Download Interaction of engineered nanoparticles with endothelial cells an

Interaction of engineered
nanoparticles with endothelial cells
an experimental approach
Francesca Re
Department of Health Sciences
University of Milano-Bicocca
[email protected]
Erice 2015
VASCULAR WALL – NANOPARTICLES
INTERACTION
ACCIDENTAL EXPOSURE
Environmental/dietary exposure
REQUIRED INTERACTION
Biomedical use of NP
VASCULAR WALL – NANOPARTICLES
INTERACTION
PASSIVE
TARGETING
ACTIVE
TARGETING
ACCIDENTAL EXPOSURE
NP-endothelial cells = aspecific
TARGETING MEDICATION
NP-endothelial cells = driven
«EPR» effect
Enhanced Permeation-Retention effect
EXAMPLES OF EC TARGET MOLECULES
 E-selectin; P-selectin
 vascular cell adhesion molecule 1 (VCAM-1)
 vascular endothelial growth factor receptor
(VEGF-R)
 leukocyte adhesion molecules (LAMs)
 Low density lipoprotein receptor LDL-R
 Transferrin receptor (Tf-R)
FACTORS THAT INFLUENCE THE
INTERNALIZATION RATES OF NP BY EC
EXPERIMENTAL MODELS OF EC
HUVEC
hCMEC/D3
human umbilical vein
endothelial cells
human brain capillary
endothelial cells
 CELLULAR UPTAKE OF NP
 MECHANISMS OF NP INTERNALIZATION
 INTRACELLULAR FATE OF NP
 NP TOXICITY
NP FEATURES
and
CELLULAR UPTAKE
SURFACE MODIFICATIONS AFFECT
THE EC UPTAKE OF NP
GOLD NANOPARTICLES
(by ICP-MS)
Albumin-coated NP
LIPOSOMES
(by radiochemical techniques)
ApoE-coated LIPOSOMES
bare NP
BARE LIPOSOMES
Au NP
[Re F, in progress]
[Bana L, Nanomedicine 2014]
ADSORPTION-MEDIATED CELLULAR
UPTAKE OF NP
TAT-1 PEPTIDE
Derived from the HIV
+
--CATIONIC ALBUMIN
LIPOSOMES
[Sancini G, Nanomed Nanotech 2013]
TAT-LIPOSOMES
LIGAND DENSITY OF NP SURFACE
AFFECTS THE EC UPTAKE OF LIPOSOMES
BARE LIPOSOMES (A)
ApoE-FUNCTIONALIZED LIPOSOMES (B-E)
[Re F, J Biotechnol 2010]
PLASMA PROTEINS-MEDIATED CELLULAR
UPTAKE OF NP
«PROTEIN
CORONA»
NPs in contact with biological fluids are rapidly covered by a selected
group of biomolecules (proteins, lipids) to form a corona
PLASMA PROTEINS-MEDIATED CELLULAR
UPTAKE OF GOLD NP
QUANTITATIVE-DEPENDENT
(by ICP-MS)
QUALITATIVE-DEPENDENT
(by SDS-PAGE)
All-MUS Striped NP
[Re F, in progress]
NP FEATURES
and
MECHANISMS OF INTERNALIZATION
NP ARE UPTAKEN BY EC SIMULTANEOUSLY
WITH DIFFERENT MECHANISMS
1-100 mm
< 120 nm
< 60 nm
< 90 nm
< 500 nm
RECEPTOR-MEDIATED CELLULAR UPTAKE
OF ApoE-LIPOSOMES
NP CELLULAR UPTAKE
LIPOSOMES
(by radiochemical techniques)
+ COMPETITOR FOR LDL-R
[Bana L, Nanomedicine 2014]
CLATHRIN-DEPENDENT CELLULAR
UPTAKE OF ApoE-LIPOSOMES
Cellular uptake
 Dynasore (Dyn) a small molecule inhibitor
+ ENDOCYTOSIS
INHIBITORS
of dynamin
 Chlorpromazine (CPZ) a cationic
amphipathic molecule that inhibits the
formation of clathrin-coated pits
[Gregori M, in progress]
NP FEATURES
and
INTRACELLULAR FATE
INTRACELLULAR FATE OF NP
ENDOSOMAL ESCAPE
Multi-vesicular bodies
Recycling
microtubule
TRANSCYTOSIS
FUNCTIONALIZED ApoE-LIPOSOMES DO NOT
CO-LOCALIZE WITH ENDOSOMES
RED = ENDO-LYSOSOMES
GREEN = NANOPARTICLES
[Sancini G, Nanomed Nanotech 2013]
TRANSCYTOSIS OF NANOPARTICLES
CELLULAR UPTAKE
(by ICP-MS)
All-MUS
Striped
TRANSCYTOSIS
(by ICP-MS)
All-MUS
Striped
[Re F, in progress]
NP FEATURES
and
TOXICITY
ApoE-LIPOSOMES TOXICITY ON EC
assessed by
MTT ASSAY
LDH RELEASE
NITRIC OXIDE PRODUCTION
NO TOXICITY WAS DETECTED
(LIPOSOMES CONCENTRATIONS 6-120 ug/ml)
C
apoptosis
**
cnt
Caspase-3
β-Actin
100
EFFECT ON NITRIC OXIDE AND ROS PRODUCTION
Zhu et al., 2011
apoptosis
GUIDELINES FOR NANOTOXICITY EVALUATION
Soenen et al, 2011
1. NANOPARTICLES DESIGNED FOR BIOMEDICAL USES
Delivery of drugs and/or imaging contrast agents
TARGETING THE VASCULAR WALL
PASSIVE
TARGETING
ACTIVE
TARGETING
VASCULAR WALL – NANOPARTICLES
INTERACTION
ACCIDENTAL EXPOSURE
NP-endothelial cells = aspecific
TARGETING MEDICATION
NP-endothelial cells = driven