% (P<0.001).
Despite prolonged QRS duration, high LVEDV at baseline was inversely related to SW
improvement during CRT (R=-0.34; P=0.042), see figure 1A. Baseline QRS duration was found to
be unrelated to SW changes during CRT (R=0.18; P=0.300), see figure 1B. LVEDV corrected QRS
duration however, proved to be significantly related to acute SW response (R=0.46; P=0.005), see
figure 1C. Conclusions: Although LV dilatation is accompanied by QRS prolongation, it is inversely
related to acute pump function improvement. Correcting QRS duration for LV dilatation might
result in improved patient selection for CRT.
Figure 1 shows scatter plots illustrating the association between: (A) LVEDV and acute SW changes during CRT; (B)
QRS duration and acute SW changes during CRT; (C) LVEDV corrected QRS duration and acute SW changes during
CRT. ICMP = ischemic cardiomyopathy; NICMP = non-ischemic cardiomyopathy
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A81
The PARP1 inhibitor ABT888 accelerates recovery from contractile dysfunction in experimental
Atrial Fibrillation
Xu Hu1, Deli Zhang1, Marit Wiersma1, Denise Van Marion1, Femke Hoogstra-Berends1, Robert H.
Henning1, Bianca J.J.M. Brundel1,2
1) Department of Clinical Pharmacy & Pharmacology, University Medical Center Groningen, University of Groningen.
2)Department of Physiology, Institute for Cardiovascular Research, VU University of Medical Center, Amsterdam.
Introduction:
Atrial Fibrillation (AF) is the most common persistent clinical tachyarrhythmia. It is associated
with metabolic and structural remodeling, resulting in contractile dysfunction. Activation of
nuclear enzyme poly (ADP-ribose) polymerase (PARP) is implicated in metabolic remodeling in
various forms of cardiomyopathies. Our previous experimental studies showed that AF-induced
metabolic remodeling and contractile dysfunction can be prevented by pretreatment with a
PARP1 inhibitor ABT888. Since most patients are diagnosed with AF when the metabolic
remodeling is already presented, it is clinically highly relevant to test whether ABT888 can
reverse remodeling. Therefore, the PARP1 inhibitor ABT888 was tested in the reversal of
contractile dysfunction in tachypaced HL-1 atrial cardiomyocytes.
Methods and Results:
HL-1 cardiomyocytes were tachypaced (TP, 5Hz) for 10hrs to induce calcium transient (CaT) loss,
followed by treatment with 40 µM ABT888 for 24hrs. TP induced significant CaT loss in
cardiomyocytes , which did not recover during a 24hrs recovery period. Interestingly,
cardiomyocytes treated with ABT888 post TP revealed significant recovery from CaT loss.
Conclusions:
Our results suggest that the PARP1 inhibitor ABT888 improved recovery from tachypacinginduced contractile dysfunction, indicating that PARP1 is involved in AF progression and
represents a possible therapeutic target for treatment. Further research is directed at elucidating
underlying mechanisms for PARP1 in AF progression and the efficiency of novel inhibitors will be
explored.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A82
CARDIAC SUBSTRATE STIFFNESS AFFECTS SARCOMERE FUNCTION
Elza D.van Deel, Max Goebel, Kim Kardux, Erik Valent, Geerten P. van Niew Amerongen Jolanda
van der Velden
Laboratory for Physiology, VU University Medical Centre, Amsterdam, the Netherlands
Introduction:
Cardiac stiffening via altered extracellular matrix (ECM) properties is predominantly described as
a passive inhibitor of cardiac function. Although ECM properties are sensed inside the cell
through costameres that link the ECM directly with the sarcomeres, little is known about the
effects of ECM stiffness on sarcomere proteins. We hypothesize that cardiac stiffening actively
impairs cardiomyocyte function by affecting sarcomere function and Ca2+ signalling.
Methods:
To explore the effects of matrix stiffness on sarcomere function, cardiomyocytes isolated from
adult rats were plated on laminin-coated polyacrlamide substrates of defined stiffness (8, 15, 50
and 100 kPa and glass). 24h after plating the cardiomyocytes were detached from their
substrates and sarcomere shortening, calcium dynamics and phospholamban (PLB)
phosphorylation were measured.
Results:
Relative sarcomere shortening as well as shortening and relaxation velocity were optimal in cells
from substrates which closely resembled the healthy heart (15 kPa), while these parameters
were impaired on more elastic (8kPa) and stiffer (50-100 kPa and glass) substrates. These findings
were not solely explained by sarcomere changes, as Ca2+ release velocity and re-uptake rate were
highest on the substrate that resembled the healthy heart, but altered in cells plated on the
stiffer substrates. Preliminary analysis reveal that the latter was at least partly explained by
reduced PLB phosphorylation in cells cultured on the stiffer substrates.
Conclusions:
Cardiomyocyte sarcomere function as well as calcium dynamics are directly affected by stiffness
of the extracellular environment and optimal under physiological conditions.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A83
Cyclophilin D ablation protects the heart against infarction and is associated with increased
end-ischemic mitochondrial hexokinase activity
Rianne Nederlof, Mark A.M. van den Elshout, Anneke Koeman, Nina C. Weber, Markus W.
Hollmann, Coert J. Zuurbier.
Laboratory of Experimental Intensive Care and Anesthesiology, Department of Anesthesiology, Academic Medical
Center, Meibergdreef 9, 1105 AZ Amsterdam.
Introduction: Both the absence of cyclophilin D (CypD) and the presence of hexokinase II (HKII)
bound to mitochondria (mtHKII) protects the heart against ischemia/reperfusion (I/R) injury.
Although research in non-cardiac cells suggests that CypD determines HKII-to-mitochondria
association, such information is currently lacking for the heart. In the present work we examine
whether CypD affects mitochondrial-hexokinase binding in normoxic, ischemic and
preconditioned isolated mouse hearts.
Methods: All experiments were performed in isolated wild type (WT) and CypD-/- mouse hearts
perfused with glucose-only or substrate-enriched (glutamine, pyruvate, lactate) perfusate,
employing I/R and ischemic preconditioning (IPC) interventions, percoll-isolated mitochondria
and protein analysis by Western blot and spectrophotometric enzyme activities techniques.
Results: In a first series of experiments, wild type (WT) and CypD-/- mouse hearts were perfused
with glucose only-perfusate and subjected to 25 min ischemia and 45 min reperfusion. At
normoxic conditions, HK in cytosol and mitochondria was similar between WT and CypD-/- hearts.
CypD ablation protected against I/R injury (42% reduction in LDH release) and strengthened
ischemic preconditioning (IPC) protective effects as compared to WT, without affecting endischemic mtHK. In a second series of experiments, hearts were perfused with additional
substrates at physiological concentrations (glucose, glutamine, pyruvate, lactate), providing a
more stable preparation. Now CypD ablation resulted in more protection against I/R injury (80%
reduction in LDH release) that was associated with increased mtHK activity, leaving no room for
additional protection by IPC.
In conclusion:In glucose only-perfused hearts, end-ischemic mitochondrial-HK binding is
independent of CypD. In contrast, mouse hearts perfused with additional substrates,
mitochondrial-HK activity is increased with deletion of CypD. This was associated with increased
protection against cardiac I/R injury. Our data suggest that under more physiological conditions,
deletion of CypD results in increased mitochondrial-hexokinase activity during prolonged cardiac
ischemia.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A84
Internet-interventions targeting cardiovascular risk factors in older people: a systematic review
and meta-analysis
Cathrien R. L. Beishuizen1, Blossom C. M. Stephan2, Willem A. van Gool1, Carol Brayne3, Ron J. G.
Peters4, Sandrine Andrieu5, Miia Kivipelto6, Hilkka Soininen7, Wim B. Busschers8, Eric P. Moll van
Charante8, Edo Richard1,9
1
Department of Neurology, Academic Medical Centre, University of Amsterdam, PO Box 22660, 1100 DD,
2
Amsterdam. Institute of Health and Society, Newcastle University Institute for Ageing, Newcastle University,
3
Newcastle upon Tyne, NE2 4AX, United Kingdom. Department of Public Health and Primary Care, Cambridge
4
Institute of Public Health, Cambridge University, Cambridge CB1 8RN, United Kingdom. Department of Cardiology,
5
Academic Medical Centre, University of Amsterdam, PO Box 22660, 1100 DD, Amsterdam. Department of
Epidemiology and Public Health, Toulouse University Hospital and Inserm U1027, F-31073, 310 00 Toulouse, France.
6
Aging Research Center and Alzheimer Disease Research Center, Karolinska Institutet, Stockholm 17177 Sweden.
7
8
Department of Neurology, University of Eastern Finland, Kuopio 70211 Finland. Department of General Practice,
9
Academic Medical Centre, University of Amsterdam, PO Box 22660, 1100 DD, Amsterdam. Department of
Neurology, Radboud University Medical Center, 6525 GA Nijmegen.
Introduction: Internet-interventions can improve single cardiovascular risk factors in adult
populations. In view of global ageing and its associated burden of cardiovascular disease(CVD),
older people form an important target population as well. In this systematic review and metaanalysis, we evaluate whether internet-interventions for cardiovascular risk factor management
reduce the risk of CVD in older people.
Methods: Embase, Medline, Cochrane and Cinahl were systematically searched from 1995
onwards for randomized controlled trials on internet-interventions targeting cardiovascular risk
factors in older populations with a mean age of at least 50 years. Outcomes regarding treatment
effects on cardiovascular risk factors (blood pressure(BP), glycated haemoglobin(HbA1C), LDLcholesterol, smoking status, weight and physical inactivity) and incident CVD were pooled with
random effects models.
Results: 57 studies(N=19,862) fulfilled eligibility criteria and 47 studies contributed to metaanalysis. No difference in incident CVD was found between groups. A significant reduction in
systolic BP(-2.66 mmHg; 95%CI, -3.81 to -1.52), diastolic BP(-1.26 mmHg; 95%CI, -1.92 to -0.60),
HbA1c level(-0.13 %; 95%CI, -0.22 to -0.05), LDL-cholesterol level(-2.18 mg/dL; 95%CI, -3,96 to 0.41), weight (-1.34 kg; 95%CI -1.91 to -0.77) and an increase of physical activity(standardized
mean difference 0.25; 95%CI, 0.10 to 0.39) in the internet-intervention group was found. The
observed effects were more profound in studies with short (<12 months) follow-up and studies
that combined the internet-application with human support.
Conclusions and relevance: Internet-interventions can improve the cardiovascular risk profile of
older people, but the effects are modest and decline with time. There is insufficient evidence for
an effect on incident CVD.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A85
Improved survival after out-of-hospital cardiac arrest and use of automated external
defibrillators.
Blom MT1, Beesems SG1, Homma PC1, Zijlstra JA1, Hulleman M1, van Hoeijen DA1, Bardai A1,2,
Tijssen JG1, Tan HL1, Koster RW1.
1
Department of Cardiology, Academic Medical Center, University of Amsterdam, P.O. Box 22660, 1100 DD,
2
Amsterdam. Interuniversity Cardiology Institute Netherlands, Utrecht.
Background:
In recent years, a wider use of Automated External Defibrillators (AEDs) to treat out-of-hospital
cardiac arrest (OHCA) was advocated in The Netherlands. We aimed to establish whether survival
with favorable neurologic outcome after OHCA has significantly increased, and if so, if this is
attributable to AED-use.
Methods:
We performed a population-based cohort study, including patients with OHCA from cardiac
causes between 2006 and 2012, excluding EMS-witnessed arrests. We determined survival status
at each stage (to emergency room, to admission, to discharge) and examined temporal trends
using logistic regression analysis with ‘year-of-resuscitation’ as independent variable. By adding
each co-variable subsequently to the regression model, we investigated their impact on the Odds
Ratio (OR) of ’year-of-resuscitation’. Analyses were performed according to initial rhythm
(shockable vs. non-shockable), and AED-use.
Results:
Rates of survival with favorable neurologic outcome after OHCA increased significantly (N=6133,
16.2% to 19.7%; P for trend=0.021), though solely in patients presenting with a shockable initial
rhythm (N=2823, 29.1% to 41.4%; P for trend<0.001). In this group, survival increased at each
stage, but was strongest in the pre-hospital phase (OR 1.11; 95% confidence interval 1.06 to
1.16). Rates of AED-use almost tripled during the study period (21.4 to 59.3%, P for trend
<0.001), thereby decreasing time from emergency-call to defibrillation-device connection
(median 9.9 to 8.0 min, P<0.001). AED-use statistically explained increased survival with
favorable neurologic outcome, by decreasing the OR of ’year-of-resuscitation’ to a non-significant
1.04.
Conclusions:
Increased AED-use is associated with increased survival in patients with a shockable initial
rhythm. We recommend continuous efforts to introduce or extend AED-programs.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A86
The Impact of the Location of a Chronic Total Occlusion in a non-infarct Related Artery on Longterm Mortality in ST-Elevation Myocardial Infarction Patients.
Joelle Elias, Loes P C Hoebers, Ivo M van Dongen, Dagmar M. Ouweneel; Bimmer E P M Claessen,
Jan J Piek, Jose P S Henriques.
Department of Cardiology, Academic Medical Center, Amsterdam.
Introduction: Several studies evaluated the impact of a CTO on short- and long term mortality in
STEMI patients. It has been speculated that the adverse prognosis could differ per coronary
location.
Methods: Between January 2000 and December 2012, a total of 480 consecutive and unselected
STEMI patients with a CTO in a non-infarct related artery were included. The primary outcome for
the present analysis was all-cause three-year mortality, evaluating the impact of the coronary
CTO and infarct location.
Results: 413 had a single CTO in a non-infarct related artery whereas 67 patients had more than
1 CTO in whom mortality was higher. In patients with a single CTO, the highest risk of mortality
was observed when the culprit lesion was located in the LAD or proximal LCX or when the CTO
lesion was located in the proximal LAD. After correction for cardiogenic shock, only age remained
a significant predictor.
Conclusion: Previously, STEMI patients with a CTO have been associated with worse prognosis
than STEMI patients without a CTO. We now show that in these patients, LAD or proximal LCX
location for the culprit lesion or proximal LAD location for the CTO lesion were associated with
the highest risk. As a result almost all CTO patients are at increased risk for mortality due to the
combination of the culprit and CTO artery location.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A87
Absolute Discharge Levels or Relative Reduction in NT-proBNP during admission for acute
decompensated heart failure as a target to reduce 6-Month Mortality?
S. Stienen, K. Salah, L. Eurlings, P. Bettencourt, JM Pimenta, M. Metra, A. Bayes-Genis, V.
Verdiani, L. Bettari, V. Lazzarini, J.P Tijssen, Y.M. Pinto and W.E. Kok.
Department of cardiology, AMC, Amsterdam and collaborating Institutes.
Background: NT-proBNP is a strong predictor for readmissions and mortality in patients
discharged after admission for acute decompensated heart failure (ADHF). Although both NTproBNP levels at discharge and relative reduction percentages have been incorporated in risk
models, it is still a matter of debate which NT-proBNP level (either absolute or relative) should be
used as pre-discharge treatment target in an ADHF population.
Methods: Our study population was assembled from 7 European ADHF cohorts, which had NTproBNP measured at admission and discharge. We defined NT-proBNP discharge targets: <1500,
<3000, <5000 and <15000 ng/L, and relative reductions from admission to discharge of >30% ,
>50% and >70%. Population attributable risk fraction (PARF)) is the proportion of all-cause
mortality in the population within 6 months after discharge that could be reduced if a risk factor
(NT-proBNP level above target) was not present. PARF was determined for each target (% ± 95%
CI) and compared with the percentage of patients attaining NT-proBNP target levels. Attainability
of targets was investigated by univariate and multivariate logistic regression analysis.
Results: A total number of 1266 patients (median age 74 (64-80),60% male) was studied. Six
month mortality was observed in 15% of patients and decreased with lower absolute discharge
levels and greater percentages reduction in NT-proBNP. For every absolute discharge level of NTproBNP, a corresponding reduction percentage of NT-proBNP was found that resulted in the
same PARF. Highest PARF was observed for <1500 ng/L at discharge or >70% reduction in NTproBNP but attainability of these targets was low (27% and 22% resp.) Strongest predictor for not
reaching these targets was admission NT-proBNP levels. Significant differences in PARF were
found between tertiles of admission NT-proBNP for absolute NT-proBNP targets, while there
were no significant differences in PARF for relative reduction NT-proBNP targets.
Conclusion: In an ADHF population, absolute or relative reductions as NT-proBNP targets are
interchangeable in the effect on PARF. Relative reduction targets show similar PARFs among
admission NT-proBNP categories while absolute targets show varying PARFs. A relative reduction
target would probably lead to the most consistent mortality reducing effect across the whole
spectrum of ADHF patients.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A88
KBTBD13: A NOVEL GENE IMPLICATED IN CARDIOMYOPATHY
Josine M. de Winter1, Maarten P. van den Berg2, Joshua Strohm3, Jan D. Jongbloed4, Vaishali
Kakkar1, Aref Najafi1, Wies Lommen1, Wilma van der Roest4, Jan van Wijngaarden5, Janneke
Timmermans6, Erik Jan Kamsteeg7, Baziel G. van Engelen8, Robbert-Jan van der Pijl1,3, Henk
Granzier3, Diederik W.D. Kuster1, Jolanda van der Velden1, Nicol C. Voermans8, Karin van
Spaendonck-Zwarts4,9* and Coen A.C. Ottenheijm1,3*
1 Dept. of Physiology, VU University Medical Center, Amsterdam,2 Dept. of Cardiology, University Medical Centre, Groningen. 3
Dept. of Cellular and Molecular Medicine, University of Arizona, Tucson AZ, USA. 4 Dept. of Genetics, University Medical Center
Groningen, Groningen, 5 Dept. of Cardiology, Deventer Hospital, Deventer. 6 Dept. of Cardiology, Radboud University Medical
Centre, Nijmegen. 7 Dept. of Human Genetics, Radboud University Medical Centre, Nijmegen. 8 Dept. of Neurology, Radboud
University Medical Centre, Nijmegen. 9 Dept. of Clinical Genetics, Academic Medical Center, University of Amsterdam,
Amsterdam. * KSZ and CACO contributed equally.
Introduction: Patients with nemaline myopathy caused by mutations in KBTBD13 have weakness
and slowness of skeletal muscles. KBTBD13 is also expressed in cardiac muscle, but a cardiac
phenotype has not yet been reported. Recently, a member of a large family with a KBTBD13
mutation visited our cardiogenetic outpatient clinic because of dilated cardiomyopathy (DCM).
Hence, we aimed to elucidate the consequences of KBTBD13 mutations on cardiac function.
Methods: A pedigree was constructed and medical reports on cardiac characteristics were
collected. Next, cardiological evaluation (ECG, echocardiography, ultrasound and MRI on
indication) and DNA-diagnostics of the Dutch founder mutation in KBTBD13 (c.1222C>T,
p.Arg408Cys) were offered to the DCM-patient and her relatives (family 1). Moreover, we
collected cardiac characteristics of three newly identified families with this particular mutation
(family 2-4). To investigate the role of KBTBD13 on the contractile function of the heart, we
engineered a Kbtbd13-knockout mouse model and performed echocardiography at rest and
during stress (i.e. administration of dobutamine) in three-month-old mice.
Results:In family 1, cardiological evaluation revealed (mild) left ventricular dysfunction or DCM in
four members, and pedigree analysis revealed three cases of sudden cardiac death. Strikingly, in
family 2-4 also at least one individual was identified with reduced left ventricular function i.e.
signs of cardiomyopathy. Mouse echocardiography revealed that during stress Kbtbd13-knockout
mice presented: a lower heart rate (-12%), lower fractional shortening (-16%) and lower ejection
fraction (-7%). Longitudinal strain rate of the left ventricle was reduced by 23% in Kbtbd13knockout mice.
Conclusion:Mutations in KBTBD13 can induce cardiomyopathy. Cardiological evaluation is
recommended for families with KBTBD13 mutations. Our novel mouse model is a useful tool for
further elucidation of the cardiac consequences of KBTBD13 mutations.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A89
Optimizing cellular delivery of HCN2/SkM1 to induce long-term biological pacing
Lucía Cócera Ortega, Anna MD Végh, Elisabeth M Lodder, Arie O Verkerk, Mischa Klerk, Erik
Oosterwerff, Vincent M Christoffels, Connie Bezzina, Hanno L Tan, Marie Jose TH Goumans,
Gerard JJ Boink
Department of Experimental Cardiology AMC, Amsterdam.
Background:
Biological pacemakers based on adenoviral overexpression of the Hyperpolarization-activated
Cyclic Nucleotide-gated channel (HCN2) and the Skeletal Muscle sodium channel (Skm1) have
shown efficient in vivo function. An important next step is the optimization of a cellular delivery
to induce long-term function. Human Cardiomyocyte Progenitor Cells (CMPCs) have shown to
efficiently deliver HCN4-based biological pacing and persist in the heart for at least 3 months
after injection. In the present study we therefore optimized electroporation protocols to
efficiently introduce HCN2/SkM1 into CMPCs.
Methods and results:
Human CMPCs were isolated from fetal hearts using magnetic beads coated with a Sca-1
antibody, cultured in non-differentiating conditions, and electroporated (Amaxa nucleofector)
with GFP, HCN2+GFP, Skm1+GFP or HCN2+Skm1+GFP. To enhance expression, a Woodchuck
hepatitis virus Posttranscriptional Regulatory Element (WPRE) motif was cloned into the CMVdriven backbones. Addition of the WPRE was highly effective in correcting the suboptimal
expression levels of HCN2 and SkM1 which resulted in an increase in the electroporation
efficiency from respectively 10% up to 80% as detected by immunocytochemistry. Functional
presence of the relevant ion currents was confirmed using perforated patch clamping at 37ºC.
The HCN2 current density at -120 mV was -32±6 (n=3), which is comparable to previously
reported values for HCN2-loaded mesenchymal stem cells.
Conclusions:
Electroporation is an efficient method to introduce relatively large ion channel genes into
CMPCS. Introduction of the WPRE motif in the expression backbone importantly increased
expression. HCN2/SkM1-loaded CMPCs hold significant promise for the introduction of long-term
biological pacing.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A90
Orphan nuclear receptor Nur77 affects cardiomyocyte calcium homeostasis and adverse
cardiac remodelling.
Lejla Medzikovic1,2, Cees A Schumacher3, Arie O Verkerk4, Elza D van Deel5, Rianne Wolswinkel3, Ingeborg
van der Made3, Natascha Bleeker1, Daniella Cakici1, Maarten MG van den Hoogenhof3, Farid Meggouh1,
Esther E Creemers3, Carol Ann Remme3, Antonius Baartscheer3, Robbert J de Winter2, Carlie JM de Vries1, E
Karin Arkenbout2,6, Vivian de Waard1*
1
2
3
4
Department of Medical Biochemistry, Department of Cardiology, Department of Experimental Cardiology, Department of
5
Anatomy, Embryology and Physiology, Academic Medical Center, University of Amsterdam, Division of Cardiology, Thoraxcenter,
6
Erasmus Medical Center, University of Rotterdam , Department of Cardiology, Tergooi Hospital, Blaricum.
Introduction: Distinct stressors may induce heart failure. As compensation, β-adrenergic
stimulation enhances myocardial contractility by elevating cardiomyocyte intracellular Ca2+
([Ca2+]i). However, chronic β-adrenergic stimulation promotes adverse cardiac remodelling.
Cardiac expression of nuclear receptor Nur77 is enhanced by β-adrenergic stimulation, but its
role in cardiac remodelling is still unclear.
Methods and results: We show high and rapid Nur77 upregulation in cardiomyocytes stimulated
with β-adrenergic agonist isoproterenol. Nur77 knockdown in culture resulted in hypertrophic
cardiomyocytes. Ventricular cardiomyocytes from Nur77-deficient (Nur77-KO) mice exhibited
elevated diastolic and systolic [Ca2+]i and prolonged action potentials compared to wild type
(WT). In vivo, these differences resulted in larger cardiomyocytes, increased expression of
hypertrophic genes, and more cardiac fibrosis in Nur77-KO mice upon chronic isoproterenol
stimulation. In line with the observed elevated [Ca2+]i, Ca2+-activated phosphatase calcineurin
was more active in Nur77-KO mice compared to WT. In contrast, after cardiac pressure overload
by aortic constriction, Nur77-KO mice exhibited attenuated remodelling compared to WT.
Conclusion: Nur77 deficiency results in significantly altered cardiac Ca2+ homeostasis and distinct
remodeling outcome depending on the type of insult. Detailed knowledge on the role of Nur77 in
maintaining cardiomyocyte Ca2+ homeostasis and the dual role Nur77 plays in cardiac remodeling
will aid in developing personalized therapies against heart failure.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A91
Lymphocytic myocarditis can lead to inflammation in the coronary arteries which may induce
an acute myocardial infarction
Linde Woudstra1,4 P. Stefan Biesbroek2,4,5 Reindert W. Emmens1,4 Lynda J. Juffermans2,4 Albert C.
van Rossum2,4 Hans W. M. Niessen1,2,4 and Paul A. J. Krijnen1,4
1
2
3
Department of Pathology , Cardiology , Cardiothoracic Surgery , VU University Medical Center, Amsterdam.; ICaR4
5
VU , Institute for Cardiovascular Research of the Vrije Universiteit of Amsterdam, Amsterdam. ICIN , Interuniversity
Cardiology Institute of the Netherlands, Utrecht.
Introduction:
Acute myocarditis can trigger coronary vasospasm, which may result in thrombus formation and
are at risk for subsequent myocardial infarction (MI). We hypothesize that LM triggers increased
inflammation in coronary lesions and increased plaque instability facilitating MI.
Methods:
The three main coronary arteries were obtained at autopsy from patients with either LM without
MI (LM), with preexisting LM and MI <6 hours old (LM+MI) and from controls. In coronary
segments containing atherosclerotic lesions (n=221), lymphocytes, macrophages, neutrophils,
mast cells and intraplaque hemorrhage (IPH) were quantified in the intima, media and adventitia
via immunohistochemistry. Plaque stability was determined via histological criteria.
Results:
The number of macrophages and neutrophils per mm2 were comparable in all patient groups.
The percentage of lymphocytes was significantly increased in the coronary arteries of LM+MI
patients compared to LM and control patients. Interestingly, the number of mast cells per mm2 in
the coronary lesions were significantly increased in LM+MI patients compared to LM and control
patients. This increase in mast cells was especially due to an increase in the media and adventitia.
The percentage of instable plaques and also IPH in LM+MI patients was significantly higher than
in LM patients and controls.
Conclusions:
This study shows increased coronary plaque infiltration of lymphocytes and mast cells in patients
with preexistent LM that develop MI, than patients with LM only or controls, coinciding with
significantly more unstable plaques and IPH. These results indicate that LM may facilitate
development of MI in a sub-population of LM patients.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A92
Remodeling of distinct sodium channel pools in the murine failing heart
M.R.Rivaud1,2, X. Lin1, E. Augullo-Pascal1, C.R.Bezzina2, C.A. Remme2, M. Delmar1
1 Division of Cardiology, New York University, New York, USA; 2 Department of Experimental Cardiology, Academic
Medical Center, Amsterdam.
Introduction:
Nav1.5, the major pore-forming protein of the cardiac sodium channel, localizes to distinct
“pools” on the cell surface. At each pool, Nav1.5 associates with region-specific molecules,
yielding complexes whose function and composition are location-specific. The remodeling of
these pools in the setting of heart failure remains to be studied.
Methods:
Mice were subjected to transverse Aortic Constriction (TAC). All data were collected 6 weeks
post-surgery. Voltage clamp recordings of sodium current (INa) were obtained in whole-cell and
cell-attached (macropatch) configurations. Sub-diffraction limit molecular imaging was carried
out by direct stochastic optical reconstruction microscopy (dSTORM).
Results:
In TAC hearts, whole-cell INa decreased by 30% (-29.2±2.2 pA/pF, n=11 vs. -40.8±3.6, n=12 in
Sham, p<0.05). Macropatch data showed reduced INa (56.3% ) in the mid-section of TAC cells (84.3±9.7 pA, n=14 vs. -193±19, n=13 in Sham, p<0,05) and in the cell end (45%; -258.1±7.4 pA,
n=5 vs. -469.5±84 pA, n=5 in Sham, p<0,05). dSTORM showed reduced average Nav1.5 cluster
size at the lateral membrane (30.7x103±14x103 nm2 vs44.1x103±14.8x103 in Sham, p<0.05) and
at the cell end (27.5x103±18.7x103 vs 40x103±18.1x103, p<0.05) of TAC cells, and a
corresponding 35% increase in cluster numbers at the cell end (2.7x10-6±0.22x10-6 vs 1.9x106±0.14x10-6 in Sham,p<0.05).
Conclusions:
We show functional remodeling of Nav1.5 leading to reduced INa in failing hearts including a
reduction in the number of large Nav1.5 clusters, and an increase in the number of clusters of
smaller size. Ongoing studies seek to define the molecular mechanisms leading to nano-structural
remodeling of Nav1.5 in the failing heart.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A93
The orphan drug 4-phenyl butyrate protects against Atrial Fibrillation by blocking ER stressinduced autophagy
M. Wiersma (University Medical Center Groningen, Groningen), R.A.M. Meijering (University Medical Center
Groningen, Groningen), X.Y. Qi (Montreal Heart Institute and Université de Montréal, Montreal, Canada), D.
Zhang (University Medical Center Groningen, Groningen), F. Hoogstra-Berends (University Medical Center
Groningen, Groningen), O.C.M. Sibon (University Medical Center Groningen, Groningen), S. Nattel, R.H.
Henning (University Medical Center Groningen, Groningen), B.J.J.M. Brundel (University Medical Center
Groningen, Groningen; VU University Medical Center, Amsterdam)
Introduction:
Atrial fibrillation (AF) is characterized by a self-perpetuating nature, driven by structural
remodelling of cardiomyocytes, resulting in electrical disturbances, contractile dysfunction, and
increasing difficulty in maintaining normal sinus rhythm. Previous work suggest an important
contribution of altered proteostasis to cardiomyocyte remodelling. Here, we explore the role of
autophagy, an important player in proteostasis via protein and organelle degradation.
Methods:
Electrophysiology and/or biochemistry were determined of HL-1 mouse atrial cardiomyocytes
and Drosophila subjected to tachypacing, in vivo dog model for AF, and atrial tissue from patients
with permanent AF.
Results: Tachypacing atrial cardiomyocytes caused a gradual and significant activation of
autophagy, evidenced by increased autophagic flux, autophagosome formation, and p62
degradation. Endoplasmic reticulum (ER) stress was responsible for autophagy-induction, as
indicated by upregulation of ER stress markers. Inhibition of ER stress and autophagy by the ER
stress inhibitor 4-phenyl butyrate prevented tachypacing-induced contractile dysfunction in
cardiomyocytes and Drosophila. Furthermore, in vivo treatment with 4-phenyl butyrate
protected atrial tachypaced dogs from electrical and contractile dysfunction. Finally, induction of
ER stress and autophagy was also found in atrial tissue of permanent AF patients, where levels of
autophagy correlated with markers of cardiomyocyte remodelling and correlated inversely with
the amount of myolysis.
Conclusion:
These results demonstrate atrial-cardiomyocyte tachycardia and clinical AF to result in ER stress
and consequent activation of autophagy and AF progression, and provide proof of concept for an
already marketed inhibitor of ER stress, the chemical chaperone 4-phenyl butyrate, to expand
the therapeutic options in AF.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A94-1 (first session)
A large permissive regulatory domain controls Tbx3 expression in the cardiac conduction
system
Henk van Weerd1, Ileana Badi1, Malou van den Boogaard1, Harmen J G van de Werken2, Wouter
de Laat2, Phil Barnett1, Vincent M Christoffels1
1
2
Department of Anatomy, Embryology & Physiology, Academic Medical Center, Amsterdam. Hubrecht InstituteKNAW and University Medical Center Utrecht, Utrecht,
Introduction:
The evolutionary conserved Tbx3/Tbx5 gene cluster encodes T-box transcription factors that play
crucial roles in the development and function of the cardiac conduction system (CCS). Tbx3 and
Tbx5 are expressed in overlapping patterns and function in strictly tissue-specific and dosedependent manners. Genetic variants have been identified in the gene desert flanking the cluster
that influence conduction system function in humans, illustrating the importance of
understanding the complex transcriptional regulation of expression of both genes.
Methods/Results:
By performing 4C-seq we generated a high-resolution model of the three-dimensional
architecture of the Tbx3/Tbx5 locus and found that its regulatory landscape is in a preformed,
permissive conformation that is similar in embryonic heart, brain and limbs. Tbx3 and its flanking
non-coding region form a 1 Mbp loop that is physically separated from neighbouring Tbx5 and
Med13l loops, suggesting enhancer sharing within the evolutionary conserved Tbx3/Tbx5 cluster
is unlikely to occur. Among the multiple sites contacting Tbx3, we identified two murine
enhancers that synergize to activate Tbx3 expression in the atrioventricular conduction system.
Using TALEN-mediated genome editing, we deleted a 70 kbp distal region that strongly contacts
Tbx3 and encompasses the mouse orthologue of a region that in humans contains variation
influencing conduction parameters.
Conclusions:
The Tbx3 locus forms an autonomous regulatory domain with multiple combinatorial regulatory
elements that control the precise pattern of Tbx3 in the CCS. Current analysis of the effect of
TALEN-mediated deletion of potential regulatory sequences, combined with the locus-wide
identification of additional regulatory sequences (STARR-seq, reporter-modified bacterial
artificial chromosomes), will add to our understanding of how the gene desert flanking Tbx3 is
involved in its regulation and how variation affecting its regulatory elements influences its
expression and conduction system function.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A94-2 (second session)
AVIATOR Initiative: Aortic Valve Insufficiency and Ascending Aorta Aneurysm International
Registry.
Authors:
Frederiek de Heer, Jolanda Kluin, Hans-Joachim Schäfers, Johanna Takkenberg, Emmanuel Lansac
on behalf of the SHVD Aortic Valve Repair working group.
Objective:
Current guidelines for patients with aortic valve insufficiency and/or ascending aorta aneurysm
are based on small single expert center case series with a limited follow-up duration. The Heart
Valve Society established the international prospective multicentre registry AVIATOR in 2013 to
provide sufficient patient numbers, to address key epidemiological and therapeutic issues and to
standardize indications for surgery as well as the place of repair versus replacement in aortic
valve surgery.
Methods:
All patients, aged 18 years or older, operated for ascending aorta aneurysm (including root
and/or supra coronary aorta and aortic dissection) and/or isolated aortic insufficiency (including
congenital mixed aortic valve disease), should be enrolled in an intention to treat design. Surgical
techniques include valve repair and replacement, valve sparing aortic root replacement and
Bentall procedures. Baseline, procedural and early outcome data are collected prospectively
during hospital stay. Thereafter annual follow-up will take place.
Results:
Currently 18 cardio-thoracic surgery centers from Belgium, Czech Republic, France, Germany,
Italy, Japan, Netherlands, Spain and Slovakia and United Kingdom enrolled in the database. They
included 529 patients thus far.
Conclusions:
The international prospective multicentre AVIATOR registry will allow for acceleration of
knowledge on the determinants of outcome in patients with aortic valve insufficiency and/or
ascending aorta aneurysm. By combining surgical experience from multiple centers, and applying
uniform definitions of echo and outcome parameters, it will become possible to provide a solid
evidence base to clarify and standardize the place of repair versus replacement in aortic valve
surgery.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A95
Measuring the power generated by a cardiac myocyte in the cardiac cycle
M. Helmes1, A.Najafi1, D. Iannuzzi2, J. van der Velden1
1
2
VUmc, department of Physiology, VU, department of Physics.
Introduction:
A method is presented to measure the power output of isolated intact cardiac myocytes.
Methods:
The mycoytes were attached to a very sensitive and fast force transducer on one side (OptiForce
form Ionoptix) and a direct drive piezo-translator on the other side to stretch or shorten the
myocyte. This allowed feedback based force control. Force clamps were used to impose a
minimum ‘pre-load’ level and maximum ‘after-load’ level between which the myocyte was
allowed to operate, mimicking the aortic and atrial pressure respectively. As long as the myocyte
reaches the after load level in systole, the loaded shortening results in a force versus length curve
that forms a work loop, analogous to the pressure volume curve in the whole heart.
Results and conclusion:
Within the measured range (0-1.5 µNewton pre-load, corresponding to sarcomere lengths of
1.86±0.07µm through 1.96±0.08µm) the end-systolic and end-diastolic force-length relations
were linear. Measuring the area within each work loop provided the external work performed by
the mycoyte for each contraction. A power curve was created by varying the after load for a
given pre-load and multiplying the work per contraction with the pacing frequency. For rat
myocytes at 37ºC the peak power output increased linearly with pre-load within the range
measured, and at a pre-load of 1.5 µNewton varied between 25 and 90 pJ.s-1. Maximum power
output was achieved at pacing frequencies between 6-8 Hz.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A96
Exome sequencing identifies a homozygous SLC22A5 mutation leading to primary carnitine
deficiency in a consanguineous Moroccan family affected by hypertrophic cardiomyopathy and
sudden cardiac death
N. Lahrouchi1, M. Mansouri2, L. Zniber3, A.V. Postma4, I. Ratbi2, A. Sefiani2, E.M. Lodder1, C.R.
Bezzina1
1. Department of Clinical and Experimental Cardiology, Academic Medical Center, Amsterdam. 2. Centre de
Génomique Humaine, Faculté de Médecine et de Pharmacie, Université Mohamed V Souissi, Rabat, Morocco 3.
Centre de Cardiologie à Rabat, Rabat, Morocco 4. Department of Anatomy, Embryology & Physiology, Academic
Medical Center, Amsterdam.
Introduction: Hypertrophic cardiomyopathy (HCM) is a severe disease associated with high
mortality and morbidity. The cause of HCM in children is poorly understood and only in one third
of children a cause can be identified. Inborn errors of metabolism account for a small fraction of
pediatric HCM.
We characterized a Moroccan consanguineous family with HCM and sudden death. Two siblings
died suddenly at the ages of 6 months and 3 years. The only living affected sibling is a 4-year old
girl with severe anemia, fatigability and HCM. Her 7-year old brother is unaffected. The aim of
this study was to elucidate the genetic cause underlying the disease in this family.
Methods: We performed exome sequencing in both unaffected parents, the affected sibling and
her unaffected brother. Given the rarity of this disorders we filtered out all variants with a
frequency >0.1% in publically available databases and in-house exome data. Based on a recessive
model of inheritance and consanguinity, we next searched for genes carrying homozygous
variants in the affected child. Variants that satisfied these criteria were validated using Sangersequencing.
Results: After the filtering process three homozygous variants were found in the affected sibling
of which one (SLC22A5, p.Tyr396X) was located in a gene associated to primary carnitine
deficiency (PCD). Both parents and the unaffected brother were heterozygous carriers of this
mutation. The findings were confirmed with Sanger sequencing. In addition, this mutation was
previously described in compound heterozygosity in a patient diagnosed with PCD.
Conclusion: Using exome sequencing we identified PCD as the underlying cause of HCM and
sudden cardiac death in this family. PCD is an autosomal recessive disorder of mitochondrial βoxidation that arises due to a defect in the high-affinity carnitine transporter, encoded for by
SLC22A5, and often presents with cardiomyopathy at young age. Interestingly, if diagnosed early,
clinical manifestations of the disorder can be completely reversed by supplementation of
carnitine. Because of this, high dose oral carnitine supplementation in the patient was started
directly after confirmation of the diagnosis.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A97
Expression of missense and truncating MYBPC3 mutations does not prevent hypercontractility
of Mybpc3-targeted knock-out mouse engineered heart tissue
Paul J.M. Wijnker1,2,3, Felix W. Friedrich1,2, Alexander Dutsch1,2, Alexandra Eder1,2, Ingra Vollert1,2,
Giulia Mearini1,2, Thomas Eschenhagen1,2, Jolanda van der Velden3, Lucie Carrier1,2
1
Department of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical
2
Center Hamburg-Eppendorf, Hamburg, Germany DZHK (German Centre for Cardiovascular Research), partner site
3
Hamburg/Kiel/Lübeck Department of Physiology, Institute for Cardiovascular Research, VU University Medical
Center, Amsterdam.
Introduction:
Hypertrophic cardiomyopathy (HCM) is a cardiac genetic disease characterized by left ventricular
hypertrophy, diastolic dysfunction and myocardial disarray. The most frequently mutated HCM
gene is MYBPC3, encoding cardiac myosin-binding protein-C (cMyBP-C), which has a central role
in the structure and function of cardiac sarcomere. Here we compared the effects of a missense
mutation (c.1591G>C) and a truncating mutation (c.2373_2374insG) in MYBPC3 on contractile
parameters of engineered heart tissue (EHT).
Methods and Results:
EHTs were generated from Mybpc3-targeted knock-out (KO) and wild-type (WT) mouse cardiac
cells. MYBPC3 WT and mutants were expressed via adeno-associated virus (AAV6) transduction
of KO-EHTs. KO-EHTs displayed higher maximal force and sensitivity to external [Ca2+] than WTEHTs. By comparing the effect of different multiplicities of infection (MOI) of AAV6-WT-MYBPC3
(30 to 300), we found that MOI100 resulted in ~73% of WT cMyBP-C level but did not prevent the
KO phenotype, whereas MOI 300 resulted in ~99% of WT cMyBP-C level and prevented the KO
phenotype. Expression of missense and truncated cMyBP-C (MOI 300) resulted in ~95% and ~33%
of protein levels, respectively. However, they did not correct the phenotype of KO-EHTs. Cotransduction of KO-EHTs with WT plus either mutant (MOI 150 each) failed to restore force to WT
level. Immunofluorescence analysis revealed correct incorporation of WT and missense cMyBP-C
in the sarcomere, while truncated cMyBP-C was found in the cytosol.
Conclusion:
KO-EHTs exhibited hypercontractility. More than ~73% of exogenous WT-cMyBP-C protein level
was required to prevent the disease phenotype of KO-EHTs, supporting haploinsufficiency as
disease-causing mechanism. In contrast to WT, both mutants did not prevent the KO-EHTs
hypercontractile phenotype, supporting their causal effect. In conclusion, EHTs can be used as a
platform to evaluate the impact of (newly identified) MYBPC3 gene variants.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A98
A regulatory domain controls the conserved Nppa-Nppb gene cluster during heart
development and stress
Irina A. Sergeeva1, Ingeborg B. Hooijkaas1, Jan M. Ruijter1, Ingeborg van der Made2, Harmen J.
van de Werken3, Esther E. Creemers2, Vincent M. Christoffels1
1
Dept. of Anatomy, Embryology and Physiology, Academic Medical Center, Amsterdam, The Netherlands,
Meibergdreef 9, 1105 AZ, Amsterdam, the Netherlands
2
Dept. of Experimental Cardiology, Heart Failure Research Center, Academic Medical Center, Amsterdam, The
Netherlands, Meibergdreef 9, 1105 AZ, Amsterdam, the Netherlands
3
Cancer Computational Biology Center, 3015 CN, Erasmus MC, Rotterdam, the Netherlands.
Introduction:
For more than two decades, Nppa and Nppb have served as paradigms for studying cardiac
developmental and stress-responsive regulation of transcription. Moreover, because of their
organization in an evolutionary conserved cluster and their near identical expression patterns,
the genes are also valuable models to study coregulation and regulatory landscape organization.
Methods:
We analyzed the in vivo chromatin conformation, epigenetic status and enhancer potential of
sequences of the locus harboring the Nppa-Nppb cluster to gain insight into its transcriptional
control.
Results:
Our data indicate that the regulatory landscape of the cluster is present within a 60 kbp CTCF
sites-flanked region, centered around Nppb. The conformation of the chromatin brings both
promoters and several potential regulatory elements in close proximity to each other. This
conformation is similar in different tissues and developmental stages. The distribution of
H3K27ac and association of Pol2 across the locus changed during cardiac hypertrophy, revealing
their potential involvement in stress-mediated gene regulation. We also generated double
reporter transgenic mice carrying modified bacterial artificial chromosomes for simultaneous
monitoring Nppa and Nppb expression. Functional analysis in vivo revealed that Nppa and Nppb
share developmental, but not stress-response enhancers responsible for their co-regulation.
Moreover, the Nppb promoter was required, but not sufficient, for hypertrophy-induced Nppa
expression.
Conclusions:
The developmental regulation and stress-response of the Nppa-Nppb cluster involves the
concerted action of different elements that act within a structurally rigid regulatory domain
relatively isolated from the surrounding genes.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A99
The inflammatory infiltrate in the endomyocardial biopsy area is increased significantly
compared to the entire myocardium in patients with lymphocytic myocarditis
Linde Woudstra1,4 Reindert W. Emmens1,4 P. Stefan Biesbroek2,4,5 Lynda J. Juffermans 2,4 Albert C.
van Rossum2,4 Hans W. M. Niessen1,3,4 and Paul A.J. Krijnen1,4
1
2
3
Departments of Pathology , Cardiology , Cardiothoracic Surgery , VU University Medical Center, Amsterdam.; ICaR4
5
VU , Institute for Cardiovascular Research, VU University Medical Center, Amsterdam, The Netherlands; ICIN ,
Interuniversity Cardiology Institute of the Netherlands, Utrecht.
Introduction: To diagnose lymphocytic myocarditis (LM) immuno-histopathological examination
of endomyocardial biopsies (EMB) is still the gold standard. Although, due to the often patchy
inflammatory infiltrate in LM, the risk of underdiagnosis in EMB is well recognized, how this
infiltrate in the endomyocardial biopsy area relates to that in the rest of the myocardium is
unknown. The aim of the study was to compare the inflammatory infiltrate in the
endomyocardium of the left ventricular posterior wall (LVPW), a common biopsy area, to that in
the entire LVPW. Moreover, we hypothesized that the use of a more common lymphocyte
marker, i.e. CD45, could increase the diagnostic sensitivity of LM.
Methods: The LVPW was obtained at autopsy of patients diagnosed with LM (n=18) and control
patients (n=10). In addition, EMB taken from LVPW of living patients with confirmed LM (n=9)
and hearts of mice with coxsackievirus B3-induced acute viral myocarditis (n=9) were used.
Macrophages (anti-CD68), T-cells (anti-CD3) and lymphocytes (anti-CD45) we quantified per mm2
in a 2.0 mm thick area of the endomyocardium of the LVPW (representing the biopsy area), the
remaining LVPW, in EMBs and in the mouse hearts.
Results: In mice with acute viral myocarditis number of CD45+ cells/mm2 was increased
compared to the number of CD3+ cells/mm2. This was also observed in the EMB. In the autopsied
hearts, using the cut–off value of ≥14 leukocytes/mm² (number of CD3+ and CD68+ cells) the
diagnosis of LM was confirmed in the biopsy area of 39% of LM patients only, while with the
combined scores of CD45+ and CD68+ cells the diagnosis of LM was confirmed in 56% of LM cases.
Importantly, a significant increase of CD45+ lymphocytes/mm2 was observed in the biopsy area
compared to the remaining LVPW for both LM and control patients.
Conclusions: This study demonstrates that the inflammatory infiltrate in the endomyocardium of
the LVPW is increased significantly compared to the ventricular wall, indicating that despite the
risk of underdiagnosis in EMB, the sampling area constitutes the highest chance for positive
pathological diagnosis. Furthermore, the sensitivity of this diagnosis may be increased with the
common lymphocyte marker CD45.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A100
Off-pump Left Ventricle Reconstruction using the Revivent Myocardial Transcatheter Anchoring
System in Patients with severe Ischaemic Cardiomyopathy
Y. Aziz, A. van Wijk, W.J.P van Boven, B.A.J.M de Mol.
The Department of Cardio-thoracic Surgery, Academic Medical Centre, Amsterdam
Introduction:
Left ventricular (LV) remodelling, a pathological process that follows myocardial infarction (MI) ,
occurs in more than 30% of patients treated with acute reperfusion therapies and
cardioprotective medications. This process leads to alteration in the size, shape, structure and
function of the ventricles. A surgical treatment to reshape the dilated LV and restore cardiac
function is the left ventricle reconstruction (LVR). By excluding the MI scar and rebuilding the
heart into its original shape and volume, the stress on the heart muscle is reduced and the
functionality is restored. The Revivent Myocardial Anchoring Transcatheter (TC) System is a new,
less-invasive method of LVR.
Methods:
The Revivent TC System will be implanted in patients with advanced ischaemic cardiomyopathy
(ICM) after anterior wall infarction. This system uses anchors that are transcatheterly implanted
into the scarred part of the heart, which when deployed excludes that area. This new technique
can be performed on the beating heart without the use of cardiopulmonary bypass and median
sternotomy.
Results:
This feasibility study is designed to investigate a new less-invasive system for the treatment of
severe ICM. The
aim of our study to analyze the characteristics of patients with severe ICM who would profit
from this new
device. The study will start-off by the end of 2015.
Conclusion:
This technique has not yet been introduced in the Netherlands and might be a remedy for
growing group of patients with ICM.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A101
Microtubule-associated protein RP/EB family member 1 modulates cardiac conduction
Portero V, Podliesna S, Veerman C, Verkerk A, Klerk M, Lodder E, Mengarelli I, Bezzina C, Remme
CA
Department of Experimental Cardiology, Academic Medical Centre, University of Amsterdam
Introduction:
Microtubule-associated protein RP/EB family member 1 (EB1 protein) encoded by the gene
MAPRE1 is part of a protein network which binds microtubules at their (+)-end extremities
underneath the cell membrane. It has been shown that EB1 regulates trafficking of connexin43
(Cx43) at the membrane and is located in adult cardiomyocytes at the intercalated discs.
Furthermore, EB1 protein is removed from intercalated discs in cardiac hypertrophy, heart failure
and in the setting of Cx43 mutations. Recent studies have also shown that EB1 protein is
implicated in the subcellular localisation of sodium channels and potassium channels in
neurones. We therefore decided to explore the electrophysiological role of EB1 expression on
cardiac sodium channels involved in cardiac conduction and arrhythmias.
Results:
eQTL experiments performed on an F2 population of mice of two separate inbred strains carrying
a sodium channel mutation showed a strong negative correlation between the expression of the
MAPRE1 gene and QRS duration on the surface ECG, suggesting a functional role for EB1 protein
in cardiac conduction. We confirmed by co-immuno precipitation technique the physical
interaction between EB1 and the major cardiac sodium channel Nav1.5. The overexpression of
EB1 protein in HEK cells transfected together with Nav1.5 led to an increase sodium current
density without affecting kinetics suggesting an underlying increased membrane trafficking of the
Nav1.5 protein.
Conclusion:
In this study we proved the functional role of EB1 protein in cardiac conduction and we highlight
its direct regulation of the sodium channel Nav1.5. We recently produced and validated an SHRNA lentivirus capable of “knocking down” EB1 in human iPS-derived cardiomyocytes (hiPS-CM).
Ongoing electrophysiological studies are aimed at characterizing the modulatory effect of
EB1/MAPRE1 on ion currents and action potential characteristics in hiPS-CM using the dynamic
clamp technique.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A102
PARP1 activation causes NAD+ depletion and contractile dysfunction in experimental and
human Atrial Fibrillation
Deli Zhang1, Xu Hu1, Marit Wiersma1, Femke Hoogstra-Berends1, Gunseli Cubukcuoglu Deniz2,3,
Serkan Durdu2,3,4, Ruchan Akar3,4, Ody C.M. Sibon5, Robert H. Henning1, Bianca J.J.M. Brundel1
1 Department of Clinical Pharmacy & Pharmacology, University Medical Center Groningen, University of Groningen 2
Ankara University Biotechnology Institute, Ankara, Turkey; 3 Ankara University Stem Cell Institute, Ankara, Turkey; 4
Department of Cardiovascular Surgery, Ankara University School of Medicine, Turkey; 5 Department of Cell Biology,
University Medical Center Groningen, University of Groningen.
Background:
Progression of Atrial Fibrillation (AF), the most common persistent clinical tachyarrhythmia, is
driven by structural and metabolic remodeling of cardiomyocytes. Activation of the nuclear
enzyme poly (ADP-ribose) polymerase (PARP) is implicated in various forms of cardiomyopathies.
Excessive PARP activation reduces ATP formation by depletion of its substrate, nicotinamide
adenine dinucleotide (NAD+). To examine whether PARP activation contributes to AF progression,
we employed experimental model systems and clinical AF.
Methods and Results:
Cultured HL-1 cardiomyocytes and Drosophila prepupa were subjected to rapid electrical
stimulation (tachypacing) to mimic atrial fibrillation. Tachypacing of HL-1 atrial cardiomyocytes
impaired contractile function (calcium transients) and induced activation of PARP1, but not
PARP2, resulting in NAD+ depletion. Replenishment of NAD+ and administration of the PARP
inhibitors 3-aminobenzamide and ABT-888 attenuated tachypacing induced impairment of
contractility in HL-1 and Drosophila prepupae heart (heart wall contractions). Consistent with
these findings, patients with AF showed a significant activation of PARP as demonstrated by
increased levels of protein-parylation, both in left and right atrial biopsies.
Conclusions:
AF causes PARP1 activation, which subsequently results in depletion of NAD+ levels and
functional loss of cardiomyocytes. Since functional loss was prevented by PARP1 inhibitors, the
findings indicate a beneficial role for PARP1 inhibitors in clinical AF.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A103
In vivo pathophysiology of hypertrophic cardiomyopathy in a highly prevalent 25 bp deletion
mutation in MYBPC3
Diederik W.D. Kuster1,2, David Barefield,1 Suresh Govindan,1 Sivaguru Mayandi,3 Ben August,4
Kyounghwan Lee,5 Rajasekaran Namakkal-Soorappan,6 Roger Craig5 and Sakthivel Sadayappan1
1
2
3
Loyola University Chicago, Maywood, IL, USA; VU University Medical Center, Amsterdam; University of Illinois at
4
5
Urbana Champaign, Urbana, IL, USA; University of Wisconsin, Madison, WI, USA; University of Massachusetts
6
Medical School, Worcester, MA, USA; University of Alabama at Birmingham, Birmingham, AL, USA.
Background: Hypertrophic cardiomyopathy (HCM) is the most common inherited cardiac
disorder. The most prevalent HCM associated mutation, is a 25 base pair (bp) deletion in the
cardiac myosin binding protein-C (cMyBP-C) gene, that results in a modified C10 domain (cMyBPCC10MUT). This mutation has >55 million carriers worldwide. However, the mechanism by which
this mutation leads to cardiomyopathy is unknown. We hypothesized that cMyBP-CC10MUT
expression directly causes contractile dysfunction and leads to the development of HCM.
Methods and results: Expression of cMyBP-CC10MUT in cultured isolated adult rat cardiomyocytes
led to improper localization and contractile dysfunction. In vitro binding studies found that
cMyBP-CC10mut lost its binding affinity to the rod region of myosin heavy chain. To further
demonstrate that cMyBP-CC10MUT is sufficient to cause HCM and contractile dysfunction in vivo,
transgenic mice were generated in which the C10 domain of cMyBP-C was modified to cMyBPCC10MUT. Transgenic mice showed significant increase in heart weight/body weight ratio by 12
weeks of age (4.43±0.11 mg/g NTG, 5.07±0.30 mg/g C10mut), compared to non-transgenic
littermates. M-mode echocardiography analysis showed hypercontractile hearts (EF: 53.4%±2.9%
NTG, 66.4%±4.7% C10mut), but early diastolic dysfunction (E/E’: 28.7±3.7 NTG, 46.3±8.4
C10mut). Histopathological analyses demonstrated increased fibrosis in cMyBP-CC10mut hearts. In
skinned cardiomyocytes from cMyBP-CC10MUT mice preserved maximum force generation was
observed but an increased Ca2+-sensitivity of force generation, compared to controls (EC50:
3.37±0.01µM NTG, 2.90±0.01µM C10mut). Protein analysis showed an highly increased presence
of cMyBP-C in the soluble fraction in cMyBP-CC10mut mice (22.5±5.1 fold increase vs NTG), which
might be explain the diastolic impairment.
Conclusion: Expression of cMyBP-CC10MUT protein is sufficient to cause contractile dysfunction and
HCM in vivo. Results from this study demonstrated the pathophysiology of the cMyBP-CC10MUT,
the most common cardiomyopathy associated gene mutation.
Abstracts in order of the poster board numbering in ARTIS see page 7 for MAP and
page 8-13 for list of presenting authors.
A104
Electrophysiological effects of late sodium current inhibitor GS967 in Scn5a-1798insD mouse
and SCN5A-1795insD hiPSC-derived cardiomyocytes
Vincent Portero1, Simona Casini1, Maaike Hoekstra1, Arie O. Verkerk2, Isabella Mengarelli1,
Connie R. Bezzina1, Luiz Belardinelli3, Sridharan Rajamani3, Marieke W. Veldkamp1, Carol Ann
Remme1
1: Department of Experimental Cardiology, Academic Medical Center, Amsterdam. 2: Department of Anatomy,
Embryology and Physiology, Academic Medical Center, Amsterdam 3: Department of Biological Sciences, Gilead
Sciences, Fremont, CA.
Background and Aim:
Selective inhibition of cardiac late sodium current (INaL) is an emerging target in the treatment of
ventricular arrhythmias. The electrophysiological effects of GS967, a potent INa,L inhibitor, were
investigated in an SCN5A overlap syndrome model of both gain (LQT3) and loss of sodium
channel function (Brugada syndrome) comprising cardiomyocytes derived from human SCN5A1795insD induced pluripotent stem cells (hiPSC-CMs) and mice carrying the homologous
mutation Scn5a-1798insD.
Methods and Results:
In line with the disease phenotype, on patch-clamp analysis, isolated mouse Scn5a-1798insD
cardiomyocytes and SCN5A-1795insD hiPSC-CMs showed reduced peak INa and action potential
(AP) upstroke velocity (Vmax) and an increased AP duration at 90% repolarization (APD90)
associated to enhanced INaL, as compared to wild-type. In mouse Scn5a-1798insD cardiomyocytes
and in SCN5A-1795insD hiPSC-CMs, GS967 (300 nM) significantly decreased APD90 and Vmax, and
selectively inhibited INaL with minimal effect on peak INa. Furthermore, GS967 prevented proarrhythmic events in mouse Scn5a-1798insD cardiomyocytes. In Langendorff-perfused isolated
mouse Scn5a-1798insD hearts, GS967 (300 nM) had no effect on ventricular activation time or
conduction velocity (as assessed by epicardial mapping).
Conclusions:
Selective inhibition of INaL by GS967 attenuated AP prolongation in mouse Scn5a-1798insD
cardiomyocytes and in SCN5A-1795insD hiPSC-CMs, thus suppressing the gain-of-function
features of this overlap syndrome mutation. Importantly, these beneficial actions of GS967
occurred in the absence of deleterious effects on sodium channel availability or cardiac
conduction, despite a pre-existing decrease in peak INa. Thus, selective inhibition of INaL
constitutes a promising pharmacological treatment of cardiac channelopathies associated with
enhanced INaL, even in overlap syndromes whereby peak INa is decreased.
Program
Jolanda Van der Velden
Mat Daemen
Layout figures page 1 & 2
Karen Folkertsma
Layout abstracts
Isabelle Vergroesen