Download ARVO 2015 Annual Meeting Abstracts 322 Ocular surface health

ARVO 2015 Annual Meeting Abstracts
322 Ocular surface health and disease
Tuesday, May 05, 2015 8:30 AM–10:15 AM
Exhibit Hall Poster Session
Program #/Board # Range: 3021–3077/D0217–D0273
Organizing Section: Cornea
Contributing Section(s): Genetics, Multidisciplinary Ophthalmic
Imaging
Program Number: 3021 Poster Board Number: D0217
Presentation Time: 8:30 AM–10:15 AM
TGFβ signaling inhibits goblet cell differentiation via SPDEF in
conjunctival epithelium
Chia-Yang Liu, Yujin Zhang. Ophthalmology, Univ of Cincinnati,
Cincinnati, OH.
Purpose: To demonstrate the signaling transduction of
TGFbRIIÇSmadsSPDEFÇMuc5/ac axis for conjunctival goblet cell
differentiation.
Methods: We employed conditional deletion of transforming growth
factor β receptor II (TGFβRII) in keratin 14-positive stratified
epithelia. Phenotypic and biochemical analyses of the resultant ocular
surface were compared to the wild-type littermates at various stages.
Results: We found that, in the absence of an external phenotype,
the ocular surface epithelium develops properly, but young mice
displayed conjunctival goblet cell expansion, demonstrating that
TGFβ signaling is required for normal restriction of goblet cells
within the conjunctiva. We observed increased expression of
SAM-pointed domain containing ETS transcription factor (SPDEF)
in stratified conjunctival epithelial cells in TGFβRII cKO mice,
suggesting that TGFβ restricted goblet cell differentiation directly by
repressing Spdef transcription. Gain-of-function of Spdef in keratin
14-derived epithelia resulted in the ectopic formation of goblet
cells in the eyelid and peripheral cornea in adult mice. We found
that Smad3 bound two distinct sites on the Spdef promoter and that
treatment of keratin-14 derived cells with TGFβ inhibited SPDEF
activation, thereby identifying a novel mechanistic role for TGFβ in
regulating goblet cell differentiation.
Conclusions: TGFbRII-Smads is the upstream regulator of Spdef
during conjunctival goblet cell differentiation.
Commercial Relationships: Chia-Yang Liu, None; Yujin Zhang,
None
Support: NIH EY21501; Research Prevent Blindness
Program Number: 3022 Poster Board Number: D0218
Presentation Time: 8:30 AM–10:15 AM
The effect of dry eye therapy on the symptoms and signs of
ptergia
Maryam Yamani1, 2, Raymond Fong1, Andrew Petti2, Matthew Petti2.
1
Raymond Fong MD, Flushing, NY; 2A Yamani MD PC, Mount
Vernon, NY.
Purpose: To evaluate the additive effect of dry eye therapy,
specifically punctal plug placement and artificial tears in the relief
of symptoms (pain, itching, foreign body sensation and tearing) and
signs (redness and punctate epithelial keratitis) of pterygia in patients
who are already being treated with non-steroidal anti-inflammatory
drops.
Methods: Twenty-two patients with pterygia were seen between May
and October of 2014. They were asked to rate the level of discomfort
in regards to the symptoms of pain, itching, foreign body sensation
(FBS) and tearing. This was done on the first visit, after a week of
non-steroidal anti-inflammatory (NSAID) drops, and finally, after at
least a week of either punctal plug placement or artificial tear therapy.
The signs of redness and punctate epithelial keratitis (PEK) were
also rated by the same physician on all visits. The result were then
analyzed.
Results: NSAID drops had no statistically significant effect on either
the symptoms or signs studied. Adding dry eye therapy to the course
of treatment significantly decreased the severity of the symptoms of
FBS (p<.05), tearing (p<.05), PEK (p<.05), and the sign of redness
(p<.05). Punctal plug placement decreased the level of discomfort
with regards to itching (p<.05), FBS (p<.05), and tearing (p<.05) as
well as redness and PEK (p<.05). Artificial tear therapy improved
tearing (p<.05) and PEK (p<.05).
Conclusions: Non-steroidal anti-inflammatory drops had no
significant effect on either the symptoms or signs of pterygia. The
addition of dry eye therapy relieved both the symptoms and signs of
pterygia with punctal plug placement being more efficacious than
artificial tear therapy.
Commercial Relationships: Maryam Yamani, None; Raymond
Fong, None; Andrew Petti, None; Matthew Petti, None
Program Number: 3023 Poster Board Number: D0219
Presentation Time: 8:30 AM–10:15 AM
Gene expression pattern comparison between pterygium and
adipose cells
Jesus Lozano1, Judith Zavala1, Victor Treviño2, Diana Vazquez1,
Pedro Romero1, Jorge E. Valdez1. 1Ophthalmology, Tecnológico
de Monterrey, Monterrey, Mexico; 2Cátedra de Bioinformática,
Tecnológico de Monterrey, Monterrey, Mexico.
Purpose: Pterygium cells are know to exhibit altered lipid
metabolism characteristics. Some of the disregulated molecules in
pterygium tissue related to inflammatory and proliferative process
are mainly expressed in adipose tissue. In this study we compared
the gene expression pattern between adipose and pterygium cells in
order to stablish the usefulness of adipose cells in the research of new
therapeutic targets for pterygium.
Methods: Eleven samples of adipose cells and eight of pterygium
cells ran with the same platform were obtained from GEO database.
The average of difference expression between samples was calculated
and a list of 535 genes with significant difference (P value above
0.01) was obtained. An over-representation analysis was made
in order to elucidate related gene ontology, pathways and protein
interactions.
Results: Among the 535 significant different genes, the nuclear
transcription factor X-box binding 1 (P value = 7.5x10-2), present in
pterygium cells, was related to a transcriptional repressor capable of
regulating the duration of an inflammatory response. LY6/PLAUR
domain containing 3 (P value = 1.1x10-13), related to cell migration
and tumor progression, was present in both cell types. CUL7 (P value
= 3.8x10-2) gene was related to the E3 ubiquitin ligase, an ubiquitinproteasome system, was only present in pterygium cells. Similarly,
ENTPD4 (P value = 1.0x10-2), was only present in pterygium cells
and was related to the hydrolysis of nucleosides on its components.
SOD1 (P value = 4.2x10-2) presence was diminished in pterygium
cells. This enzyme is related to catalysis process in the dismutation
of the toxic superoxide radical. On the other hand, SEC14L3 (P value
= 6.3x10-4), related to a phosphatidylinositol transfer essential for
biogenesis of golgi-derived transport vesicles, was only present in
adipose cells.
Conclusions: We found marked differences between the gene
expression pattern of adipocytes and pterygium cells, mainly in
genes related to inflammation and ubiquitination processes. However
adipocytes and pterygium cells expressed common genes related
to proliferation and migration processes. This analysis provides
a general idea of the genetic pattern similatities and differences
between the two cell types. Further studies with more samples will
©2015, Copyright by the Association for Research in Vision and Ophthalmology, Inc., all rights reserved. Go to iovs.org to access the version of record. For permission
to reproduce any abstract, contact the ARVO Office at [email protected].
ARVO 2015 Annual Meeting Abstracts
elucidate specific adipocyte molecules useful for the research of new
therapeutic targets for pterygium.
Commercial Relationships: Jesus Lozano, None; Judith Zavala,
None; Victor Treviño, None; Diana Vazquez, None; Pedro
Romero, None; Jorge E. Valdez, None
Program Number: 3024 Poster Board Number: D0220
Presentation Time: 8:30 AM–10:15 AM
Eye Whitening Using Subconjunctival Injections
Michael Campion1, Robert J. Campion MD2, Vincent M. Campion3.
1
Ophthalmology, Southwestern Eye Center, Mesa, AZ; 2St. Bernards
Medical Center, Jonesboro, AR; 3Ross University School of
Medicine, Miramar, FL.
Purpose: Recent reports have alerted eye care providers and
the general public about the danger of certain eye whitening
procedures that involve a combination of removing generous areas
of conjunctiva, cautery, laser and/or using antimetabolites. This
study explores the possibility of an alternative procedure involving
subconjunctival injections which could be used to whiten the eyes
and avoid the risks associated with previously described techniques.
Methods: Cadaver ocular specimens were obtained from a
reputable medical animal supplier. Tattoo ink was obtained from
a commercially available source. Photography was used for
documentation. The subconjunctival space was entered using a 25
gauge needle and balanced salt solution was injected to balloon up
the conjunctiva. The ink was then injected into the subconjunctival
space and spread throughout with external pressure applied with a
cotton tip applicator. Eyes were observed for six months. A similar
procedure was performed on live mice and rabbits. Attention was
paid to the appearance, homogeneity, longevity and tolerance of
the ink. Lastly the procedure was performed on a human subject.
Attention was directed to possible side effects. All live eyes were
given postoperative topical antibiotics.
Results: All cadaver eyes accommodated the ink in the
subconjunctival space. The ink could be easily spread throughout the
eye in a homogeneous fashion. The ink remained visibly apparent
throughout the study period of six months. Live animal models
demonstrated that the ink was tolerated by the eye with no signs of
inflammation. The ink would cover up the deeper scleral vessels but
the superficial conjunctival vessels were still visible. The effect of
the procedure remained during the entire study period of six months.
The human subject reported mild transient tearing, light sensitivity
and seepage of ink during the first five days but it quickly abated after
that time period. Visual acuity, intraocular pressure, cell and flare,
lack of corneal staining, and lid position were unchanged throughout
the study period. The appearance of the human eyes post operatively
was consistent for 24 months.
Conclusions: This cadaveric, live animal and human study
demonstrates the possibility of utilizing subconjunctival injections as
a means to whiten eye appearance and avoid the risks associated with
previously used techniques.
Commercial Relationships: Michael Campion, 8,382,726 (P);
Robert J. Campion MD, None; Vincent M. Campion, None
Program Number: 3025 Poster Board Number: D0221
Presentation Time: 8:30 AM–10:15 AM
The role of extracellular matrix (ECM) protein composition on
the adhesion and growth of conjunctival epithelium
ARUNI MAKULOLUWA, Rachel Williams, Stephen B. Kaye,
Rosalind M. Stewart. EYE AND VISION SCIENCE DEPARTMENT,
UNIVERSITY OF LIVERPOOL, Liverpool, United Kingdom.
Purpose: There is a need to understand how the ECM may affect cell
behaviour. This will guide development of substrates for conjunctival
reconstruction. We carried out a laboratory study to establish the
ECM deposited by conjunctival cells and determine the behaviour
of these cells when seeded onto pre-adsorbed proteins modelling the
natural ECM composition.
Methods: ECM proteins deposited by a conjunctival cell line
(HCjE-Gi) were assessed by immunofluorescence (IF) at days
5-42. Maximum adsorption of fibronectin, collagen IV and laminin
111 onto tissue culture plastic (TCP) was determined by ELISA.
Adhesion and growth of HCjE-Gi cells seeded onto uncoated TCP
and pre-adsorbed proteins up to the saturation points were assessed
by DAPI staining. HCjE-Gi-secreted ECM was isolated using 1%
NH4OH to remove the cells at days 5-42. Cellular growth on this
secreted ECM was determined until 7 days by DAPI staining and
counted using Image J. Statistical analysis was performed using
1-way ANOVA and independent t-tests.
Results: Laminin α3 was present abundantly from day 5 onwards,
fibronectin was present earlier but reduced with time. Collagen IV
was only detected from day 21. Maximum adsorption of fibronectin
and collagen IV onto TCP were reached from a 5μg/mL solution
and laminin 111 from a 0.5μg/mL solution. Adhesion of cells was
increased by pre-adsorption of fibronectin (p<0.05) and collagen IV
(p<0.05), however, this was unaffected by pre-adsorption of laminin
111. Cellular growth was not affected by pre-adsorption of fibronectin
or laminin 111, but was increased by both pre-adsorption of collagen
IV (p<0.05), or ECM secreted (p<0.05) for greater than 5 days.
Conclusions: The presence of secreted ECM or collagen IV alone
enhances the growth of conjunctival cells. Substrates or substrate
coating with these components could enhance ex-vivo expansion of
conjunctival cells for ocular surface reconstruction.
Commercial Relationships: ARUNI MAKULOLUWA, None;
Rachel Williams, None; Stephen B. Kaye, None; Rosalind M.
Stewart, None
Program Number: 3026 Poster Board Number: D0222
Presentation Time: 8:30 AM–10:15 AM
Viral double-strand RNA strengthens the barrier function in
immortalized conjunctival epithelium
Yuriko Ban1, 3, Rie Oda2, Shigeru Kinoshita3. 1Nantan General
Hospital, Nantan, Japan; 2Faculty of Life and Medical Sciences,
Doshisha University, Kyotanabe, Japan; 3Department of
Ophthalmology, Kyoto Prefectural University of Medicine, Kyoto,
Japan.
Purpose: To investigate the response to polyinosinic:polycytidylic
acid [Poly(I:C)], an analog of viral double-stranded RNA produced
during viral replication in the barrier function of immortalized
conjunctival epithelium.
Methods: Immortalized conjunctival epithelial cells were cultured
on 12-mm Transwell filters at a density of 4x105 cells/cm2. The
cultured cells were then stimulated with 2.5mg/ml, 25mg/ml, and
250mg/ml of Poly(I:C). Transepithelial electrical resistance (TER)
was then measured using endohm electrodes (World Precision
Instruments, LTD., Hertfordshire, UK). After 3 hours of exposure
to Poly(I:C), the expressions of tight junction-related protein ZO-1,
occludin, and claudin-1,-4, and -7 were analyzed by Western blotting.
Immunoreactive bands were visualized by chemiluminescence, and
densitometry analysis was then performed.
Results: After 3 hours of exposure to Poly(I:C), TER was increased
in a dose-dependent manner (control: 51.6±6.9Ωcm2, 2.5mg/ml
stimulation: 64.4±2.9Ωcm2, 25mg/ml stimulation: 114.9±9.6Ωcm2,
and 250mg/ml stimulation: 309.2±34.7Ωcm2). Poly(I:C) challenge
also increased the TER in a time-dependent manner (0hour:
53.9±1.3Ωcm2, 1hour: 119.4±7.4Ωcm2,: 2hours 194.4±14.7Ωcm2,
and 3hours: 241.8±19.9Ωcm2 by 25mg/ml stimulation). Claudin-4
©2015, Copyright by the Association for Research in Vision and Ophthalmology, Inc., all rights reserved. Go to iovs.org to access the version of record. For permission
to reproduce any abstract, contact the ARVO Office at [email protected].
ARVO 2015 Annual Meeting Abstracts
expression was decreased, yet no change was observed in the other
tight junction-related proteins.
Conclusions: The findings of this study show that Poly(I:C)
challenge, which mimics a viral infection, increased the barrier
function of ocular surface epithelia, and decreased claudin-4
expression due to a strengthening of the barrier. Thus, we theorize
that the increased barrier function must be a kind of defense reaction
to viral infection.
Commercial Relationships: Yuriko Ban, None; Rie Oda, None;
Shigeru Kinoshita, None
Program Number: 3027 Poster Board Number: D0223
Presentation Time: 8:30 AM–10:15 AM
Evaluation of redox status in human conjunctival epithelial cells
exposed to diesel exhaust particles (DEP)
Romina M. Lasagni Vitar1, Julia Tau2, Claudia G. Reides1, Alejandro
Berra2, Sandra M. Ferreira1, Susana Llesuy1. 1General and Inorganic
Chemistry Division, IBIMOL (UBA-CONICET), Faculty of
Pharmacy and Biochemistry, University of Buenos Aires, Buenos
Aires city, Argentina; 2Ocular Investigation Laboratory, Pathology
Department, Faculty of Medicine, University of Buenos Aires,
Buenos Aires city, Argentina.
Purpose: Environmental pollution causes adverse effects on human
health and the eyes are particularly vulnerable to these effects, as
they are constantly exposed to the environment. The aim of the
present study was to evaluate the redox status in human conjunctival
epithelial cells after the incubation with diesel exhaust particles
(DEP).
Methods: The incubation was performed at different concentration of
DEP (10, 50 and 100 μg/mL) for 24 hs and the following parameters
were evaluated: reactive oxygen and nitrogen species production;
protein oxidation; antioxidant enzymes activities (superoxide
dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx),
glutathione S-transferase (GST) and glutathione reductase (GR));
non-enzymatic antioxidant levels measured as total antioxidant
capacity (TRAP) and reduced (GSH) and oxidized (GSSG)
glutathione. One-way ANOVA test and Dunnett’s test as a post hoc
test were used for statistical analysis.
Results: The cells exposed to 50 and 100 μg/mL of DEP showed
a significant increase in reactive oxygen species production (24%,
p<0.01 and 45%, p<0.001, respectively) and in reactive nitrogen
species production (257%, p<0.01 and 514%, p<0.001, respectively)
compared to the control group. They also exhibited an increase in the
activities of SOD (84% and 108%, p<0.001, respectively), GPx (41%
and 50%, p<0.05 and p<0.01, respectively) and GST (42% and 45%,
p<0.01, respectively). A decay in GR activity was observed (17%
and 37%, p<0.05 and p<0.01, respectively), meanwhile CAT levels
remained unchanged. DEP100 group also presented an increase in
protein oxidation (58%, p<0.05). Furthermore, TRAP was significant
reduced in both groups (42%, p<0.05 and 58%, p<0.01, respectively)
as well as the GSH/GSSG ratio. There was no significant difference
between DEP10 and control groups in all the measurements.
Conclusions: The increase of reactive oxygen and nitrogen species
production, the decrease in non-enzymatic antioxidants and the
increase in the antioxidant enzymes activities are consequences of
the exposure to DEP. These results suggest that the alteration of
redox status could be a possible mechanism of damage in the human
conjunctival epithelial cells exposed to this particular matter.
Commercial Relationships: Romina M. Lasagni Vitar, None;
Julia Tau, None; Claudia G. Reides, None; Alejandro Berra,
None; Sandra M. Ferreira, None; Susana Llesuy, None
Support: University of Buenos Aires Grant 01/796BA
Program Number: 3028 Poster Board Number: D0224
Presentation Time: 8:30 AM–10:15 AM
Congenital conjunctival choristomas
Kelley J. Bohm, Gary J. Lelli, Ana G. Alzaga Fernandez, Jessica B.
Ciralsky. Ophthalmology, Weill Cornell Medical College, New York,
NY.
Purpose: To report the outcomes of our unique surgical technique for
removal of congenital conjunctival choristomas.
Methods: Retrospective case series of seven consecutive patients
who underwent congenital conjunctival choristoma removal at our
institution and who had at least 1 month of follow-up care. In all
cases, a conjunctival sparing technique was used. For extensive
lesions, only the visible portion was excised. All cases were
performed in conjunction with an oculoplastic surgeon to help
preserve vital anatomy, including rectus muscles, lacrimal glands,
and eyelid structures. Outcome measures included visual acuity,
complications, and cosmetic appearance, as assessed by photography
and reported satisfaction (parent and patient (when applicable)).
Results: Seven eyes of seven subjects, aged 13 months to 28 years,
underwent surgical removal of a congenital conjunctival choristoma
at our institution. Of these lesions, six involved a portion of the
cornea and limbus, two involved the lateral canthus, and four were
located in the area of a rectus muscle. Three patients had Goldenhar’s
syndrome. For patients old enough to undergo visual acuity testing,
average pre-operative and most recent visual acuity measurements
were 0.46 and 0.29, respectively. The average change in visual
acuity revealed an improvement of -0.17 logMAR. All subjects
(patients and/or parents) reported cosmetic improvement. No serious
complications were noted.
Conclusions: In all patients in our series, our surgical technique
preserved visual acuity and improved the cosmetic appearance of the
eye. Furthermore, there were no serious complications in any case.
Commercial Relationships: Kelley J. Bohm, None; Gary J. Lelli,
None; Ana G. Alzaga Fernandez, None; Jessica B. Ciralsky, None
Program Number: 3029 Poster Board Number: D0225
Presentation Time: 8:30 AM–10:15 AM
Shifting and sliding of medial-inferior conjunctival flap for
surgical treatment of Pterygium
Ignazio Zucca, Mariella Ulleri, Alberto Cuccu, Stefania Piludu,
Maria Silvana Galantuomo, Maurizio Fossarello. Ophthalmology,
University of Cagliari, Cagliari, Italy.
Purpose: To describe a novel surgery technique in the treatment of
primary pterygium and to evaluate its safety and rate of recurrence.
Methods: Retrospective case series. Between January 2012 and
October 2013, escluded summer months, 128 patients, of which 103
were men and 25 women, all living in Sardinia (Italy) were operated
on for a primary nasal pterygium. The mean age was 55,61 ± 9,93
(range 33-75). Pterygium excision was combined with a sliding and
shifting congiuntival flap technique. The flap harvested from the
medial inferior bulbar conjunctiva was placed over the bare sclera
and glued with Tisseel (Baxter AG, Vienna), a human tissue glue.
Topical and oral steroids and antibiotic eyedrops were prescribed, and
the eye was patched for 1 day.
Results: Patients were followed up for at least 1 year, and
the outcomes and recurrence rates were evaluated. In case of
fibrovascular regrowth, it was manifested by third month of follow
up, but progression arrested before sixth month control. Six patients
showed a corneal invasion between 1 and 2.5 mm beyond the limbus,
in 1 patient the new fibrovascular tissue was 4.5 mm . The recurrence
rate estimated at 1 year after surgery was 5.46%. No intraoperative
complication was observed. The mean surgery time was 10.30’’ ±
1.30” minutes.
©2015, Copyright by the Association for Research in Vision and Ophthalmology, Inc., all rights reserved. Go to iovs.org to access the version of record. For permission
to reproduce any abstract, contact the ARVO Office at [email protected].
ARVO 2015 Annual Meeting Abstracts
Conclusions: The sliding and shifting congiuntival flap technique is
a safe alternative in pterygium surgery for preventing recurrence. It is
technically easier than congiuntival autograft and reduces operative
time. Moreover the low rate of recurrence could be positively
influenced by the reduced inflammatory reaction due to the absence
of a congiuntival suture.
Commercial Relationships: Ignazio Zucca, None; Mariella Ulleri,
None; Alberto Cuccu, None; Stefania Piludu, None; Maria Silvana
Galantuomo, None; Maurizio Fossarello, None
Program Number: 3030 Poster Board Number: D0226
Presentation Time: 8:30 AM–10:15 AM
Clinical analysis for subconjunctival hemorrhage
Hiroaki Ozaki, Jane Y. Huang, Eiichi Uchio. Ophthalmology,
Fukuoka Univ Sch of Medicine, Jyonan-ku, Japan.
Purpose: Subconjunctival hemorrhage (SCH) is a relatively common
disease which is characterized by an acute onset of bleeding under
the bulbar conjunctiva. However, the etiology and the cause are not
defined. We have investigated the age, gender, incidence, seasonal
occurrence, location, extent, and medical history for patients with
SCH.
Methods: The study was designed as prospective from April 2011
to March 2013. A total number of the outpatients was 69058 in the
study. A total of 743 patients with SCH aged 1–91 years was enrolled.
There were 286 men and 457 women. The average age was 57.9
± 20.4 years old. The excluding criteria was SCH associated with
ocular injections and/or ocular surgeries. The age, gender, incidence,
seasonal occurrence, location, extent and medical history of the
patients were determined.
Results: The incidence of SCH outpatients was 1.1%(743/69058).
SCH was more common in female, and the male to female ratio
of SCH was 2:3. SCH was found on the right eye in 355 patients,
on the left eye in 388 patients and on the both eyes in 9 patients.
Seasonal variation was not statistically significant. The incidence
of SCH showed an increase with age. The peak onset of SCH was
between 71 and 80 years. The extent of SCH was more commonly
seen within one quadrant around corneal limbs(58.5%). About
23.5% of eyes had involvement of two quadrants to entire conjuncti
bulbi. The main related factors for SCH were hypertension (24.3%),
heperlipemia(15.1%) and diabetes(9.6%). Idiopathic SCH was found
in 345 patients(48.5%). The extent of SCH had no correlation with
age or medical history.
Conclusions: SCH is more commonly seen in female and in the
temporal side of the conjunctiva. SCH was commonly seen between
71 and 80 years of age and the incidence increased with age. The
extension of SCH was not related to the medical history. Our results
suggest that SCH could be an age-related disease.
Commercial Relationships: Hiroaki Ozaki, None; Jane Y. Huang,
None; Eiichi Uchio, None
Program Number: 3031 Poster Board Number: D0227
Presentation Time: 8:30 AM–10:15 AM
Ophthalmia Neonatorum (ON) in Angola: Results of the first
attempt to develop a prophylaxis program
Jose-Carlos Pastor1, Isabel Alexandre2, Mar Justel3, Prudencio
Martinez1, Raul Ortiz de Lejarazu3. 1IOBA-Campus Miguel Delibes,
University of Valladolid, Valladolid, Spain; 2IONA, Luanda, Angola;
3
Department of Microbiology and Immunology, Hospital Clinico
Universitario de Valladolid, Valladolid, Spain.
Purpose: To study the efficacy of povidone-iodine (P-I) prophylaxis
for ophthalmia neonatorum (ON) in Angola and to document the
maternal prevalence and mother-to-child transmission rates of three
important infectious agents
Methods: After ethical approval by Agostinho Neto University
committee the study was conducted at General Augusto N’Gangula
Specialized Hospital and Health Center of Samba, both in Luanda,
Angola from 7-Dec-2011 to 22-Nov-2012. Inclusion criteria
consisted of healthy children weighing at least 2.3 kg and a gestation
period of at least 37 weeks. Endocervical samples from mothers
(n=312) and newborn conjunctival smears (n=245) were analyzed
by multiplex polymerase chain reaction (PCR) for Chlamydia
trachomatis (CT), Neisseria gonorrhoeae (NG), and Mycoplasma
genitalium (MG). After ophthalmological examination, the newborns
were randomized into a non-interventional group (no local treatment)
and an interventional group that received a drop of P-I 2.5%
bilaterally after conjunctival smear collection. Mothers were trained
to identify signs of ON and attend a follow-up visit 7-10 days after
discharge
Results: Forty-four (17.9%) newborns had pathologic ocular
findings, and 11 (4.5%) had clinical signs consistent with ON at the
time of delivery. Maternal PCR was positive for MG (n=19), CT
(n=8), and NG (n=2) (one mother had two microorganisms: CT and
MG).. Eight newborns were positive for CT (n=4), MG (n=3), and
NG (n=1). Mother-to-child transmission rates were 50% for CT and
NG and 10.5% for MG. Ten of the positive mothers had risk factors
for M-C-T. No relationship could be established between external
signs of acute conjunctivitis and maternal genito-urinary infections.
Only 16 of the newborns were returned for a follow-up visit
Conclusions: The lack of maternal compliance prevented the
successful testing of prophylactic P-I efficacy in ON prevention.
Nevertheless, we documented the prevalence and mother-to-child
rates for CT, NG, and MG. These results emphasize the need to
educate the Angolan public on the origins of ON and to develop an
effective prophylaxis program
Commercial Relationships: Jose-Carlos Pastor, None; Isabel
Alexandre, None; Mar Justel, None; Prudencio Martinez, None;
Raul Ortiz de Lejarazu, None
Support: Agencia Española de Cooperación Internacional y
Desarrollo. Ministerio de Asuntos Exteriores.
Program Number: 3032 Poster Board Number: D0228
Presentation Time: 8:30 AM–10:15 AM
Risk of Recurrence and Surgical Complications of Pterygium
Surgery performed by Ophthalmologists-in-Training
Gabriel Luna, Priya Janardhana, Wuqaas Munir, Vanessa Vazquez,
Arvind Devanabanda. Ophthalmology, Boston University Medical
Center, Boston, MA.
Purpose: To investigate the surgical outcomes of primary pterygium
surgery performed at Boston University Medical Center by
ophthalmology residents, under the supervision of a single attending.
Methods: Retrospective chart review, from January 2006-November
2012, in which 66 patients qualified. Statistical analysis was
performed using SAS 9.1.3 (SAS Institute Inc).
Results: There were no infections noted post-operatively and one
pterygium recurrence at 3 months post-operatively, associated with
noncompliance of prednisolone acetate 1% eye drops. We also found
that there is no correlation with surgical experience and surgical
length of time of pterygium surgery performed by ophthalmology
residents.
Conclusions: Surgical outcomes of pterygium surgery performed by
residents have a low risk of recurrence and complications.
Commercial Relationships: Gabriel Luna, None; Priya
Janardhana, None; Wuqaas Munir, None; Vanessa Vazquez,
None; Arvind Devanabanda, None
©2015, Copyright by the Association for Research in Vision and Ophthalmology, Inc., all rights reserved. Go to iovs.org to access the version of record. For permission
to reproduce any abstract, contact the ARVO Office at [email protected].
ARVO 2015 Annual Meeting Abstracts
Program Number: 3033 Poster Board Number: D0229
Presentation Time: 8:30 AM–10:15 AM
Conjunctiva fixation to the posterior sclera for the treatment of
conjunctivochalasis
Ari Shinojima, Mitsuko Yuzawa. Ophthalmology, Surugadai Nihon
University Hospital, Tokyo, Japan.
Purpose: There are reports describing operative techniques for
conjunctivochalasis with and without resection of the conjunctiva.
If there is excessive conjunctival tissue, it is difficult to attach 8-0
Vicryl® thread (Johnson & Johnson, NJ, USA) directly at the site
8 mm posterior to the corneal limbus, as in the previously reported
method (Otaka and Kyu, Am J Ophthalmol, 2000). Therefore, we
endeavored to develop a simple non-resection surgical technique for
conjunctivochalasis with simultaneous conjunctival tacking.
Methods: Patients over 60 years of age who had been diagnosed with
conjunctivochalasis were enrolled in this study. The mean age (±SD)
of our patients was 77.1 ± 5.9 years (range, 64–83 years). In total, 23
eyes of 14 patients (3 men, 5 eyes; 11 women, 18 eyes) underwent
operative treatment of conjunctivochalasis. Topical anesthetic eye
drops including 0.4% oxibuprocaine and 4% lidocaine were instilled.
The lower bulbar conjunctiva was initially scooped 4mm posterior to
the corneal limbus, and stitches were then fixed with careful tacking
8 mm posterior to the corneal limbus, in order to avoid restriction of
the lower gaze. We used 8-0 Vicryl® for the stiches, allowing focal
inflammation and firm attachment of the fornical conjunctiva to the
globe. The mean follow-up period (±SD) was 234.4 ± 179.9 days
(range, 55– 536 days).
Results: The tear meniscus was maintained normal in all eyes
(n=23). No recurrence of conjunctivochalasis was observed during
the follow-up period.
Conclusions: Our operative method was successful and there were
no recurrences during follow-up.
Commercial Relationships: Ari Shinojima, None; Mitsuko
Yuzawa, None
Program Number: 3034 Poster Board Number: D0230
Presentation Time: 8:30 AM–10:15 AM
Inhibition of pterygium fibroblasts proliferation by Siempre viva
plant (Sedum dendroideum) extracts
Judith Zavala1, Pedro Romero1, Diana Vazquez1, Bertha Barba1,
Jorge Valdez1, Jorge E. Valdez1, 2. 1Ophthalmology Research
Chair, Tecnologico de Monterrey, Monterrey, Mexico; 2Tec Salud,
Tecnologico de Monterrey, Monterrey, Mexico.
Purpose: The only treatment available for pterygium is surgery,
however this procedure faces high recurrence rates. Sedum
dendroideum is an endemic mexican plant known to possess
high concentrations of flavonoids. We tested the effect of Sedum
dendroideum extracts on NIH3T3 and pterygium fibroblasts
proliferation rate.
Methods: Pterygium fibroblasts obtained from human surgically
removed samples and NIH3T3 fibroblasts were cultivated with
DMEM-F12 media supplemented with 0.2% DMSO. S. dendroideum
extracts (methanolic, chloroformic, hexanic, hexane/acetone, and
ether) were obtained from dried leaves and stems. Viability assays
were performed by triplicate 2 h and 24 h after adding 500, 1000,
1500, and 2000 mg/mL of each extract to NIH3T3 fibroblasts
cultures. The extract that demonstrated higher proliferation inhibitory
activity was tested on pterygium fibroblasts at same time points and
concentrations.
Results: Ether, hexane, and hexane/acetone extracts showed the
highest anti-proliferative activity in NIH3T3 fibroblasts. After 2h,
ether extract showed an LD50 of 1550 mg/mL, while hexane/acetone
showed an LD50 of 2000 mg/mL. After 24 h, ether exhibited an LD50
of 545 mg/mL, hexane of 476 mg/mL, and hexane/acetone of 815 mg/
mL. Given that there was no significant difference between ether
and hexanic extract activity in NIH3T3 fibroblasts at 24 h (p value =
0.324), ether extract was chosen to be tested in pterygium fibroblasts.
This extract showed an LD50 of 1600 mg/mL after 2h, and of 807 mg/
mL after 24 h.
Conclusions: Ether extracts from S. dendroideum plant inhibit the
proliferation of NIH3T3 and pterygium fibroblasts. Further analysis
of the effect of ether extract on levels of molecules related to
pterygium pathogenesis along with the characterization of the extract
will provide more information about the potential of this Mexican
endemic plant as therapeutic approach for pterygium treatment.
Commercial Relationships: Judith Zavala, Tecnologico de
Monterrey (P); Pedro Romero, Tecnologico de Monterrey (P);
Diana Vazquez, Tecnologico de Monterrey (P); Bertha Barba,
Tecnologico de Monterrey (P); Jorge Valdez, Tecnologico de
Monterrey (P); Jorge E. Valdez, Tecnologico de Monterrey (P)
Program Number: 3035 Poster Board Number: D0231
Presentation Time: 8:30 AM–10:15 AM
High level of carbonic anhydrase 1 in pterygium
Kaili Wu, Qian Yang, Xiuping Liu, Liping Lin, Feng Zou. Zhongshan
Ophthalmic Center, State Key Laboratory of Ophthalmology, Sun
Yat-sen University, Guangzhou, China.
Purpose: Carbonic anhydrase 1 (CA1)catalyze the reversible
hydration of carbon dioxide and participate in a variety of biological
processes.The aim of this study was to determine the level of CA1
in pterygial tissue, using normal human conjunctival samples as a
control.
Methods: Water soluable proteins of pterygium and normal human
conjunctiva were extracted and underwent SDS-polyacrimide gel
electrophoresis (SDS-PAGE). Intensive bands of pterygial proteins
were identified by MALDI-TOF-MS/MS. CA1, one of identied
proteins, was examined by Westernblot and immunohistochemical
staining, using normal human conjunctival samples as the control.
Results: On SDS-PAGE gel, several bands of pterygial extracts
were more intensive than that of normal conjunctiva. One band
was identified as CA1 by MALDI-TOF-MS/MS. Western blot
analysis reveraled that CA1 level increased in pterygial tissue when
compared with normal conjuctiva. Immunohistochemical staining
revealed that CA1 was more intensively stained in pterygial tissue
than that observed in normal conjunctival tissue. In pterygial tissue
that was characterized with neovascularization and congestion,
intensive staining appeared in red blood cells, vascular wall cells
and fibroblast cells as well as in some epithelial cells. While none
neovascularization and epithelial staining was found in normal
conjunctiva.
Conclusions: A high level of CA1 existed in pterygial tissue, which
was mainly resulted from the neovascularization and congestion of
pterygium.
Commercial Relationships: Kaili Wu, None; Qian Yang, None;
Xiuping Liu, None; Liping Lin, None; Feng Zou, None
Support: NSFC grant No. 81170827
Program Number: 3036 Poster Board Number: D0232
Presentation Time: 8:30 AM–10:15 AM
Findings of several immunohistochemical stains and
transmission electron microscopy (TEM) according to severity of
conjunctivochalasis
Woo C. Park, Jang Won Seo, Jeong Bum Bae, Choi Woo seok.
ophthalmology, Dong-A Univ. Hospital, Busan, Korea (the Republic
of).
©2015, Copyright by the Association for Research in Vision and Ophthalmology, Inc., all rights reserved. Go to iovs.org to access the version of record. For permission
to reproduce any abstract, contact the ARVO Office at [email protected].
ARVO 2015 Annual Meeting Abstracts
Purpose: This study compared and analyzed the correlation between
the severity of conjunctivochalasis and the findings of several
immunohistochemical stains and TEM.
Methods: A prospective study was conducted on 24 eyes from 23
patients with conjunctivochalasis who had undergone excisional
biopsies between June 2013 and January 2014. The severity
of conjunctivochalasis was categorized as LIPCOF Grade.
H&E staining, Verhoeff-Van Gieson (VVG) staining, D2-40
immunohistochemistry, TUNEL assay, and transmission electron
microscopy (TEM) were used to examine excised conjunctival
specimens.
Results: Conjunctivochalas subjects were classified by severity
degrees of mild to severe, using LIPCOF grade. Increased severity of
conjunctivochalasis increased in the infiltration of inflammatory cells
and reduced collagen fiber density on H&E staining, and it increased
the degeneration of elastic fibers on VVG staining (p<0.05). Increases
in the number of lymphatics and dilated lymphatics were observed
on D2-40 immunohistochemistry (p<0.05). TUNEL assay revealed
an increase in cell apoptosis with increasing conjunctivochalasis
severity (p<0.05). TEM revealed apoptosis of conjunctival epithelial
cells, destruction of nuclei, a decrease in intercellular cohesion
and destruction of inter-digitation, and marked accumulation of
degenerated elastic fibers.
Conclusions: Severity-dependent associations were confirmed
using H&E staining, VVG staining, D2-40 immunohistochemistry,
TUNEL assay, and TEM in conjunctivochalasis. The results of our
immunohistochemical studies will be the foundation of further
studies in the pathogenesis of conjunctivochalasis.
Commercial Relationships: Woo C. Park, None; Jang Won Seo,
None; Jeong Bum Bae, None; Choi Woo seok, None
Program Number: 3037 Poster Board Number: D0233
Presentation Time: 8:30 AM–10:15 AM
Pterygium and evidence of UV exposure: conjunctival
autofluorescence and cytological changes
Radhika Tandon, Sangeetha Vij, Namrata Sharma, Seema Sen. Dr R
P Centre for Ophthal Sci, All India Inst of Med Sciences, New Delhi,
India.
Purpose: To study the presence and pattern of conjunctival
autofluorescence in patients with pterygium and the underlying
cytological changes by impression cytology .
Methods: 75 patients with pterygium and 75 age and sex matched
controls were recruited. History, routine clinical examination and
relevant investigations were done. Parameters noted were grading
of pterygium, presence and pattern of conjunctival autofluorescence.
Impression cytology of the nasal conjunctiva was done both in
cases and controls. The same process was repeated after 6 months.
Progression of the pterygium was assessed clinically and also from
the photographs with the help of Image J software.
Results: Conjunctival autofluorescence was present in 51 % of cases
with pterygium. The predominant pattern was autofluorescence at
the leading edge (65.9%). Autofluorescence was detected in 83.3%
cases of grade 3 pterygia, 64.7% cases of grade 2 pterygia and 28.9%
of grade 1 pterygia ( p value <0.0005). By impression cytology,
conjunctival epithelial cells were found in 60% of cases. Squamous
metaplasia was significantly more frequently found in grade 3
pterygia (45%) compared to grade 2 (11.8%) and absent in grade 1
pterygia (p value< 0.0005).
Conclusions: The presence and pattern of conjunctival
autofluorescence was related to both the grade and impression
cytology changes especially squamous metaplasia.
Commercial Relationships: Radhika Tandon, None; Sangeetha
Vij, None; Namrata Sharma, None; Seema Sen, None
Program Number: 3038 Poster Board Number: D0234
Presentation Time: 8:30 AM–10:15 AM
Aldehyde dehydrogenase (ALDH) inhibition is a novel potential
therapy for conjunctival fibrosis in ocular mucous membrane
pemphigoid (OcMMP)
John K. Dart1, 2, Sarah Dale2, Julie T. Daniels2, Valerie P. Saw1,
2
, David Abraham3. 1Corneal and External Disease, Moorfields
Eye Hospital, London, United Kingdom; 2Ocular Biology and
Therapeutics, UCL Institute of Ophthalmology, London, United
Kingdom; 3Research Department of Inflammation, Royal Free and
University College Medical School University College London Royal Free Campus, London, United Kingdom.
Purpose: We have previously shown that OcMMP human
conjunctival fibroblasts (HCF) maintain their phenotype in culture.
We hypothesize that in vitro models of OcMMP HCF activity may
predict conjunctival scarring mechanisms in-vivo and allow us to
identify profibrotic targets, and analyse their therapeutic potential. We
have used gene expression, proteomics and western blots of whole
OcMMP conjunctival tissue, and OcMMP HCF, to identify putative
conjunctival scarring inhibitors. We investigated the therapeutic
potential of these inhibitors in our in vitro HCF model.
Methods: Gene expression on whole OcMMP conjunctiva and on
cultured OcMMP HCF was carried out by microarray to identify
over expressing genes in OcMMP. Western blots were used to
confirm over expression of targets in OcMMP fibroblasts. Kinexus
whole tissue proteomics, enabled identification of downstream
signaling molecules associated with the target protein. A range of
in vitro HCF assays including collagen secretion and production,
matrix contraction, proliferation, chemotaxis, and myofibroblast
differentiation were used to analyse the role of target proteins.
Results: Gene expression studies identified 14 overexpressed genes
in OcMMP. ALDH1A3 was chosen for further study, as being
potentially relevant to fibrosis pathways. Protein phosphorylation
profiling, in whole OcMMP conjunctiva, identified NF-κBp50, p53
and Bcl-XL as potential ALDH1A3 signaling pathways. Western blot
of MMP fibroblasts showed increased ALDH1A3. OcMMP HCF
showed increased collagen production, decreased matrix contraction,
reduced rates of proliferation, and myofibroblast differentiation.
Addition of both DEAB (diethylaminobenzaldehyde) and disulfiram,
two selective ALDH inhibitors, restored the functionality of OcMMP
HCF to that of Control HCF. Conversely, adding retinoic acid
(a downstream product in the ALDH pathway) to Control HCF
promoted OcMMP like HCF dysfunction.
Conclusions: These findings taken together show that ALDH1A3 is
a novel target involved in the scarring process in OcMMP. In addition
they demonstrate that ALDH inhibition may provide a therapeutic
approach to fibrosis with application in OcMMP that should be
evaluated in other scarring disorders. Disulfiram is licenced for
human use, and can be repurposed for this indication.
Commercial Relationships: John K. Dart, UCL Business (P);
Sarah Dale, UCL Business (P); Julie T. Daniels, UCL Business (P);
Valerie P. Saw, None; David Abraham, UCL Business (P)
Support: Fight for Sight PhD studentship Ref: 1818; Anonymous
donors through Moorfields Eye Charity and Moorfields Eye Hospital
Special Trustees Grants ST 11 11D, ST11 11B, ST 13 04 E
©2015, Copyright by the Association for Research in Vision and Ophthalmology, Inc., all rights reserved. Go to iovs.org to access the version of record. For permission
to reproduce any abstract, contact the ARVO Office at [email protected].
ARVO 2015 Annual Meeting Abstracts
Program Number: 3039 Poster Board Number: D0235
Presentation Time: 8:30 AM–10:15 AM
The role of IL-13 in the pathogenesis of ocular mucous
membrane pemphigoid
Suryanarayana Rayapureddi1, 2, Jill Norman1, Valerie P. Saw3, 4,
David Abraham2, John K. Dart3, 4. 1UCL Centre for Nephrology,
University College London, London, United Kingdom; 2UCL Centre
for Rheumatology and Connective Tissue Diseases, University
College London, London, United Kingdom; 3Corneal and External
Disease, Moorfields Eye Hospital, London, United Kingdom; 4Ocular
Biology and Therapeutics, UCL Institute of Ophthalmology, London,
United Kingdom.
Purpose: Ocular mucous membrane pemphigoid (OcMMP),
a severe inflammatory autoimmune disease is characterised by
conjunctival fibrosis leading to blindness. Current therapies fail
to halt the progressive conjunctival scarring and there is a need
for novel anti-fibrotic therapies. We previously reported the high
expression of Interleukin-13 (IL-13) in OcMMP conjunctiva (Saw et
al., Am J Pathol. 2009). We hypothesize that IL-13 plays a role in the
pathogenesis of OcMMP, activating human conjunctival fibroblasts
(HCF) to promote scarring, and that it may be a new therapeutic
target.
Methods: HCFs were grown from conjunctival biopsies
obtained from OcMMP patients and age-matched controls (5
patients/group). Expression of IL-13 receptors on HCFs was
examined by flow cytometry. The effects of IL-13 on HCF
function were measured using established fibrogenic assays: cell
proliferation (BrdU incorporation), migration (scratch wound
assay), collagen gel contraction and myofibroblast differentiation
(expression of α-Smooth Muscle Actin by Western blotting
and immunofluorescence). Gene expression profiling (Illumina
Transcriptome Arrays) was done to identify IL-13 target genes.
Cytokine production in response to IL-13 was assessed by Multiplex
ELISA of HCF conditioned media.
Results: Both IL-13 receptors (IL-4α/IL-13Rα1; IL13Rα2) are
expressed on HCFs with high levels of IL-13Rα1 (both groups) and
variable IL13Rα2 expression (control-low; OcMMP-variable). IL-13
dose-dependently (5 – 50ng/ml) inhibits control HCF proliferation
(p<0.05 in 4/5) but had no significant effect on OcMMP cells. IL-13
stimulates collagen gel contraction by control HCFs (% change
after 7-days; p=0.025) but not by OcMMP cells (p=0.074). IL-13
had no effect on HCF migration or myofibroblast differentiation
in either group. Both pro-inflammatory (Interleukin-6, Monocyte
Chemoattractant Protein-1) and pro-fibrotic proteins (which regulate
collagen biosynthesis and extracellular matrix (ECM) stability) were
induced by IL-13 in both groups.
Conclusions: Our current data suggest that, although IL-13 has
no direct pro-fibrotic effects on HCFs, IL-13 produced by T-cells
may act to promote inflammation and sustain fibrosis through the
induction of inflammatory cytokines and proteins involved in ECM
synthesis and stability. Taken together, these data suggest that IL-13
inhibition may be an effective topical therapy to limit scarring in
OcMMP patients.
Commercial Relationships: Suryanarayana Rayapureddi, None;
Jill Norman, None; Valerie P. Saw, None; David Abraham, None;
John K. Dart, None
Support: UCL Grand Challenges Studentship, Anonymous donors
through Moorfields Eye Charity and Moorfields Eye Hospital Special
Trustees Grants ST 11 11D, ST11 11B, ST 13 04 E
Program Number: 3040 Poster Board Number: D0236
Presentation Time: 8:30 AM–10:15 AM
Conjunctival epithelial cells (CEC) morphology in hematological
patients before and after hematopoietic stem cell transplantation
(HSCT). A Cytological and Confocal Microscopy Study
Giuseppe Giannaccare1, Piera Versura1, Nicoletta Testoni2, Gaia
Ameli2, Michela Fresina1, Francesca Bonifazi2, Giuseppe Bandini2,
Emilio C. Campos1. 1DIMES, Ophthalmology Unit, Alma Mater
Studiorum University of Bologna, Bologna, Italy; 2Haematology
Institute “L.A. Seragnoli”, DIMES, Alma Mater Studiorum
University of Bologna, Haematology Institute “L.A. Seragnoli”,
Bologna, Italy.
Purpose: To analyze CEC morphology in hematological patients
before and after HSCT and to correlate possible modifications to
HSCT parameters and ocular surface impairment
Methods: Prospectively collected data from 33 patients (20 F; 13 M)
who had received HSCT were retrospectively analyzed upon local
ethical committee authorization. CEC collected before the pre-op
conditioning regimen (V0) and 1 month (V1), 3 months (V2), 6
months (V3) and 12 months (V4) post- op were analyzed by in vivo
confocal microscopy (IVCM), scraping and impression cytology,
immunocytochemistry (ICC) and interphase FISH analysis with
X and Y chromosome probes in donor/recipient sex-mismatched
cases. Data were related to recipient age, donor type and age, HSC
source, T-cell depletion intensity, sex mismatch, Dry Eye (DE) onset,
systemic and ocular graft versus-host disease (GVHD)
Results: No patient developed ocular GVHD. DE was diagnosed
in 13 patients already at V0 and in 7 patients after HSCT (medium
interval 3.8±3.1 months from HSCT to DE onset). Squamous
metaplasia with decrease/absence of goblet cells was found already at
V0. Giants polyploid (GP) cells were never observed at V0 but were
detected in 18 patients only at V1 with density 10-45% vs total CEC.
GP cells reached a density peak at V2 and V3 and were found up
to V4 in 7 of 26 surviving patients who were living without disease
relapse. GP cells were also detected by IVCM as randomly grouped
enlarged superficial cells. No polyploid cell was observed in bone
marrow aspirate at the same post-op intervals. FISH analysis did
not show any donor chromosome; ICC showed a CK19 positivity
suggesting a conjunctival recipient origin. GP cell density was found
to be related to squamous metaplasia degree (r=0.572,p<0.01) and
T-cell depletion intensity (r=0.43,p<0.05)
Conclusions: This is the first study diagnosing DE in hematological
patients already before HSCT and conditioning regimen. Further,
GP cells were detected after HSCT in conjunctiva for the fisrt time
by IVCM. These cells have been shown to increase in number as a
response to adverse circumstances as chemotherapy but their role
has been poorly investigated in human conjunctiva. In this study GP
cells appeared to be related to conditioning regimen and squamous
metaplasia representing a possible marker of post-HSCT ocular
surface dryness
Commercial Relationships: Giuseppe Giannaccare, None; Piera
Versura, None; Nicoletta Testoni, None; Gaia Ameli, None;
Michela Fresina, None; Francesca Bonifazi, None; Giuseppe
Bandini, None; Emilio C. Campos, None
Program Number: 3041 Poster Board Number: D0237
Presentation Time: 8:30 AM–10:15 AM
Performance of comparative studies between healthy
conjunctiva, SCC tissue and a conjunctival SCC cell line
Bettina Muller, Bernadette Brockmann-Ahmed, Henning Thomasen,
Marina Schock, Klaus-Peter Steuhl, Daniel Meller. Ophthamology,
University Duisburg Essen, Essen, Germany.
©2015, Copyright by the Association for Research in Vision and Ophthalmology, Inc., all rights reserved. Go to iovs.org to access the version of record. For permission
to reproduce any abstract, contact the ARVO Office at [email protected].
ARVO 2015 Annual Meeting Abstracts
Purpose: Our aim was to understand the squamous cell carcinoma
(SCC) from the human bulbar conjunctiva in more detail. Therefore,
we analyzed healthy conjunctival tissue in comparison to the SCC
tissue and the cell line PeCa-UKHb-01 derived out of SCC of the
bulbar conjunctiva. We set the focus to genes involved in tumor
development, stem cell characteristics and in tissue specific ones.
Methods: For the comparison of conjunctival tissue, tumor tissue and
PeCa-UkHb-01 a qRT-PCR based on the TaqMan® Assays as duplex
reactions was performed. Beforehand, the endogenous control genes
were tested and GAPDH as well as PPIA were suited the best ones.
Tissue specific genes of interest were EpCAM, K3, K15, K13, K19
and CD90. The cancer and stem cell genes of interest were ABCG2,
EGFR, KIT, Myc, Nestin, Notch1, Oct4, and SOX2, also NANOG a
gene known for stem cell characteristics. Furthermore, Pax6 a gene in
eye development was analyzed. Additional information was obtained
by immunofluorescent staining.
Results: The expression pattern of EGFR, KIT, Myc, Nestin,
NANOG and Oct4 did not show any conspicuousness. Notch1
expression was lower in PeCa-UkHb-01. Only the conjunctival
tissues had higher Pax6 expression. ABCG2 expression was higher
in the cell line. The expression pattern of Sox2 was different, in the
conjunctival tissue low, in the malignant tissue low and high and
PeCa-UkHb-01 in the middle. The expression of EpCAM was in the
cell line higher than in the tissue samples. K3 served as a control for
the purity of the samples. K13 and K15 expression was observed
in all samples. The conjunctival samples as well as PeCa-UkHb-01
showed expression of K19, none was seen in the tumor tissue
samples. The other way around was it with CD90.
Conclusions: Through this study we could show that there was
a variance between conjunctival tissue and tumor tissue in the
genes CD90 and K19. Sox2 needs further analyses to prove that
there could be a difference between distinguished tumor samples.
A variation between cell line and tissue samples was witnessed for
ABCG2, EpCAM and Notch1. Pax6 expression was equal for the
malignant tissue and the cell line. It is a gene for eye development
and therefore it was expected that both of these samples do not
show any expression. Further genes need to be analyzed to find one
which shows similarity between the cell line and tumor but not to the
healthy conjunctival tissue.
Commercial Relationships: Bettina Muller, None; Bernadette
Brockmann-Ahmed, None; Henning Thomasen, None; Marina
Schock, None; Klaus-Peter Steuhl, None; Daniel Meller, None
Program Number: 3042 Poster Board Number: D0238
Presentation Time: 8:30 AM–10:15 AM
Polymeric Shield to Reduce Conjunctival Scarring and Facilitate
Re-Access for Multistage Surgical Procedures
John J. Whalen1, 2, Yi Zhang1, 2, Paulo Falabella1, 2, Niki Bayat4, Mark
E. Thompson3, Mark S. Humayun1, 2. 1USC Eye Institute, University
Of Southern California, Los Angeles, CA; 2Institute for Biomedical
Therapeutics, University Of Southern California, Los Angeles,
CA; 3Chemical Engineering and Materials Science, University Of
Southern California, Los Angeles, CA; 4Chemistry, University Of
Southern California, Los Angeles, CA.
Purpose: Intervention of combat-related open globe injuries is
typically managed in two-stages: first the casualty is stabilized at
a critical care facility, and later, complete surgical intervention is
performed after transportation to a base hospital with full surgical
facilities. Casualties wait more than 24 hours before the second,
follow-up intervention. We evaluate, in vivo (rabbit), a nonresorbable polymer patch placed sub-conjunctival as a shield to
prevent fibrosis before full intervention.
Methods: Sub-conjunctival pockets(superior and inferior) were
created in one eye (OD) of New Zealand pigmented rabbits by
making a 5mm incision 3mm from and tangential to the limbus. A
sterile (ETO), flexible polymer substrates (Area: <100mm2) was
inserted into the superior pocket as a shield with the conjunctiva
drawn back over and held in place with a suture. Examinations for
infection or lacerations were conducted daily for 14 days, at which
time the implants were removed. Half of the animals were euthanized
and the treated eye was enucleated and fixed to compare fibrosis of
the superior (patch insert) vs. inferior (no insert) pockets. Remaining
rabbits were followed up for two additional weeks (Day-28) with the
superior pocket shields removed. At Day-28 the remaining rabbits
were euthanized and the treated eyes were enucleated and fixed for
histology to assess fibrosis 14-days post-shield removal. This study
was conducted in compliance with the ARVO Statement for the Use
of Animals in Ophthalmic and Visual Research.
Results: Shield placement caused no irritation or inflammation postplacement. Shield removal at Day-14 caused no additional trauma
and post-removal it enabled easy re-access to the scleral surface.
Fibrosis at Day-14 showed differences between superior (shielded)
and inferior (unshielded) pockets.
Conclusions: Temporary placement of polymeric shields between the
sclera and external tissues prevents fibrosis between apposed tissues,
allowing easier follow-up access as in the case of two-stage ocular
interventions for combat casualties. This research was conducted
with support from the Vision Research Program of the United States
Telemedicine and Advanced Technology Research Center (TATRC)
under contract # W81XWH-12-1-0314 and Research to Prevent
Blindness.
Commercial Relationships: John J. Whalen, None; Yi Zhang,
None; Paulo Falabella, None; Niki Bayat, None; Mark E.
Thompson, None; Mark S. Humayun, None
Support: TATRC Grant W81XWH-10-2-0076
Program Number: 3043 Poster Board Number: D0239
Presentation Time: 8:30 AM–10:15 AM
Immunohistochemical Analysis of the Conjunctiva in Peripheral
Ulcerative Keratitis
Douglas A. Lyall1, Deepa Anijeet1, Kaleena B. Michael1, Fiona
Roberts2, Kanna Ramaesh1. 1Ophthalmology, NHS Greater Glasgow
and Clyde, Glasgow, United Kingdom; 2Pathology, NHS Greater
Glasgow and Clyde, Glasgow, United Kingdom.
Purpose: Peripheral ulcerative keratitis is a sight-threatening
inflammatory disease of the cornea characterized by stromal thinning
and adjacent conjunctival inflammation that may be associated with
several systemic autoimmune conditions. We wished to identify types
of inflammatory cells infiltrating the conjunctiva and characterize the
cytokeratin (CK) profile of the conjunctiva in patients with peripheral
ulcerative keratitis by performing immunohistochemical analysis on
conjunctival biopsies taken during episodes of active ocular surface
inflammation.
Methods: Prospective, non-comparative, interventional case series.
Three patients (2 male, 1 female, mean age 50 years) with active
peripheral ulcerative keratitis (Figure 1) underwent conjunctival
recession as part of their management. Immunohistochemical staining
of the excised tissue was performed.
Results: In all cases it was shown that the lymphocytic population
was predominantly CD3 (Figure 2A) and CD4 (Figure 2B) positive
T cells. In one case there were more CD8 positive than CD4 positive
cells. In two cases CD8 positive cells (Figure 2C) were only present
in small amounts. In all cases there were only small numbers of
CD20 positive B cells (Figure 2D). A background population of
histiocytes and a small number of macrophages with CD68 were also
©2015, Copyright by the Association for Research in Vision and Ophthalmology, Inc., all rights reserved. Go to iovs.org to access the version of record. For permission
to reproduce any abstract, contact the ARVO Office at [email protected].
ARVO 2015 Annual Meeting Abstracts
identified. Cytokeratin profiling of the conjunctiva found that CK 19
was diffusely present throughout the epithelium and CK 3 and 12
were only present in the more superficial layers.
Conclusions: The predominant cells infiltrating the conjunctiva
during peripheral ulcerative keratitis are CD3 and CD4 positive T
cells. When considering systemic therapy to treat this condition,
clinicians should consider using treatments specifically targeting this
group of cells. Therapies targeting other cells, such as B cells, would
be less effective as such cells are only present in small numbers.
Peripheral ulcerative keratitis in a 25-year-old female
Immunohistochemical staining of conjunctival biopsy (A) CD3
positive T cells, (B) CD4 positive T cells, (C) CD8 positive T cells,
and (D) CD20 positive B cells
Commercial Relationships: Douglas A. Lyall, None; Deepa
Anijeet, None; Kaleena B. Michael, None; Fiona Roberts, None;
Kanna Ramaesh, None
Program Number: 3044 Poster Board Number: D0240
Presentation Time: 8:30 AM–10:15 AM
Effect of antioxidant on conjunctiva exposed to high glucose
Soon-Suk Kang, Jin Hyoung Park. Ophthalmic, Seoul Asan Medical
Center, Seoul, Korea (the Republic of).
Purpose: To evaluate the effects of N-acetylcystein (NAC), which is
known to inhibit ROS-dependent apoptosis, on high glucose-induced
ROS, apoptosis, inflammation, and delayed-wounding closure in
primary cultured human conjunctival epithelial cells (pHCECs) and
regulatory effects of cleaved caspase-3, Bax, NF-kB, and IL-6 and
TNF-α on this process.
Methods: High glucose-induced ROS generation was measured
using 2’,7’-dichlorofluorescein diacetate (DCFH-DA). Effects
of NAC for high glucose-induced apoptosis were investigated in
pHCECs using Annexin-V and PI staining and cleaved caspase-3
and Bax expression levels using immunoblotting. To evaluate
inflammatory response, IL-6 and TNF-α expression levels were
quantified by multiplex cytokine analysis and NF-kB activation and
IkB-α degradation were assessed by Western blot. The effects of
NAC on high glucose-delayed conjunctival epithelial wound healing
were assessed by Scratch-induced directional wounding assay.
Results: Compared with untreated control and normal glucose, 5
mM, high glucose at 25 mM stimulated ROS generation, apoptosis,
inflammatory cytokines, and delayed wound healing in pHCECs. The
addition of NAC markedly abolished the high glucose-induced ROS,
Annexin-PI-positive cells, levels of cleaved caspase-3 and Bax, and
IL-6 and TNF-α. Also, NAC prevented high glucose-delayed wound
healing.
Conclusions: High glucose promotes apoptosis by affecting
mitochondria-dependent caspase activity, through elevated ROS, a
process that can be reversed by the antioxidant NAC. These findings
demonstrated that NAC have a beneficial effect on conjunctival
epithelial cell wound healing, anti-apoptosis, and anti-inflammation.
Commercial Relationships: Soon-Suk Kang, None; Jin Hyoung
Park, None
Support: NRF-2012R1A1A2003278
Program Number: 3045 Poster Board Number: D0241
Presentation Time: 8:30 AM–10:15 AM
Evaluation of a Smart Phone-Based Ocular Imaging System for
the Assessment of Conjunctival Hyperemia
Peter Corcoran1, Richard Abelson2, Keith J. Lane3, Endri Angjeli3,
George W. Ousler1, Paul J. Gomes4. 1Dry Eye, R & D, Ora, Inc.,
Andover, MA; 2Statistics and Data Corporation, Tempe, AZ; 3R & D,
Ora, Inc., Andover, AZ; 4Allergy, Ora, Inc., Andover, MA.
Purpose: Conjunctival hyperemia can vary greatly based on the
external environment, time of day, and intermittent nature of ocular
surface diseases. In-office slit lamp examinations are the gold
standard for evaluation of conjunctival hyperemia, but are infrequent
snap shots that do not provide a global assessment of redness severity
and duration, especially in the context of evaluating drug efficacy
and duration of effect. We developed and tested a smart-phone based
image capture system that the subject can carry out independently to
capture conjunctival redness away from the clinician’s office.
Methods: An ocular imaging device and instructional software
application were developed for a Samsung Galaxy S5 smart phone.
Twenty images from 10 eyes were evaluated in this proof-of-concept
study. Healthy volunteers self-captured images of both eyes using
the smart phone system. A trained clinician then graded the severity
of ocular hyperemia (0-4 scale) in these subjects at the slit lamp.
All images were graded manually by trained investigators, and
automatically, using a validated automated computer program1.
Pearson’s Correlations were calculated between live grading and
images graded by the clinician, and between live grading and images
graded automatically.
Results: Previous results1 demonstrated that inter-investigator
correlations of manual image grading varied between 0.68<r<0.83
(mean r=0.77; N=99 images). In this study, correlation between
live grading at the slit lamp and manual image grading was r=0.76.
Correlation between live grading and automatic grading of images
was r=0.65.
©2015, Copyright by the Association for Research in Vision and Ophthalmology, Inc., all rights reserved. Go to iovs.org to access the version of record. For permission
to reproduce any abstract, contact the ARVO Office at [email protected].
ARVO 2015 Annual Meeting Abstracts
1 Rodriguez JD, Johnston PR, Ousler GW, Smith LM, Abelson
MB. Automated grading system for evaluation of ocular redness
associated with dry eye Clinical Ophthalmology 2013:7 1–8.
Conclusions: These results indicate that a smart phone-based
imaging system, with subsequent automated or clinical grading of
redness, correlates well to live-doctor assessments. This novel device
provides subject-captured data as yet never collected in real-time
environmental assessments of conjunctival hyperemia, and such a
tool might prove valuable in the evaluation of drugs that modify
redness, or as an adjunct to cell-phone based clinical trial diary
platforms.
Commercial Relationships: Peter Corcoran, Ora, Inc. (E);
Richard Abelson, SDC, Inc. (E); Keith J. Lane, Ora, Inc. (E);
Endri Angjeli, Ora, Inc. (E); George W. Ousler, Ora, Inc. (E); Paul
J. Gomes, Ora, Inc. (E)
Program Number: 3046 Poster Board Number: D0242
Presentation Time: 8:30 AM–10:15 AM
Acrolein exposure severely damages cornea, eyelids, conjunctiva
and causes vision loss in rabbits in vivo
SUNEEL Gupta1, 3, Ajay Sharma1, 3, Charles R. Brown1, 3, Elizabeth A.
Giuliano1, 3, Prashant Sinha1, 3, Rajiv R. Mohan1, 2. 1Harry S Truman
Memorial Veterans’ Hospital, Columbia, Missouri, Columbia, MO;
2
Mason Eye Institute, School of Medicine, University of Missouri,
Columbia, MO; 3Veterinary Medicine and Surgery, College of
Veterinary Medicine, University of Missouri, Columbia, MO.
Purpose: Acrolein was used recently in Syria and during World War
I as a chemical weapon. It caused severe eye, skin and lung problems
besides many casualties. At present, no animal model exists, to study
acrolein’s toxicity to eye. We sought to develop an in-vivo rabbit
model to understand mechanisms causing blindness by assessing
damages to the cornea, conjunctiva, eyelids and sclera, and identify
potential targets for developing antidotes for acrolein’s toxicity.
Methods: New Zealand White rabbits were used. Eyes were exposed
to varied acrolein conc (0.5M, 2M, 6M or 10M) for 1 min. Slit-lamp
biomicroscopy and eye examination were performed before and after
acrolein exposure at 0h, 4h, 24h, 3day, 7day, 10day and 14day and
record to analyze ocular inflammation and corneal pathology. Cornea,
eyelid and sclera tissues were harvested and cryo-preserved for
histology and various assays. H&E and immunofluorescence study
investigated the role of macrophages, neutrophils and lymphocytes,
keratocyte density, in damage cornea and other ocular tissues. Tissue
lysates were used to quantify oxidative stress, glutathione peroxidase,
superoxide dismutase, total lipid peroxidation and eicosanoids.
Results: Acrolein caused severe inflammation, irritation, and
damage to cornea, eyelids and sclera within 15 min of exposure.
This pathology and vision loss enhanced over time. The damages to
cornea, eyelid and conjunctiva were dose dependent. High acrolein
concentrations (6M and 10M) caused severe morbidity, thus animals
were humanely euthanized after 4h or 24h. Dramatically increased
leukocyte invasion, leukotriene LTB4 (2.5 fold) and reactive oxygen
species levels (5 fold) and remarkably decreased glutathione levels
(40-60%; p <0.01) were detected in eyelid and corneal tissues.
All acrolein-exposed eyes developed eyelid and meibomian gland
dysfunction, corneal fibrosis and neovascularization by day5;
pathology deteriorated by day14. H&E and immunofluorescence
detected exceptionally increased levels of TGFβ, VEGF, keratocyte
death, invading-vessels, and neutrophils in corneal sections.
Conclusions: Acrolein causes severe acute ocular inflammation,
corneal damage and vision loss. TGFβ, VEGF, leukotrienes and
glutathione are attractive targets to counter acrolein’s toxicity. More
in-vivo studies are warranted.
Commercial Relationships: SUNEEL Gupta, None; Ajay Sharma,
None; Charles R. Brown, None; Elizabeth A. Giuliano, None;
Prashant Sinha, None; Rajiv R. Mohan, None
Support: The Ruth M. Kraeuchi Missouri Endowment Chair
Ophthalmology Fund. Partially from the RO1EY17294 National
Eye Institute, NIH, Bethesda, Maryland, USA and 1I01BX00035701
Veteran Health Affairs, Washington DC USA grants.
Program Number: 3047 Poster Board Number: D0243
Presentation Time: 8:30 AM–10:15 AM
Angiographic Characteristics of the Pterygia and Restoration
of the Structure After Limbal Conjunctival Autograft
Transplantations
Seung Hoon Yoo, Jin Kwon Chung. Ophthalmology, Soonchunhyang
University Hospital, Seoul, Korea (the Republic of).
Purpose: Pterygia recently have been thought to originate
from limbal stem cells. In past, few studies have evaluated
the angiographic features of the affected limbus. We studied
prospectively to evaluate the perioperative angiographic
features of the limbus in patients with pterygia and assessed
limbal reconstruction after limbal-conjunctival autograft (LCA)
transplantation in terms of vascular remodeling.
Methods: Thirty one eyes with pterygium of 31 patients and
sound eyes of 28 normal participants were enrolled in this study.
To investigate the angiographic characteristics of the pterygium,
indocyanine green angiography (ICGA) was performed for each
participant preoperatively and at 1 week, and 1 and 3 months
postoperatively. The perioperative angiographic features of the
pterygium were compared with control group. The structural
changes of the Marginal corneal vascular arcades (MCAs) and LCA
were quantitatively assessed postoperatively in terms of vascular
density and lacunarity using a semiautomated computational
tool(AngioTool).
Results: Deteriorated MCAs that extended beyond the pterygium
head were observed in the pterygium group. The pterygium had a
dual blood supply from the conjunctival and episcleral circulations.
In terms of limbal reconstruction, the engorged reperfusion vessels
arose from the adjacent episcleral vessels along the limbus at 1 week
postoperatively. The reconstructed MCAs had begun to appear at 1
month postoperatively and became apparent 3 months postoperatively
in 26 (83.9%) of 31 eyes of the pterygium group, resulting in a
successful clinical outcome. Higher vascular density and lower
lacunarity were measured in the limbus and the graft at 3 months than
at 1 month (P < 0.001 for all), which indicated fine reorganization of
the reconstructed vessels.
Conclusions: The MCAs were deteriorated in the affected
limbus, and the pterygium head had a dual blood supply from the
conjunctival and episcleral circulations. The remodeling of the LCA
and affected limbus continued up to 3 months postoperatively.
Commercial Relationships: Seung Hoon Yoo, None; Jin Kwon
Chung, None
Program Number: 3048 Poster Board Number: D0244
Presentation Time: 8:30 AM–10:15 AM
Minor Ipsilateral Simple Limbal Epithelial Transplantation
(Mini-SLET) for Pterygium Treatment
Erick Hernandez-Bogantes1, Arturo J. Ramirez-Miranda1, Guillermo
Amescua2, Alejandro Navas1, Alexandra Abdala-Figuerola1, Andrew
G. Olivo-Payne1, Enrique O. Graue1. 1Cornea and Refractive Surgery,
Conde de Valenciana, Mexico City, Mexico; 2Cornea and Refractive
Surgery, Bascom Palmer Eye Institute, Miami, FL.
©2015, Copyright by the Association for Research in Vision and Ophthalmology, Inc., all rights reserved. Go to iovs.org to access the version of record. For permission
to reproduce any abstract, contact the ARVO Office at [email protected].
ARVO 2015 Annual Meeting Abstracts
Purpose: To describe a novel surgical technique for pterygium
removal taking advantage of the properties of the amniotic membrane
and limbal epithelial stem cells.
Methods: A total of 13 patients with primary, recurrent or mixture
pterygium, with nasal or temporal location underwent a resection of
pterygium with conventional technique. The bare sclera was covered
with an amniotic membrane with fibrin sealant. Resection of 2x2 mm
of limbal epithelial stem cells at the superior or inferior palisades of
Vogt was performed. The epithelial limbal strip was cut into 8-10
pieces and placed in a linear fashion in the affected limbal area being
covered with a second amniotic membrane in a sandwich technique
with the use of fibrin sealant. Finally a soft contact lens was left in
place.
Results: After 4 months of follow-up, all patients were fully
epithelialised with a complete adaptation of the amniotic membrane.
The epithelial limbal stem cells remained in place and became
transparent. There were neither sight threatening complications nor
early signs of recurrence.
Conclusions: This technique requires less tissue than the
conventional conjunctival autograft and offers the advantages of
epithelial stem cells, which in a long-term may reduce the rate of
recurrence significantly.
in a controlled environment under standard housing conditions (5060% relative humidity). After five days of treatment, tear production
and ocular surface damage were accessed using phenol red thread
test (PRT) and corneal fluorescein staining respectively. Following
this, the eyes and lids were removed and processed for frozen tissue
sectioning. Goblet cell counts were obtained after periodic acid schiff
staining and the level of MMP9 on the ocular surface was determined
by immunohistochemistry.
Results: Both wild-type and MyD88 KO animals who underwent
experimental dry eye showed significant increases in ocular surface
staining (p<0.02), and a significant reduction in tear production via
PRT test when compared to untreated wild-type controls (p<0.01).
In addition, experimental dry eye MyD88 KO mice experienced
a significant decrease in ocular surface staining when compared
to wild-type mice under experimental dry eye (p<0.03). Goblet
cell counts were significantly increased in MyD88 animals who
underwent experimental dry eye when compared to their wild-type
counterparts (p<0.01).
Conclusions: This preliminary data suggests that TLRs play a role
in modulating ocular surface damage resulting from dry eye. It is
also known that TLRs play a protective role in pathogen recognition
enabling an innate immune response to microbes. When attempting
to alleviate dry eye symptoms it is important to maintain an ability
to fight pathogens, as such determining the exact role of TLRs in dry
eye inflammation is imperative for developing novel treatments that
modulate inflammation without increasing the risk for infection.
Commercial Relationships: Justin Courson, None; Carolina
Lema, None; Rachel L. Redfern, None
Support: EY023638
Preoperative and postoperative clinical photographs of 2 eyes that
underwent mini-SLET. A, preoperative photograph of a 47-year-old
male carpenter (case 9) with a history of bilateral pterygium. B, 1 day
alter the surgery. C, 1 week. D, 1 month. E, 3 months. F, Preoperative
photograph of a 26-year-old student (case 6) with bilateral pterygium,
the mini-SLET was performed in the temporal lesion. G, 1 day after
the surgery. H, 1 week. I, 1 month. J, 3 months.
Commercial Relationships: Erick Hernandez-Bogantes, None;
Arturo J. Ramirez-Miranda, Carl Zeiss Meditec (C); Guillermo
Amescua, None; Alejandro Navas, None; Alexandra AbdalaFiguerola, None; Andrew G. Olivo-Payne, None; Enrique O.
Graue, None
Program Number: 3050 Poster Board Number: D0246
Presentation Time: 8:30 AM–10:15 AM
Evaluation of a Single Thermal Pulsation Treatment for Dry Eye
and Meibomian Gland Dysfunction and Likelihood of Positive
SJO Test
Krysta Goslin1, Craig Czyz1, 2, Alice Epitropoulos2, 1. 1Ophthalmology,
Ohio State University, Grand Rapids, MI; 2Ophthalmology, The Eye
Center, Columbus, OH.
Purpose: Evaporative dry eye from meibomian gland dysfunction
(MGD) is the most common form of dry eye. Thermal pulsations
(LipiFlow®, Tearscience, Morrisville, NC) is the only FDA approved
treatment for Evaporative Dry Eye. The device uses a single
12-minute thermal pulsation to treat MGD. This study assesses if
LipiFlow® improves subjective and objective variables relating to
dry eye and MGD and if there is a correlation to Sjögren’s Syndrome.
Methods: Retrospective study of patients who received a single
LipiFlow treatment between October 2013 and October 2014.
Preoperative lipid layer thickness, degree of meibomian gland
dropout, symptom SPEED scores (Standard patient of eye dryness
[SPEED]), subjective comments, tear breakup time (TBUT), and
Meibomian Gland Evaluation Score (MGES) were recorded. Thirty
patients were also tested for Sjogren’s syndrome. SPEED Score,
TBUT, MGES and subjective comments were also obtained between
postoperative weeks 6-8. The Wilcoxon matched-pairs signed rank
test was used to compare preoperative and postoperative SPEED
scores and MGES and the paired t-test was used for preoperative and
postoperative TBUT for patients who underwent Lipiflow treatment.
Male and female outcomes were compared with the Mann Whitney
test.
Results: One hundred forty-six eyes, of 20 males and 66 females
(86 patients) were studied, with an average age of 63.5 years (SD
12.8). There was a statistically significant difference (p<0.05)
between preoperative and postoperative SPEED scores, TBUT, and
Program Number: 3049 Poster Board Number: D0245
Presentation Time: 8:30 AM–10:15 AM
The Role of Toll-like Receptors in Experimental Dry Eye
Inflammation
Justin Courson, Carolina Lema, Rachel L. Redfern. College of
Optometry, University of Houston, Houston, TX.
Purpose: We hypothesize that toll-like receptors (TLRs) play
a role in dry eye inflammation. We have previously shown that
hyperosmolar stress modulates TLR expression and the release
of endogenous damage associated molecular patterns (DAMPs)
which may activate TLRs to produce cytokines and matrix
metalloproteinases (MMPs). In support of this, we have shown that
topical TLR agonists induce corneal ulceration in mice undergoing
experimental dry eye. In this current study, we use MyD88 knock out
(loss of functioning TLRs) mice to investigate the role of TLRs in dry
eye pathology.
Methods: Experimental dry eye (EDE) was induced in six to eight
week old wild-type and MyD88 knock out (KO) C57BL/6 mice
by scopolamine hydrobromide injections and environmental stress
(10-20% humidity and constant air draft). Untreated mice were kept
©2015, Copyright by the Association for Research in Vision and Ophthalmology, Inc., all rights reserved. Go to iovs.org to access the version of record. For permission
to reproduce any abstract, contact the ARVO Office at [email protected].
ARVO 2015 Annual Meeting Abstracts
MGES. There was also a difference in postoperative SPEED scores
(p=0.0425) when comparing males and females but no preoperative
(p=0.123) difference. Of the 30 patients tested for Sjögren’s (Sjö™,
NICOX, Inc, Fort Worth, TX), 43% were positive (compared with
26% of the general dry eye population testing positive for Sjögren’s
syndrome biomarkers). Eighty-six percent of patients experienced a
subjective improvement in symptoms post treatment.
Conclusions: This study demonstrates the effectiveness of a single
12-minute thermal pulsation treatment in patients with dry eye from
MGD. The efficacy is evidenced by the improvement in postoperative
SPEED scores, TBUT, and MGES. There was a strong correlation
between positive Sjö™ tests and MGD, indicating an autoimmune
component contributing to symptoms experienced.
Commercial Relationships: Krysta Goslin, None; Craig Czyz,
None; Alice Epitropoulos, Allergan (C), Bausch and Lomb (C),
TearScience (C)
Program Number: 3051 Poster Board Number: D0247
Presentation Time: 8:30 AM–10:15 AM
Analysis of Meibum Before and Following Intraductal
Meibomian Gland Probing for Eyes with Obstructive Meibomian
Gland Dysfunction
Naohiko Nakayama, Motoko Kawashima, Minako Kaido, Reiko
Ishida, Reiko Arita, Sachiko Inoue, Kazuo Tsubota. Department of
Ophthalmology, Keio University School of Medicine, Shinjuku,
Japan.
Purpose: Obstructive meibomian gland dysfunction (o-MGD) is
a common lid margin disorder which causes a sense of stubborn
eye discomfort, yet our current symptomatic treatment is often
ineffective. Intraductal meibomian gland probing, an invasive orifice
penetration and intraductal probing was previously reported to
alleviate symptoms of o-MGD, although its effect on the meibum has
not been described. We hypothesize that this intervention increases
the amount of lipid flow and thus yields improvement in the ocular
symptoms.
Methods: Patients with o-MGD, which was refractory to
conventional management underwent intraductal meibomian gland
probing. Meibum and clinical outcome were evaluated before the
procedure and at the 1-month postoperative visits. Meibum analysis
was performed by the Meibometer®, which can quantify lipid
volume objectively. The condition of tear film was evaluated by
DR1 and meibomian gland loss was analyzed by noncontact infrared
meibography. Lid margin findings (tarsal hyperemia, telangiectasia,
mucocutaneous junction, plugging, thickening or irregularity),
tear break-up time (BUT), fluorescein score, meibum viscosity
(Shimazaki grade) and ocular symptoms were also assessed.
Results: Six lid margins of three refractive o-MGD eyes underwent
intraductal meibomian gland probing. There were no complications.
At the postoperative visits, all cases showed improvement of meibum
lipid levels (446 to 1376, 757 to 802, 396 to 571 meibometer units,
respectively) and meibum viscosity (grade 3 to 0, 3 to 1, 3 to 2,
respectively). Two cases showed an improvement of tear BUT (2 to
5, 0 to 6 sec, respectively). Morphological change of the meibomian
gland was not observed in any cases.
Conclusions: Intraductal meibomian gland probing seems to
provide improvement of lipid levels of the meibomian glands, and
it is an effective and promising treatment for o-MGD resistant to
conventional treatment.
Commercial Relationships: Naohiko Nakayama, None; Motoko
Kawashima, None; Minako Kaido, None; Reiko Ishida, None;
Reiko Arita, None; Sachiko Inoue, None; Kazuo Tsubota, None
Clinical Trial: 000014463
Program Number: 3052 Poster Board Number: D0248
Presentation Time: 8:30 AM–10:15 AM
Effect of Eye Shampoo for Obstructive Meibomian Gland Disease
Hirotaka Tanabe, Minako Kaido, Motoko Kawashima, Reiko Ishida,
Kazuo Tsubota. Ophthalmology, Keio University School of Medicine,
Tokyo, Japan.
Purpose: To investigate the efficacy of eyelid margin cleansing
with Eye Shampoo (MediProduct©, Japan), a lid hygiene product,
for obstructive obstructive meibomian gland dysfunction (MGD)
patients.
Methods: Thirty-seven eyes of 37 patients (10 males, 27 females,
aged 28 to 86 years [67.4±13.6 years]) diagnosed with obstructive
MGD were enrolled throughout this prospective clinical study.
All subjects were instructed to cleanse the eyelid margin with Eye
Shampoo (MediProduct©, Japan), the lid hygiene product, at least
once daily. Lid margin abnormalities, meibum properties, and tear
stability were observed before and 1 month after the initiation of
this study. Meibography was performed on all subjects to observe
morphological characteristics of the meibomian gland before the
treatment. A questionnaire relating to the severity of MGD symptoms
was administered after the treatment. Scores relating to the ocular
conditions, i.e., subjective symptoms improvement, corneal and
conjunctival fluorescein staining scores, tear break up time (TBUT),
lid margin lissamine green staining scores, mucocutaneous junction
(MCJ) by fluorescein, vascular dilatation, lid margin deformation,
plugging, meibum status, and meibomian gland scores by
meibography were evaluated. We adhered to the tenets of Helsinki,
and the ethics committee in Keio University School of Medicine
approved the protocol.
Results: There were significant improvements in TBUT, lid margin
lissamine green staining scores, and meibum status (P<0.05,
respectively). There were no significant improvements in corneal
and conjunctival fluorescein staining scores, MCJ by fluorescein,
vascular dilatation, lid margin deformation, and plugging (P^0.05,
respectively). As to subjective symptoms, there were improvements
in 29 subjects, and no improvements in 8 subjects. Meibography
revealed 31 subjects had normal meibomian glands (based on the
aggregate scores of both upper and lower lids), and 6 subjects had
meibomian gland atrophy and there were significant improvements
in the subjective symptoms, TBUT, and lid margin lissamine green
staining scores (P<0.05, respectively) in both groups, but meibom
status (P<0.05) only in the group with normal meibomian glands.
There were no obvious problems in all subjects throughout the study.
Conclusions: Lid margin cleansing with Eye Shampoo could be one
of the effective lid hygiene options.
Commercial Relationships: Hirotaka Tanabe, None; Minako
Kaido, None; Motoko Kawashima, None; Reiko Ishida, None;
Kazuo Tsubota, MediProduct (F)
Clinical Trial: UMIN000014459
Program Number: 3053 Poster Board Number: D0249
Presentation Time: 8:30 AM–10:15 AM
Quantitative measurement of conjunctival hyperemia and tear
film properties in glaucoma patients using the Keratograph 5M
Alfonso L. Sabater1, Anne-Marie Bleau1, Adriano Guarnieri1,
Vanesa Antón1, Gema Rebolleda2, Javier Moreno-Montanes1.
1
Ophthalmology, Clinica Universidad de Navarra, Pamplona-Iruna,
Spain; 2Hospital Ramón y Cajal, Madrid, Spain.
Purpose: To determine the effect of ocular hypotensive drops on
conjunctival hyperemia and tear film properties in glaucoma patients
using the Keratograph 5M (Oculus).
Methods: The conjunctival hyperemia (CH) and non-invasive
break-up time (NIBUT) in 200 glaucoma patients and 60 patients
©2015, Copyright by the Association for Research in Vision and Ophthalmology, Inc., all rights reserved. Go to iovs.org to access the version of record. For permission
to reproduce any abstract, contact the ARVO Office at [email protected].
ARVO 2015 Annual Meeting Abstracts
after glaucomatous filtration surgery (without other topical eyedrops) were quantitatively measured and compared to 50 control
patients. Only one eye of each patient was included. In addition, we
determined the corneal localization of tear film rupture. We analyzed
other parameters: the OSDI score, the number of antiglaucomatous
drugs, the number of eye drops administrated per day and the total
time of glaucomatous treatment. These parameters were compared
with CH and NIBUT using non-parametric statistical test.
Results: The mean (SD) CH was 1.69 (0.59) in glaucomatous
eyes, 1.49 (0.48) in operated eyes and 1.39 (0.38) in normal eyes
(p<0.05). In glaucomatous eyes, the CH was related to the number
of medication administrated (p=0.031). CH returned to normal
values after glaucoma surgery (p<0.05). OSDI scores were higher in
glaucomatous patients as compared to control patients (p<0.05), and
remained elevated in the operated group. In all patients, OSDI score
correlated with CH levels (p<0.05).
The mean (SD) NIBUT was 7.30 (5.45) in glaucomatous group, 8.38
(6.68) in operated eyes and 11.03 (6.99) in control group (p<0.05).
NIBUT values were significantly lower in the glaucoma surgery
group compared to control group (p=0.041). The tear breakup
location was in the inferior and nasal corneal area with a mean value
of 8.31 (1.11) sec in the treated group, and was in the temporal area
with 13.72 (1.54) sec in the control group (p<0.01).
Conclusions: The HC was higher in glaucomatous eyes than in
normal and operated group, indicating the irritating effect of the
medication on the ocular surface. The correlation between CH and
OSDI score confirms a greater discomfort associated with hyperemia.
The NIBUT was affected in glaucoma patients, especially in the
treated group. The localization of the ocular tear-film rupture was in
the inferior area, similar to that found in the dry-eye disease.
Commercial Relationships: Alfonso L. Sabater, None; AnneMarie Bleau, None; Adriano Guarnieri, None; Vanesa Antón,
None; Gema Rebolleda, None; Javier Moreno-Montanes, Sylentis
(F)
Support: RETICS-OFTARED, Instituto de Salud Carlos III,
Proyecto RD07/0062.
Program Number: 3054 Poster Board Number: D0250
Presentation Time: 8:30 AM–10:15 AM
Ocular Surface Cooling as a Potential Stimulus for Blinking
Wing Li1, 2, Andrew D. Graham2, Meng C. Lin1, 2. 1Vision Science
Graduate Group, University of California Berkeley, Berkeley, CA;
2
Clinical Research Center, University of California Berkeley School
of Optometry, Berkeley, CA.
Purpose: Animal models have identified the activation of corneal
cold receptors as the noxious stimulus primarily responsible for
tearing and could also be the potential stimulus for blinking. Previous
studies have found that a faster tear breakup time is associated with
an increased blink rate but there has been a lack of direct examination
on how cooling of the ocular surface is linked to the blink process. In
this study, we aim to determine if the ocular surface cooling (OSC)
rate is associated with how long an individual can refrain from
blinking.
Methods: Subjects were acclimated to the exam room for 10 minutes
before measurements. They were asked to open their eyes and refrain
from blinking for as long as they could, even if they experienced
discomfort, while the ocular surface temperature (OST) was
measured. Each measurement period was termed the maximum interblink period (MIBP). A total of ten measurements were taken with
four successive blinks separating each period. OST was measured
using an infrared thermographer (FLIR A655sc) and the OSC rate
was analyzed using commercially available software.
Results: 17 subjects (11 females, 6 males) completed the study. The
mean OSC rate in the study was .0470C/seconds (range: 0.00020.34770C/seconds) with a mean MIBP of 17.2 seconds (range:
1.1-90.5 seconds). Using a linear mixed effects model with a natural
log-transformed MIBP, increased OSC rate was significantly related
to a shorter MIBP (p<0.001). Based on the statistical model, it is
estimated that an individual would be able to refrain from blinking
for an additional 16.0 seconds if they had the minimum OSC rate
found in the study (0.00020C/seconds) when compared to someone
with the maximum OSC rate (0.34770C/seconds).
Conclusions: Our results show that the OSC rate was related to how
long an individual could refrain from blinking, suggesting that OSC
may be a potential stimulus for a blink.
Commercial Relationships: Wing Li, None; Andrew D. Graham,
None; Meng C. Lin, None
Program Number: 3055 Poster Board Number: D0251
Presentation Time: 8:30 AM–10:15 AM
Inflammatory manifestations of Ocular Graft versus Host Disease
in the Cornea, Eyelid and Conjunctiva
Stephanie Duffort1, Henry Barreras2, Robert Levy2, 1, Victor L.
Perez1, 2. 1Ophthalmology, Bascom Palmer Eye Institut, Miami, FL;
2
Immunology and Microbiology, University of Miami Miller School
of Medicine, Miami, FL.
Purpose: Graft-versus host disease (GVHD) remains a major
complication of allogeneic hematopoietic stem cell transplantation
(HSCT) which is frequently accompanied by ocular involvement.
We previously reported that in recipient mice following experimental
MHC-matched minor histocompatibility-mismatched HSCT, ocular
GVHD involves the presence of donor T cells in the cornea and
pathologic changes in this compartment. The present study was
directed to determine the kinetics of donor allo-reactive T cell
presence in the cornea and investigate the presence of macrophage
infiltration into the ocular adnexal tissue in recipient mice undergoing
experimental allogeneic HSCT.
Methods: Experimental HSCT Model: After high-dose TBI, C3H.
SW (H2b) mice were transplanted with T cell depleted bone marrow
(TCD-BM) alone from B6 (H2b) EGFP transgenic mice or alloreactive T cells from B6 (H2b) CD45.1 transgenic mice. Recipients
were monitored weekly for signs of systemic and oGVHD. In vivo
fluorescent intravital microscopy was performed to track EGFPlabeled TCD-BM cells. Cornea, lids and conjunctiva were harvested
weekly beginning 2 weeks post-HSCT for evaluation by histology,
immunohistochemistry (IHC) and flow-cytometry.
Results: Mice transplanted with donor TCD-BM+T cells underwent
weight loss and began exhibiting clinical signs of systemic GVHD.
EGFP-labeled TCD-BM cells were detectable in the lid margin,
conjunctiva and cornea of animals undergoing systemic GVHD
by week two after transplantation and showed massive increased
fluorescence by week six. Allo-reactive T cells were also present at
week two which peaked at week four. IHC confirmed the presence of
T cells and macrophage infiltration in the lid margin and conjunctiva.
By week six, lid pathology showed significantly elevated donor
inflammatory cell infiltrate consisting primarily of macrophages and
T cells compared to control (TCD-BM only) recipients.
Conclusions: GVHD is a complex systemic disorder involving
multiple tissues including the eye. The present study demonstrated
that following HSCT, allo-reactive T cells appear first in the lid
margin, conjunctiva and cornea and subsequently the recruitment
of macrophages in the ocular adnexa can be identified followed by
tissue damage.
Commercial Relationships: Stephanie Duffort, None; Henry
Barreras, None; Robert Levy, None; Victor L. Perez, None
©2015, Copyright by the Association for Research in Vision and Ophthalmology, Inc., all rights reserved. Go to iovs.org to access the version of record. For permission
to reproduce any abstract, contact the ARVO Office at [email protected].
ARVO 2015 Annual Meeting Abstracts
Support: Physician Scientist RPB Grant, R01 EY018624, P30
EY014801 from National Institute of Health, National Eye Institute,
Bethesda, MD, USA and Research to Prevent Blindness
Program Number: 3056 Poster Board Number: D0252
Presentation Time: 8:30 AM–10:15 AM
Expression of WW domain-containing Oxidoreductase WOX1 in
Pterygium
Yi-Hsun Huang1, 2, Sung-Huei Tseng1. 1Ophthalmology, National
Cheng Kung University Hospital, Tainan, Taiwan; 2Institute of
Clinical Medicine, National Cheng Kung University, Tainan, Taiwan.
Purpose: Pterygia was traditionally regarded as a degenerative
disease, but some tumor-like features, such as a propensity to
invade normal tissue, high recurrence rates following resection, and
the coexistence with secondary premalignant lesions, suggest that
pterygia might be a premalignant tissue.The WW domain containing
oxidoreductase (WWOX) gene was recently identified as a candidate
tumor suppressor gene, thus, in this study, we investigated WWOX
expressions in pterygium.
Methods: Pterygium tissues were obtained from patients (n=16,
primary=8, recurrent=8) who received surgical excisions. Each
tissue was further divided into head and body region. WWOX
expressions were examined by immunohistochemistry, western blot
and quantitative polymerase chain reaction. For comparison, normal
superior temporal bulbar conjunctivas were used as controls.
Results: Up-regulation of WWOXs and Tyr33 phosphorylation
WWOXs was observed in the head region of all pterygium
specimens. In the head and body of pterygium, WWOXs expressions
were significantly different from controls. Additionally, WWOX
expression is higher in the recurrent pterygia than in the primary
pterygia.
Conclusions: Increased WWOX expression, especially in the head
region, is an up-regulation in response to the invasiveness of the
pterygium. Our result indicated that WWOX might play a role in
pterygium progression and recurrence.
WWOX expressions in primary pterygium
WWOX expressions in recurrent pterygium
Commercial Relationships: Yi-Hsun Huang, None; Sung-Huei
Tseng, None
Program Number: 3057 Poster Board Number: D0253
Presentation Time: 8:30 AM–10:15 AM
Corneal abrasion following anesthesia for non-ocular surgical
procedures.
A case-control study
Anais Carniciu1, Melissa Fazzari2, Pauline Tabibian2, Priti Batta2, 3,
Ronald C. Gentile2, 3, James Grendell2, Collin Brathwaite2, Nazanin
Barzideh2, 3. 1Stony Brook University School of Medicine, Stony
Brook, NY; 2Winthrop-University Hospital, Mineola, NY; 3The New
York Eye and Ear Infirmary of Mount Sinai, New York, NY.
Purpose: To identify and characterize the risk factors associated
with corneal abrasions following anesthesia for non-ocular surgical
procedures at a single institution.
Methods: The medical, surgical, and anesthesia records of
consecutive patients with a corneal abrasion following anesthesia that
underwent ophthalmology consultation between 6/2011 and 11/2013
were reviewed. Data collected included demographics, medical
history, duration and type of surgery, and patient positioning during
surgery. Similar data was collected for a comparative group of agematched controls from the same time period. Analysis was performed
using an age-stratified logistic regression model evaluating the
association between corneal abrasion occurrence and gender, diabetes
status, duration of surgery, surgical position (supine vs. lateral vs.
prone), and type of procedure (head and neck vs. other procedures).
Odds ratios (ORs) and their 95% confidence intervals (CIs) were
calculated as a measure of the association between corneal abrasions
and the variables tested.
Results: Thirty-seven consecutive patients with post-operative
corneal abrasions were identified over the 2.5-year period and
compared to 101 age-matched controls (202 eyes). Corneal abrasions
were associated more often with procedures lasting 3 hours or more.
Forty-eight percent of the patients with a corneal abrasion had
surgeries lasting ≥ 3 hours compared to 18% of the patients in the
control group without a corneal abrasion. The adjusted odds of a
corneal abrasion is 5.3 times greater for patients having surgery for
≥ 3 hours (univariate OR=4.2, 95% CI:1.9-9.5, p<0.001). Gender,
diabetes status, surgical procedure or position were not found to be
associated with the occurrence of a corneal abrasion.
Conclusions: Longer duration of the surgical procedure is a risk
factor for a corneal abrasion. Surgeons and anesthesiologists should
be aware of this risk factor and take extra precautions to prevent this
complication, especially when surgery takes longer than three hours.
Commercial Relationships: Anais Carniciu, None; Melissa
Fazzari, None; Pauline Tabibian, None; Priti Batta, None; Ronald
C. Gentile, Alcon Laboratories, Inc. (C); James Grendell, None;
Collin Brathwaite, None; Nazanin Barzideh, None
Program Number: 3058 Poster Board Number: D0254
Presentation Time: 8:30 AM–10:15 AM
A Phase 3 Clinical Study to Evaluate the Efficacy and Safety
of Fixed Combination of Azithromycin and Dexamethasone in
DuraSite, and Dexamethasone alone in DuraSite in the Treatment
of Blepharitis
Kamran Hosseini1, 2, Richard Lindstrom3. 1Clinical, InSite Vision Inc,
Alameda, CA; 2KMG Pharma, Mountain View, CA; 3Minnesota Eye
Consultants, Bloomington, MN.
Purpose: To evaluate the efficacy and safety of the combination
and single agents, dexamethasone and azithromycin in DuraSite
vehicle (AzaSite). The study was also designed to investigate disease
response to therapy in combination with lid hygiene using lid scrubs.
Furthermore, the recurrence rate of the disease was to be investigated
during a 6 month follow up period.
©2015, Copyright by the Association for Research in Vision and Ophthalmology, Inc., all rights reserved. Go to iovs.org to access the version of record. For permission
to reproduce any abstract, contact the ARVO Office at [email protected].
ARVO 2015 Annual Meeting Abstracts
Methods: In this randomized, double masked, multicenter, 4-arm,
parallel-group, comparative study subjects received: dexamethasoneazithromycin combination (n=305), dexamethasone (n=298), AzaSite
(n=155), or DuraSite Vehicle (n=149). Subjects started the study with
a weeklong lid hygiene regimen followed by the application of 1
drop of study medication onto the eyelid(s) twice a day for 14 days.
Subjects were assessed on Day 4 during the 2-week dosing period,
and at day 15 and Month 1, 2, 3, 4, 5, and 6 during the 6 month
post-treatment evaluation phase.. The clinical signs (eyelid redness,
eyelid swelling, lid-debris) and eyelid irritation were evaluated.
Furthermore, a Patient Report Outcome instrument designed in
collaboration with UCLA and FDA was employed to assess the
patient’s satisfaction level during the study.
Results: Subjects randomized to the dexamethasone-azithromycin
combination arm and dexamethasone arm had a statistically
significantly (p < 0.05) greater reduction in blepharitis scores when
compared to AzaSite and/or Vehicle arms. The proportion of subjects
with the eyelid irritation score of 0 at Day 15 were significantly (p
< 0.05) higher in those dexamethasone containing arms as well.
Adverse events were reported in approximately 26% of the patients,
with an equal distribution among the corticosteroid containing
and Vehicle arms, with approximately 21% in the AzaSite arm.
Intraocular pressure increase was observed in the combination (2%),
dexamethasone alone (1%), and AzaSite (1.3%) arms.
Conclusions: The potent anti-inflammatory properties of
dexamethasone formulated in the long lasting DuraSite vehicle
appear to be efficacious in treating these important signs and
symptoms of blepharitis, which was maintained for up to 6 months
for a vast majority of the patients. Additionally, no significant safety
findings were reported.
Commercial Relationships: Kamran Hosseini, InSite Vision, Inc
(E); Richard Lindstrom, InSite Vision, Inc (C)
Clinical Trial: NCT01408082
Program Number: 3059 Poster Board Number: D0255
Presentation Time: 8:30 AM–10:15 AM
Tear osmolarity in keratoconus and allergic patients in Southern
Brazil
Leda das Neves Almeida Sandrin1, Ruth M. Santo1, Joyce H.
Takiuti1, Fábio F. Castro2. 1Ophthalmology, Faculdade de Medicina
da Universidade de São Paulo, São Paulo, Brazil; 2Allergy and
Immunology, Faculdade de Medicina da Universidade de São Paulo,
São Paulo, Brazil.
Purpose: Many studies showed ocular surface damage as well as the
presence of pro-inflammatory cytokynes in allergic and keratoconus
(KC) patients. Increased tear osmolarity is associated to ocular
surface inflammation in dry eye disease.We tested the hypothesis
that tear osmolarity and ocular surface disease index (OSDI) score
were increased and correlated positively in keratoconus and allergic
conjunctivitis (AC) when compared to control subjects (C).
Methods: This cross-sectional case control study enrolled ninetyeight patients (38 female,60 male) aged from 3 to 57 years (y):10
healthy C (median 21.5y[11;28]), 40 KC (median 11.5y [3;56]),
48 AC (median 31y [11;57]).The exclusion criteria were contact
lens wearers, use of eye drops in the last seven days, glaucoma,
pregnancy, breastfeeding and previous ocular surgery. Complete
eye examination were done. Lab-on-chip tear osmolarity (TearLab
Osmolarity System,San Diego,CA) measures were taken once in
both eyes, during afternoon period in a controlled temperature and
air humidity room. Placido-disk-based topography (EyeSysC) were
performed and power measures for K maximum (Kmax) were taken.
Patients filled in an OSDI questionnaire. ANOVA, Bonferroni and
Pearson’s tests were used to check the correlation between variables.
Results: The three groups were similar in gender (p=0,812), but
statistical significant differences were observed in age and Kmax,
which were higher (p<0,001;p<0,05) in keratoconus patients.No
difference in average tear osmolarity values observed among the
groups (p=0,058-0,064)).However, average tear osmolarity value was
higher in all groups (right eyes 310.82 +/-11.37;left eyes 313.65 +/14.52) when compared to previous data on osmolarity in healthy eyes
(302 +/- 9.7 – 18.2 mOsm/L). No correlation was observed between
OSDI score and tear osmolarity in the groups. C subjects had the
lower OSDI score and a statistical significant difference in OSDI
scores among the groups (p<0,05) was observed.
Conclusions: Our results could not find differences in average
tear osmolarity values among healthy eyes, KC and AC eyes.
Furthermore, there was no correlation between OSDI scores and tear
osmolarity.However, OSDI was able to distinguish healthy subjects
from AC and KC.Higher tear osmolarity values found in this study
might be related to particular biogeographic features of the region
where our patients live and more studies are necessary to elucidate
these findings.
Commercial Relationships: Leda das Neves Almeida Sandrin,
None; Ruth M. Santo, None; Joyce H. Takiuti, None; Fábio F.
Castro, None
Program Number: 3060 Poster Board Number: D0256
Presentation Time: 8:30 AM–10:15 AM
Assessment of 3 questionnaires for diagnosis of Ocular Surface
Disease in a Tertiary Care Ophthalmology Center in Mexico City.
Daniela Alvarez, Jaime D. Martinez, Nallely Ramos-Betancourt.
ASOCIACION PARA EVITAR LA CEGUERA EN MEXICO IAP,
Mexico City, Mexico.
Purpose: The purpose of this study was to assess 3 questionnaires
for diagnosis of Ocular Surface Disease (OSD) in a Tertiary Care
Ophthalmology Center in Mexico City.
Methods: 120 consecutive patients seen in an ophthalmologic
center in Mexico City for the first time from September 2013 to
November 2014 underwent a comprehensive examination, including
measurement of tear film break-up time (TBUT), fluorescein staining
classified by Oxford scheme, and Schirmer test type 1. Symptoms of
OSD were evaluated by the Ocular Surface Disease Index (OSDI),
Dry eye Questionnaire (DEQ-5) and McMonnies questionnaire.
Symptomatic OSD was defined as having an OSDI score ≥55, DEQ-5
score ≥12, or Mcmonnies >14. Clinical OSD was defined as having a
Schirmer test <5, TBUT <5, or staining >1.
Results: Mean patient age was 41.86 (±16.4) years (range, 18-80);
64 (53.3%) patients were female. The frequency of symptomatic
OSD based on the OSDI score ≥55 was 21.7%, DEQ-5 was 24.2%,
and Mcmonnies was 21.7%. Frequency of patients with two positive
signs and one positive questionnaire was 11.7% with OSDI score
≥55, 18.3% with DEQ-5, and 15% with McMonnies. McMonnies and
DEQ5 questionnaires had a moderate correlation wirh one another,
and OSDI a low correlation compared to the other questionnaires
(p-value <0.05). McMonnies and DEQ5 questionnaires are more
sensitive than OSDI questionnaire.
Conclusions: This is the first study to assess 3 questionnaires
for diagnosis of Ocular Surface Disease in a Tertiary Care
Ophthalmology Center in Mexico City. Frequency of OSD based on
questionnaire, and two signs plus one positive questionnaire were
really close to one another.
Commercial Relationships: Daniela Alvarez, None; Jaime D.
Martinez, None; Nallely Ramos-Betancourt, None
©2015, Copyright by the Association for Research in Vision and Ophthalmology, Inc., all rights reserved. Go to iovs.org to access the version of record. For permission
to reproduce any abstract, contact the ARVO Office at [email protected].
ARVO 2015 Annual Meeting Abstracts
Program Number: 3061 Poster Board Number: D0257
Presentation Time: 8:30 AM–10:15 AM
Lipid deposition and oxidized-low density lipoprotein in
pterygium
Dulei Zou1, 2, Liying Zhang1, 2, Changkai Jia1, 2, Tingting Liu1, Xumin
Shang2, Fan Yang2, Zuguo Liu1, 2, Wei Li1, 2. 1Eye Institute of Xiamen
UniversityFujian Provincial Key Laboratory of Ophthalmology
& Visual Science, Xiamen, China; 2Xiamen University affiliated
Xiamen Eye Center, Xiamen, China.
Purpose: Previous study has shown abnormal cholesterol metabolism
and lipid peroxidation in pterygium, this study was to investigate
whether lipid deposition and apolipoprotein peroxidation exist in
pterygium.
Methods: Primary pterygium specimens were cut into two parts
along the central long axis for frozen and paraffin embedding,
respectively. Semi-quantitative reverse transcription–polymerase
chain reaction ( RT-PCR ) and western blot ( WB ) were used to
detect low density lipoprotein receptor ( LDL-R ), Lectin-like
oxidized low density lipoprotein receptor-1 ( LOX-I ) and scavenger
receptor that binds phosphatidylsedne and oxidized lipoprotein (
SR-PSOX ). Lipid staining( sudan black B and oil red O ), specific
lipid extraction and filipin staining were used to investigate lipid
deposition. Immunofluorescence (IF) or Immunohistochemistry (
IHC ) was used to apolipoprotein ( including APOA1, B and E ),
malonaldehyde-low density lipoprotein ( MDA-LDL ), oxidized-low
density lipoprotein ( OX-LDL ), LDL-R, LOX-1 and SR-PSOX.
Results: We found that lipid staining, filipin staining, APOA1 and
APOE were positive in the head area of pteygium. MDA-LDL was
positive in the whole pterygium tissue. OX-LDL was weak positive
in the superficial parts of pterygium. LOX-1 was expressed in small
amount of the samples. LDL-R and PS-OX was not expressed in
pterygium.
Conclusions: Lipid, especially esterified cholesterol, APOA1, and
APOE were deposited at head area of pterygium. Two different
oxidized levels of LDL expressed in pterygium in different level.
Lipid deposition and apolipoprotein peroxidation may be engaged in
pterygium pathological progress.
Commercial Relationships: Dulei Zou, None; Liying Zhang,
None; Changkai Jia, None; Tingting Liu, None; Xumin Shang,
None; Fan Yang, None; Zuguo Liu, None; Wei Li, None
Program Number: 3062 Poster Board Number: D0258
Presentation Time: 8:30 AM–10:15 AM
The clinical evaluation of treatment for the ocular complications
from Stevens-Johnson syndrome and toxic epidermal necrolysis
Qin Long. Ophthalmology, Peking Union Medical College Hospital,
Chinese Academy of Medical Sciences, Beijing, China.
Purpose: To evaluate the treatment strategies for the ocular
complications from Stevens-Johnson syndrome(SJS) and toxic
epidermal necrolysis(TEN).
Methods: Ten patients with SJS/TEN were included in the study, the
severity of illness score (SIS) was identified using SCORTEN. The
severity of eye score (SES) was evaluated according to conjunctival
hyperemia, conjunctival exudate, conjunctival pseudomembrane,
corneal erosion, eyelid involving and lid margin involving. The
treatment strategies were made according to the SIS and SES
individually.
Results: The mean SIS was (n=10,1.40±1.89), the mean SES was
(n=20,7.00±3.58). According to the SIS and SES, patients at their
acute stage received systemic methylprednisolone (1.5mg-2mg/Kg)
W/O immunoglobulin(0.4g/Kg)for 3-5 days; ophthalmic cortical
steroid (0.1% or 1%) W/O 0.05%-1% Cyclosporine A eye drops to
inhibit ocular inflammation; for the eyes with severe ocular surface
involvement, two eyes received amniotic membrane application,
ten eyes received autoserum therapy. All the patients had artificial
tear application to clean the conjunctival exudate and lubricate
ocular surface. The convalescent treatment included topical 0.05%
Cyclosporine A, artificial tear application or bandage contact lens
wearing. After follow-up for 6-15 months, the ocular complication
included moderate to severe dry eye (6 eyes), symblepharon (4 eyes),
corneal neovascularization (2 eyes) and persistent corneal epithelial
defection (4 eyes).
Conclusions: Sufficient systemic and ocular cortical steroid
treatment and prompt amniotic membrane application in the patients
with severe ocular involvement appear to result in reducing the
acute ocular complications secondary to SJS/TEN. The late ocular
complications were significantly related to the SES, but not SIS.
Commercial Relationships: Qin Long, None
Program Number: 3063 Poster Board Number: D0259
Presentation Time: 8:30 AM–10:15 AM
Effects of Quercetin in a Mouse Model of Experimental Dry Eye
JaeWook Yang1, Ha Na Oh1, Jihyun Lee2, Chae Eun Kim2. 1Dept
of Ophthalmology, InJe University Busan Paik Hosp, Busan;
2
University, Ocular Neovascular Disease Research Center, Busan,
Korea (the Republic of).
Purpose: To evaluate the treatment effect of quercetin on dry eye in
a mouse model.
Methods: 0.5% quercetin eye drops were prepared by diluting 10%
quercetin with phosphate buffered saline (PBS). Experimental dry
eye model was induced in NOD.B10.H2b mice by dessicating stress.
The mice were divided into three groups according to the treatment
regimen : DS 10D (only desiccation stress for 10 days) ; PBS group;
and Quercetin group. Tear volume and corneal irregularity scores
were measured at 3, 5, 7 and 10 days after treatment. Haematoxylin
and eosin staining(H&E), periodic acid-Schiff staining(PAS) and
immunohistochemistry(IHC) were performed at the end of the
experiment.
Results: Quercetin group showed significantly increased tear volume
and decreased corneal irregularity score compared with PBS group
(p<0.05). Quercetin group kept smoothe corneal surface without
corneal epithelial detachment in H&E and indicated increased goblet
cell density in PAS. Quercetin group also showed significant declines
of TNF-α, MMP-2, MMP-9, ICAM-1 and VCAM-1 levels in IHC of
the lacrimal gland compared with PBS group.
Conclusions: Quercetin is thought to be helpful in improving the
ocular surface disorders of dry eye not only by restoring the corneal
epithelial detachment but also by increasing tear volume and goblet
cell density. Furthermore, quercetin is effective in the treatment of
dry eye related to inflammation.
Commercial Relationships: JaeWook Yang, None; Ha Na Oh,
None; Jihyun Lee, None; Chae Eun Kim, None
Support: This study was supported by a grant from the Korea
Healthcare Technology R&D Project of the Ministry of Health and
Welfare Affairs, Republic of Korea (grant #: HI12C0005).
Program Number: 3064 Poster Board Number: D0260
Presentation Time: 8:30 AM–10:15 AM
Extended Soft Bandage Contact Lenses Therapy for Ocular
Graft-Versus-Host Disease
Narae Ko1, Yichen Sun1, 3, Yoshihiro Inamoto2, Stephanie Lee2, Tueng
Shen1, 4. 1Ophthalmology, University of Washington, Seattle, WA;
2
Fred Hutchinson Cancer Research Center, Seattle, WA; 3Graduate
Institute of Clinical Medicine, National Taiwan University, Taipei,
Taiwan; 4Biomedical Engineering, University of Washington, Seattle,
WA.
©2015, Copyright by the Association for Research in Vision and Ophthalmology, Inc., all rights reserved. Go to iovs.org to access the version of record. For permission
to reproduce any abstract, contact the ARVO Office at [email protected].
ARVO 2015 Annual Meeting Abstracts
Purpose: Ocular Graft-versus-Host Disease (GVHD) affects 40-60%
of patients with allogeneic hematopoietic stem cell transplantation.
The most common symptoms, including photophobia, eye pain, and
blurring of vision, can have a large impact on daily living. We applied
extended soft bandage contact lens (BCL) on symptomatic ocular
GHVD patients and assessed its clinical outcome.
Methods: A prospective, phase II study was performed with
Institutional Review Board approval. Patients older than 18 years
old with diagnosis of GHVD defined by the NIH criteria and ocular
symptoms with NIH eye score of 2 or greater were recruited.
Extended soft bandage contact lenses were placed with antibiotic
coverage for at least 1 month. Visual acuity and slit lamp microscopy
exam were performed on initial presentation, at 2 and 4 weeks of
treatment. Statistical analysis of visual acuity was done using onetailed Z-test.
Results: Thirty-eight eyes of 19 patients with ocular GVHD
underwent contact lens placement. The mean Snellen acuity before
treatment was 20/31(LogMar 0.18). Visual acuity at 2 weeks of
contact lens wear was 20/27 (LogMar 0.13, p<0.001) and 20/30 at
4 weeks (LogMar 0.17, p>0.05). Twenty-four out of 25 eyes (96%)
which initially presented with conjunctival inflammation (chemosis,
injection, or staining) showed improvement after 2 and 4 weeks of
BCL wear. Twenty-three of 30 eyes (76.7%) with punctate epithelial
defect and 8 out of 15 (53.3%) eyes with filamentous keratopathy
showed improvement after 2 and 4 weeks of BCL wear.
Conclusions: Visual acuity improved in patients with ocular GVHD
after 2 weeks of soft BCL wear but the improvement was no longer
seen at 4 weeks. Conjunctival inflammation, punctate epithelial
defect and filmanetous keratopathy all improved after BCL wear.
These results support the use of BCL in patients with ocular GHVD.
Commercial Relationships: Narae Ko, None; Yichen Sun, None;
Yoshihiro Inamoto, None; Stephanie Lee, None; Tueng Shen, None
Clinical Trial: NCT01616056
Program Number: 3065 Poster Board Number: D0261
Presentation Time: 8:30 AM–10:15 AM
Evaluation of the Safety of Eye Wash with EyebonR Washing
Liquid
Hiroshi Fujishima1, Masaki Iwashita1, Hiroko Yano2, Miho Tanaka2,
Yasuko Santo2, Murat Dogru3. 1Ophthalmology, Tsurumi Univ School
of Dental Med, Yokohama, Japan; 2R&D Health Division, Kobayashi
Pharmaceutical Co., Ltd, Ibaraki, Japan; 3Ophthalmology, Tokyo
Dental College, Ichikawa, Japan.
Purpose: In Japan, eyewash is a popular remedy for symptoms
associated with contact lens wear and for the treatment of cedar
pollen related allergic conjunctivitis. EyebonR (Kobayashi
Pharmaceutical Co, Osaka, Japan) is a non-preserved antiinflammatory and anti-histaminic washing liquid containing
aminocapron, tocopherol acid ester, vitamin B6, chlorphenylamine,
chondroitin acid ester, and dipotassium glycyrrhizate. In this study,
we evaluated the safety of EyebonR in rabbits and human volunteers.
Methods: EyebonR eye wash was performed 9 times per a day
for 30 seconds in rabbits in one eye with isotonic sodium chloride
solution (IS) used as control for the other eye. Fluorescein and
lissamine corneal staining scores, and in vivo confocal microscopy
examinations of the cornea were performed to evaluate phenotypic
and inflammatory changes before and after eye wash. In the
human study, non-preserved EyebonR, EyebonR with preservatives,
another commercial washing solution (Rohto RizeR, Japan), and
IS with preservatives were applied in one eye for 30 seconds 6
times a day. Fluorescein and lissamine corneal staining scores, and
fluorophotometry for corneal epithelial permeability were assessed
before and after eye wash. Animal studies were conducted in
accordance with ARVO guidelines for animal experiments. Human
study was IRB reviewed and followed the tenets of the Declaration
of Helsinki. Non-paired t tests were used for statistical analyses. A p
value less than 5% was considered statistically significant.
Results: In the rabbit study, there was no statistical difference
between EyebonR and IS wash in relation to vital staining scores,
corneal epithelial cell density and inflammatory cell numbers in in
vivo confocal microscopy (p>0.05). In the human study, IS with
preservatives showed significantly higher corneal staining scores than
other solutions (p<0.05). EyebonR with preservatives also showed a
significantly higher staining but was significantly lower than IS with
preservatives (p<0.05).
Conclusions: Eye wash liquids with preservatives cause damage on
the ocular surface but preservative free liquids show significantly
lesser vital staining scores and less perturbation of corneal epithelial
permeability•barrier function. EyebonR eye wash appears to be a safe
modality with promising future applications in dry eye and allergic
eye disease.
Commercial Relationships: Hiroshi Fujishima, Kobayashi
Pharmaceutical Co., Ltd. (F); Masaki Iwashita, None; Hiroko
Yano, Kobayashi Pharmaceutical Co, Ltd (E); Miho Tanaka,
Kobayashi Pharmaceutical Co., Ltd (E); Yasuko Santo, Kobayashi
Pharmaceutical Co., Ltd (E); Murat Dogru, None
Support: Educational Grant of Otsuka Pharmaceutical Co., Ltd,
Grant of Lion Co, Ltd, Grant from Senju Pharmaceutical Co.,
Ltd, Grant of Alcon Pharmaceutical Co., Ltd, and grant of Santen
Pharmaceutical Co., Ltd.
Clinical Trial: 1127
Program Number: 3066 Poster Board Number: D0262
Presentation Time: 8:30 AM–10:15 AM
Intratarsal injection of kenacort in the treatment of severe cases
of VKC
Sihem Lazreg1, Didier Renault2, Moncef Khairallah3, Maya Benali
abdellah1. 1Blida, Cabinet Dr Lazreg, Blida, Algeria; 2Laboratories
thea, Clear not ferrand, France; 3University hospital, Monastir,
Tunisia.
Purpose: To treat refractory cases of Vkc with intratarsal injections
of kenacort
Methods: We treated Severe cases of Vkc that have already
experienced different anti allergic treatments and topical steroids
with frequent relapses and dependancies to steroids with intratarsal
injection of 40 mg of dexametasone, the follow up visits were
performed at D0, D 3, D7 and D30 ( slit lamp examination, corneal
staining, ocular pressure )
Results: 60 severe cases of Vkc 45 males, mean age 10,4+/- 3,5years,
90% bilateral, and 100% of corneal involvement . 70% mixed forms
and 15% of tarsal forms, the mean follow up was 20+/-7 months . at
D3 we had a decrease of all ocular signs (photophobia, redness and
pruritis) at D7 decrease of corneal staining and trantas nods, and at
d30, total remission of the Vkc, the mean duration of the efficacy of
the treatment was 10.4+/- 2.6 months, no adverse event was observed
.
Conclusions: Intratarsal injection of steroids is very effective
in severe and resistant cases of Vkc, especially in our countries
where this disease is very severe and frequent and when topical
cyclosporine is not available .
©2015, Copyright by the Association for Research in Vision and Ophthalmology, Inc., all rights reserved. Go to iovs.org to access the version of record. For permission
to reproduce any abstract, contact the ARVO Office at [email protected].
ARVO 2015 Annual Meeting Abstracts
evaluated for proportion of activity to develop a similar metric or
“leakage index.”
Results: Thirty-two eyes of 32 patients with diabetic retinopathy
or retinal venous occlusions (RVO) were included in this study. All
images were successfully analyzed with the automated algorithm. All
eyes (100%) displayed macular and/or peripheral leakage, while only
18 of 32 eyes presented with ischemia. Of the 32 eyes, the leakage
index was a mean of 4.4% (range: 0.08-13%). Of the 18 eyes with
ischemia, the mean ischemic index was 13.7% (range: 2.9-37.7%),
most of which was peripheral.
Conclusions: UWFA in diabetic retinopathy and RVO can be
analyzed with a computer-based segmentation algorithm to delineate
areas of ischemia and leakage. Significant pathology is noted in the
retinal periphery that may overlooked with standard angiography.
Improving our understanding of the importance and relative location
of these features (e.g., far/mid-periphery, macula) may be critical in
understanding disease burden, treatment response, and prognosis.
Further research is needed to enhance higher order identification
of various types of leakage (e.g., perivascular, generalized,
microaneurysms).
Figure 1: Automated segmentation samples of diabetic retinopathy
revealing leakage (green) and ischemia (blue).
Commercial Relationships: Sihem Lazreg, None; Didier Renault,
None; Moncef Khairallah, None; Maya Benali abdellah, None
Program Number: 3067 Poster Board Number: D0263
Presentation Time: 8:30 AM–10:15 AM
Automated Quantitative Analysis of Leakage and Ischemia for
Ultra-widefield Angiography in Retinal Vascular Disease
Carley Tanchon, Sunil K. Srivastava, Justis P. Ehlers. Cole Eye
Institute, Cleveland Clinic, Chagrin Falls, OH.
Purpose: To develop and establish an automated method for
objective quantitative analysis of leakage and ischemia parameters in
ultra-widefield angiography.
Methods: An IRB-approved retrospective review of patients
with retinal vascular disease who had undergone ultra-widefield
angiography by the Optos 200Tx System was performed. Two
independent reviewers examined the ultra-widefield fluorescein
angiograms (UWFA) for image quality and limited artifacts.
Two images were selected for each eye, one early and one late.
A computerized automated algorithm was developed to identify
areas of leakage and ischemia with co-registration of the two image
timepoints. Manual segmentation was also performed with Image J
software for validation of the computerized algorithm. Quantification
of ischemia was based on proportion of areas identified within the
overall retinal area, “ischemic index.” Similarly, leakage was also
Figure 2: Automated segmentation samples of RVO revealing leakage
(green) and ischemia (blue).
Commercial Relationships: Carley Tanchon, None; Sunil K.
Srivastava, Allergan (R), Bausch and Lomb (C), Bausch and Lomb
(R), Bioptigen (P), Synergetics (P); Justis P. Ehlers, Alcon (C),
Bioptigen (C), Bioptigen (F), Bioptigen (P), Genentech (F), Leica
(C), Leica (F), Synergetics (P), Thrombogenics (C), Thrombogenics
(F), Zeiss (C), Zeiss (F)
Support: NIH/NEI K23-EY022947-01A1 (JPE); Ohio Department
of Development TECH-13-059 (JPE)
©2015, Copyright by the Association for Research in Vision and Ophthalmology, Inc., all rights reserved. Go to iovs.org to access the version of record. For permission
to reproduce any abstract, contact the ARVO Office at [email protected].
ARVO 2015 Annual Meeting Abstracts
Program Number: 3068 Poster Board Number: D0264
Presentation Time: 8:30 AM–10:15 AM
Normative Values for Corneal Nerve Morphology Assessed Using
Corneal Confocal Microscopy: A Worldwide Normative Dataset
Mitra Tavakoli1, Maryam Ferdousi1, Ioannis Petropolus1, Andrey
Zhivov8, Dan Ziegler2, Ken Romanchuk3, Bruce Perkins4, Gordon
Smith5, Nathan Efron6, Rayaz A. Malik1, 7. 1Centre for Endocrinology
& Diabetes, University of Manchester, Manchester, United
Kingdom; 2Institute for Clinical Diabetology, German Diabetes
Center at Heinrich Heine University, Leibniz Center for Diabetes
Research, Düsseldorf, Germany; 3Alberta Children’s Hospital,
Alberta Children’s Hospital Research Institute, University of
Calgary, Canada, Calgary, AB, Canada; 4Division of Endocrinology,
Leadership Sinai Centre for Diabetes, University of Toronto, Toronto,
Canada, Toronto, ON, Canada; 5Department of Neurology, University
of Utah, Salt Lake City, USA, Salt Lake City, UT; 6Institute of
Health Biomedical Innovation, QUT, Brisbane, Australia, Brisbane,
QLD, Australia; 7Weill Cornell Medical School Qatar, Doha, Qatar;
8
Department of Ophthalmology, University of Rostock, Rostock,
Germany, Rostock, Germany.
Purpose: Corneal confocal microscopy (CCM) is a novel diagnostic
technique for the detection of nerve damage and repair in a range
of peripheral neuropathies, in particular diabetic neuropathy. To
enable clinical translation and wider use of this technique, normative
reference values are required. We have therefore undertaken a
multicenter collaboration to provide worldwide age adjusted
normative values of corneal nerve fibre parameters.
Methods: 1965 corneal nerve images from 343 healthy volunteers
were pooled from six clinical academic centers. All subjects
underwent examination with the Heidelberg Retina Tomograph
corneal confocal microscope. Images of the central corneal sub-basal
nerve plexus were acquired by each centre using a standard protocol
and analyzed by three trained examiners using manual tracing and
semi-automated software (CCMetrics). Age-trends were established
using simple linear regression, and normative corneal nerve fiber
density (CNFD), corneal nerve fiber branch density (CNBD), corneal
nerve fiber length (CNFL) and corneal nerve fiber tortuosity (CNFT)
reference values were calculated using quantile regression analysis.
Results: There was a significant linear age-dependent decrease in
CNFD (-0.164 no/mm2 per year for men,P<0.01 and -0.161 no/
mm2 per year for women, P<0.01). There was no change with age
in CNBD (+0.192 no/mm2 per year for men, P=0.26 and -0.050 no/
mm2 per year for women; P=0.78). CNFL decreased in men (-0.045
mm/mm2 per year, P=0.07) and women (-0.060 mm/mm2 per year,
P=0.02). CNFT increased with age in men (+0.044 per year, P<0.01)
and women (+0.046 per year, P<0.01). Height, weight and BMI did
not influence the 5th percentile normative values for any corneal
nerve parameter.
Conclusions: This study provides robust worldwide normative
reference values for corneal nerve parameters to be used in research
and clinical practice in the study of diabetic and other peripheral
neuropathies.
Commercial Relationships: Mitra Tavakoli, None; Maryam
Ferdousi, None; Ioannis Petropolus, None; Andrey Zhivov, None;
Dan Ziegler, None; Ken Romanchuk, None; Bruce Perkins, None;
Gordon Smith, None; Nathan Efron, None; Rayaz A. Malik, None
Support: • Manchester: This research was funded by the National
Institutes of Health (R1-05991), JDRF International (R106784)
and JDRF International (27-2008- 362). Support from the NIHR
Wellcome Trust Clinical Research Facility is acknowledged. •
Brisbane: This research was funded by the JDRF International grant
(8-2008-362). • Utah: This research was funded by R01-DK064814
and by the National Center for Advancing Translational Sciences of
the NIH 1ULTR001067. • Düsseldorf: This work was supported by
the Ministry of Science and Research of the State of North RhineWestphalia (MIWF NRW) and the German Federal Ministry of
Health (BMG). • Calgary: Part of this work was supported by the
JDRF International grant (17-2008-1032). • Toronto: This research
was funded by the JDRF International grant 17-2008-715 and
Canadian Diabetes Association Operating Grant OG-3-3123-BP.
Program Number: 3069 Poster Board Number: D0265
Presentation Time: 8:30 AM–10:15 AM
Neurotrophic keratopathy: a new therapeutic approach using a
novel matrix regenerating agent
Joao M. Gil1, Marta G. Guerra2, 1, Esmeralda Costa1, 3, Andreia
Rosa1, 3, Maria J. Quadrado1, Joaquim N. Murta1, 3. 1Ophthalmology,
Centro Hospitalar e Universitário de Coimbra, Coimbra, Portugal;
2
Ophthalmology, Centro Hospitalar Tondela Viseu, Viseu, Portugal;
3
Medicine Faculty of the University of Coimbra, Coimbra, Portugal.
Purpose: Neurotrophic keratopathy (NK) is a sight-threatening
degenerative corneal disease, characterized by reduced corneal
sensitivity, spontaneous epithelial breakdown and impairment of
corneal healing. Treatment is usually of poor or transient efficacy and
rehabilitation of the corneal healing response is rarely accomplished.
Cacicol® is a new regenerating agent (RGTA) designed to mimic
heparan sulphates, protecting from proteolysis and enabling
growth factors and cytokines to act on the injury. We performed a
prospective, non-controlled, interventional study to evaluate the
efficacy of Cacicol® in the treatment of NK.
Methods: Patients with persistent corneal neurotrophic ulcers, not
responding to other therapies, were treated with RGTA eye drops. All
started with one instillation per week, increased to two instillations
per week in the absence of ulcer area decrease. During follow-up,
slit-lamp examination, anterior segment photography and bestcorrected visual acuity were analysed. Ulcer evolution was evaluated
using software image analysis of slit-lamp photographs. Ulcer area
was calculated as proportion of total corneal area. Grading of images
was made by an independent reader, blinded to patient’s identity and
clinical status.
Results: We included 17 eyes of 17 patients. All patients except one
(94.12%) had complete ulcer healing within an average of 3.53±2.17
weeks – ranging from 1 to 8.14 weeks. The healing rate after 7 days
of treatment was 23.53% (n=4), followed by 41.18% (n=7) by day
14, 52.94% (n=9) after 21 days and 70.59% (n=12) within the first
month. Mean ulcer area decreased significantly from 13.81±16.16%
on the baseline to 4.52±7.59% on the 7th day (n=13, p=0.004) and
to 0.46±1.21% on the 14th day (n=10, p=0.04). Mean ulcer area
then stabilized – 0.79±1.32% on the 3rd week (n=8, p=0.43) and
0.48±0.33 on the 1st month (n=5, p=0.351). By the end of the 2nd
month all ulcers had healed except for one, occupying 2.10% of
corneal area. The mean number of instillations until complete healing
was 5.25±3.38. No neovascularization was observed but moderate
corneal scarring was noticed. Despite successful reepithelization,
improvement in visual acuity was not significant (p=0.157). One case
of recurrence (5.88%) ocurred one month after healing. No systemic
or local side effects were noticed.
Conclusions: RGTA (Cacicol®) appears to be an effective and safe
new alternative in the management of NK.
©2015, Copyright by the Association for Research in Vision and Ophthalmology, Inc., all rights reserved. Go to iovs.org to access the version of record. For permission
to reproduce any abstract, contact the ARVO Office at [email protected].
ARVO 2015 Annual Meeting Abstracts
Commercial Relationships: Joao M. Gil, Thea Portugal (F); Marta
G. Guerra, None; Esmeralda Costa, None; Andreia Rosa, None;
Maria J. Quadrado, None; Joaquim N. Murta, None
Program Number: 3070 Poster Board Number: D0266
Presentation Time: 8:30 AM–10:15 AM
Poly(ADP-ribose) polymerase inhibition improves corneal
epithelial innervation and wound healing in diabetic corneas
Yong-Soo Byun1, 2, Borami Kang3, Young-Sik Yoo1, 2, Choun-Ki Joo1,
2 1
. Ophthalmology and Visual Science, Catholic University of Korea,
College of Medicine, Seoul St. Mary’s Hospital, Seoul, Korea (the
Republic of); 2Catholic Institute for Visual Science, Seoul, Korea
(the Republic of); 3Division of Endocrinology and Metabolism,
Department of Internal Medicine, Catholic University of Korea,
College of Medicine, Seoul St. Mary’s Hospital, Seoul, Korea (the
Republic of).
Purpose: We evaluated the effect of PARP inhibitor,
1,5-isoquinolinediol (ISO), on the alteration of corneal epithelial
innervation in diabetes.
Methods: ISO (3mg/kg, intraperitoneal) or vehicle was administered
to streptozotocin (STZ)-induced diabetic rats for 4 weeks. The
epithelial innervation, epithelial wound healing, and corneal sensation
were evaluated in diabetic rats treated vehicle (DM rats), diabetic rats
treated with ISO (DM-ISO rats), and healthy non-diabetic (non-DM)
rats. The density of epithelial innervation was calculated separately
as nerve terminals and subbasal nerve plexus by analyzing the images
of whole mount corneas stained with β-tubulin antibody. To compare
the epithelial wound healing, the images of the epithelial defect were
obtained at 0, 18, 36 h after creating a 4 mm wound on the cornea and
the healed area of epithelial defect were measured. Corneal sensitivity
test was conducted using a Cochet-Bonnet handheld esthesiometer.
Additionally, PARP1 and poly(ADP-ribosyl)ated polymers (pADPr)
as its products, were identified in trigeminal ganglions (TGs) by
Western blot analysis and immunofluorescence staining.
Results: In DM rats, the density of epithelial nerve terminals (5.57
± 0.94%) and subbasal nerve plexus (22.08 ± 1.78 mm/mm2) was
significantly reduced, as compared to DM-ISO rats (8.64 ± 1.42%,
30.82 ± 2.01 mm/mm2) or non-DM rats (9.02 ± 1.14%, 34.77 ±
4.45 mm/mm2). The healed area (%) of epithelial defect at 18, 36 h
was significantly lower in DM rats (23.8 ± 5.2%, 53.2 ± 4.6%) than
in DM-ISO rats (43.2 ± 1.4%, 75.8 ± 2.2%) or non-DM rats (48.1
± 8.6%, 86.1 ± 3.3%). The corneal sensitivity was also decreased
in DM rats, not in DM-ISO rats. There were no differences in all
parameters between DM-ISO and non-DM rats. The abundance of
pADPr was increased in the TGs of DM rats, whereas not in the TGs
of DM-ISO or non-DM rats.
Conclusions: Diabetic corneas showed loss of epithelial innervation,
along with delayed epithelial healing and decreased corneal
sensitivity. PARP inhibition alleviated these diabetes induced
alterations in corneal epithelium.
Commercial Relationships: Yong-Soo Byun, None; Borami Kang,
None; Young-Sik Yoo, None; Choun-Ki Joo, None
Program Number: 3071 Poster Board Number: D0267
Presentation Time: 8:30 AM–10:15 AM
Comparative quantitative assessment of the human corneal
sub-basal nerve plexus by histologic staining and in vivo confocal
microscopy
Rudolf F. Guthoff1, Bhavani S. Kowtharapu1, Marina Hovakimyan1,
Stephan Allgeier2, Bernd köhler3, Karsten Winter4, Thomas Stahnke1,
Carl F. Marfurt5, Oliver Stachs1. 1Ophthalmology, University
of Rostock, Rostock, Germany; 2Institute for Applied Computer
Science/Automation, Karlsruhe Institute of Technology, Karlsruhe,
Germany; 3Karlsruhe Institute of Technology, Institute for Applied
Computer Science, Karlsruhe, Germany; 4Translational Centre for
Regenerative Medicine, University of Leipzig, Leipzig, Germany;
5
Indiana University School of Medicine-South Bend, South Bend, IN.
Purpose: The corneal sub-basal nerve plexus (SBP) offers a potential
biomarker for early diagnosis of various neurodegenerative diseases
such as diabetic peripheral neuropathy. Changes in SBP can be
assessed in vivo by non-invasive confocal laser scanning microscopy
(CLSM) observations and quantified using specific morphometric
features. However, the ability of CLSM to adequately resolve
intimate details of SBP anatomy remains incompletely understood.
The present study is designed to compare data from histological
staining of the SBP with data from in vivo CLSM to validate the
differences in parameters related to SBP quantification in healthy
humans.
Methods: The present study was conducted on 3 large scale SBP
images (area=1.68 mm2) of healthy subjects generated using a
CLSM mosaicking technique described earlier and 3 large scale
histologically stained regions of human corneal whole mounts
(area=1.65 mm2) prepared by beta-III tubulin staining method. A set
of parameters for each image section including corneal nerve fiber
length (CNFL), corneal nerve fiber density (CNFD), average corneal
nerve single fiber length (CNSFL), average weighted corneal nerve
fiber tortuosity (CNFTo), corneal nerve branch density (CNBD),
corneal nerve connection points (CNCP) and average weighted
corneal nerve fiber thickness (CNFTh) was calculated using a
dedicated, published algorithm and compared.
Results: Experiments showed considerable increases in CNFD
(259.6 vs. 1345.9 nerve fibers/mm2), CNFL (20.33 vs. 44.03 mm/
mm2), CNBD (140.6 vs. 768.3 branches/mm2) and CNCP (33.43 vs.
56.86 connections/mm2) values derived from histological staining
compared with values derived from CLSM. In contrast, CNFTh (2.22
vs. 1.35 mm) and CNFSL values were higher (74.76 vs. 33.16 mm)
in the CLSM data. No significant difference was observed in CNFTo
(0.085 vs. 0.091 mm-1).
Conclusions: We conclude from these data that the high parameter
bias for many quantitative SBP features is due to the higher
resolution of the histological staining method compared to in vivo
CLSM (1-2 mm). The latter method enables only imaging of SBP
nerve fiber bundles but not of single fibers, which are only displayed
in histology. In vivo CLSM facilitates SBP in vivo quantification for
clinical staging, but the resulting values are biased due to inherent
limitations in image resolution.
Commercial Relationships: Rudolf F. Guthoff, None; Bhavani
S. Kowtharapu, None; Marina Hovakimyan, None; Stephan
Allgeier, None; Bernd köhler, None; Karsten Winter, None;
Thomas Stahnke, None; Carl F. Marfurt, None; Oliver Stachs,
None
©2015, Copyright by the Association for Research in Vision and Ophthalmology, Inc., all rights reserved. Go to iovs.org to access the version of record. For permission
to reproduce any abstract, contact the ARVO Office at [email protected].
ARVO 2015 Annual Meeting Abstracts
Program Number: 3072 Poster Board Number: D0268
Presentation Time: 8:30 AM–10:15 AM
The impact of retinal detachment surgery and endolaser on
corneal sensation, corneal sub-basal nerve plexus, and corneal
epithelium.
Nacim Bouheraoua, linda hrarat, Otman Sandali, Jad Akesbi,
Isabelle Goemaere, Vincent M. Borderie, Laurent Laroche.
Ophthalmology, CHNO des Quinze-Vingts, Paris, France.
Purpose: To assess the effects of vitrectomy with 360° endolaser on
the human corneal sub-basal nerve plexus (SBNP) and to investigate
correlations between corneal sub-basal nerve (SBN) density, corneal
sensitivity, and corneal epithelial condition.
Methods: Prospective, observational, non-randomized study. 15
eyes of 15 patients who underwent vitrectomy with 360° endolaser
for retinal detachment (RD group) and 15 eyes of 15 patients
who underwent vitrectomy for macular hole with no endolaser
(MH group). Density of corneal sub-basal nerves, central corneal
sensitivity, and corneal epithelium thickness were assessed.
Results: Compared with baselines values, the mean sub-basal nerve
density (p<0.001), the mean corneal sensitivity (p<0.001), and the
mean corneal epithelial thickness (p=0.006) significantly decreased
6 months after surgery in the RD group. Conversely, there were no
significant differences in the mean sub-basal nerve density (p=0.34),
mean corneal epithelial thickness (p=0.19), and mean corneal
sensitivity (p=0.42) between pre-operative and 6-month values in
the MH group. The corneal sub-basal nerve density significantly
correlated with the corneal sensitivity (r2=0.47; p=0.004) and the
corneal epithelial thickness (r2=0.42; p=0.006). The post-operative
decrease in corneal sensitivity poorly correlated with the corneal
epithelial thickness (r2=0.23; p=0.045). The post-operative decrease
in corneal nerve density correlated with the laser energy delivered
during surgery (r2=0.50; p=0.002).
Conclusions: Sub-basal nerve density decreases after vitrectomy
combined with 360° endolaser. Surgeons should take care of the
long ciliary nerves intraoperatively and, when possible, avoid heavy
confluent treatment at these sites, without compromising adequate
retinopexy. When corneal anesthesia occurs, it is important to make
an early diagnosis and to start promptly an appropriate treatment to
minimize the risk of severe neurotrophic ulcer.
Commercial Relationships: Nacim Bouheraoua, None; linda
hrarat, None; Otman Sandali, None; Jad Akesbi, None; Isabelle
Goemaere, None; Vincent M. Borderie, None; Laurent Laroche,
None
Program Number: 3073 Poster Board Number: D0269
Presentation Time: 8:30 AM–10:15 AM
A novel animal model of corneal neurotization in a Thy1-GFP+
rat: studying a novel solution to neurotrophic keratitis
Joseph Catapano1, 2, Michael Willand2, Uri Elbaz3, 4, Tessa Gordon2,
Asim Ali3, 4, Gregory H. Borschel1, 2. 1Surgery, University of Toronto,
Toronto, ON, Canada; 2Plastic and Reconstructive Surgery, The
Hospital for Sick Children, Toronto, ON, Canada; 3Ophthalmology
and Vision Sciences, The Hospital for Sick Children, Toronto, ON,
Canada; 4Ophthalmology and Vision Sciences, The University of
Toronto, Toronto, ON, Canada.
Purpose: Corneal anesthesia renders the cornea susceptible to occult
injury leading to neurotrophic keratitis, progressive scarring and
vision loss. Standard medical therapy often fails to prevent disease
progression. New surgical techniques involving nerve transfers and
grafts to reinnervate the cornea in humans has restored protective
sensation and improved visual acuity. Establishing an animal
model is necessary to investigate factors that mediate corneal nerve
regrowth. Here we present a surgical model of corneal denervation
in aThy1-GFP+ rat model, which can then be used to surgically
reinnervate the cornea.
Methods: The Thy1-GFP+ rat strain expresses green fluorescent
protein (GFP) in all neurons and axons, permitting reliable imaging
of corneal nerves. A perilimbal incision was used to identify the
ciliary nerves which were then transected lateral to the optic nerve
and inferior to the lateral rectus muscle. Seven days following
surgical denervation, confocal microscopy was used to image both
the denervated cornea and contralateral intact cornea with a 10x
objective.
Results: All rats tolerated the procedure well. Corneal nerve imaging
in the Thy1-GFP+ rat strain with confocal microscopy permitted
tracing of stromal, sub-basal and epithelial nerves within the cornea
(Figure 1A). Seven days after injury and transection of the ciliary
nerves, denervation was complete (Figure 1B).
Conclusions: Surgical corneal neurotization improves corneal
sensation and ocular defense mechanisms, protecting vision in
patients who would otherwise lose vision. We have demonstrated
that the Thy1-GFP+ rat cornea can be effectively denervated through
a minimally invasive intraocular approach and that corneal nerves
can be reliably image with whole mount confocal microscopy. The
Thy1-GFP+ rat can serve as the basis for a model to investigate
corneal nerve regrowth following surgical neurotization, permitting
the investigation and identification of treatments that may further
potentiate nerve regrowth and improve clinical results.
Figure 1. (A) GFP+ expression allows reliable tracing of corneal
nerves with confocal imaging. (B) 7 days following transection
there is complete loss of axons. Dotted lines indicate the periphery
of the sample. (Note: cornea has been cut in four locations to enable
imaging in one focal plane).
Commercial Relationships: Joseph Catapano, None; Michael
Willand, None; Uri Elbaz, None; Tessa Gordon, None; Asim Ali,
None; Gregory H. Borschel, None
Support: Canadian Institute of Health Research, Plastic Surgery
Foundation
Program Number: 3074 Poster Board Number: D0270
Presentation Time: 8:30 AM–10:15 AM
CO2-Induced Corneal Nociceptor Stimulation Reveals
Neuropeptide Changes in a Novel Murine Pain Model
Yashar Seyed-Razavi1, Pedram Hamrah1, 2. 1Department of
Ophthalmology-Harvard Medical School, Schepens Eye Research
Institute/Massachusetts Eye and Ear, Boston, MA; 2Department of
Ophthalmology-Harvard Medical School, Cornea and Refractive
Surgery Service-Massachusetts Eye and Ear Infirmary, Boston, MA.
Purpose: The cornea is highly innervated with sensory nociceptors,
the stimulation of which result in the sensation of pain and defensive
reflex actions, such as blinking and tear secretion. Following
stimulation, nociceptors also release neuropeptides which in
turn can affect cell proliferation and migration, vasodilation and
©2015, Copyright by the Association for Research in Vision and Ophthalmology, Inc., all rights reserved. Go to iovs.org to access the version of record. For permission
to reproduce any abstract, contact the ARVO Office at [email protected].
ARVO 2015 Annual Meeting Abstracts
permeability of vessels, and promote wound healing after injury. To
our knowledge, there have been no studies examining neuropeptide
expression alterations in the cornea following induction of pain and
in the absence of tissue injury.
Methods: A modified version of the Belmonte CO2 esthesiometer
was developed for murine use to investigate nerve stimulation
in the absence of injury. In short, one or three series of pulses of
100% CO2 gas, triplicates of 5 seconds (3x5 seconds) duration each,
were applied to the central cornea of mice with 10 minute intervals
between stimuli. Blink was induced manually every 2 minutes during
the procedure to avoid dryness. Corneas were excised 10 minutes
following final CO2 application and processed for neuropeptide
gene quantification by real-time PCR. Neuropeptides analyzed
include substance P (SP), nerve growth factor (NGF), urocortin and
secretoneurin (SN). Naïve unstimulated corneas served as controls.
Results: CO2 administration to the central cornea successfully
stimulated polymodal nociceptors of the murine cornea where
a blinking response indicative of pain was observed in all mice.
Substance P mRNA expression significantly increased in both the one
and three CO2 pulse groups (2.1 and 4.1 fold change respectively,
p<0.05) compared to non-stimulated control corneas with SP
expression increasing with the number of pulses; this increase was
not significant (p=0.07). Further, NGF expression increased (1.7
fold, p<0.01), whereas urocortin and secretoneurin expression levels
decreased following CO2 stimulation as compared to non-stimulated
control corneas (0.5 fold change, p<0.01 and 0.4 fold change, p<0.05,
respectively). Fluorescein staining of the cornea revealed these
changes were not due to epithelial damage or dry eye.
Conclusions: Our study demonstrates immediate changes in corneal
neuropeptides including increase in SP and NGF expression levels
after CO2 induced pain. This murine model may serve as the basis for
future studies into understanding the pathophysiology of peripheral
pain.
Commercial Relationships: Yashar Seyed-Razavi, None; Pedram
Hamrah, None
Support: NIH R01-EY022695 (PH), Research to Prevent Blindness
Career Development Award (PH), Falk Medical Research Trust (PH),
MEEI Foundation (PH)
Program Number: 3075 Poster Board Number: D0271
Presentation Time: 8:30 AM–10:15 AM
Pigment epithelial derived factor (PEDF) plus docosahexaenoic
acid (DHA) induce a dual response of the immune system and
increase nerve regeneration in HSV-1-infected corneas
Maria S. Cortina1, Jiucheng He2, Neumann Donna3, Farhana
Musarrat3, Azucena H. Kakazu2, Haydee E. Bazan2. 1ophthalmology
and visual sciences, University of Illinois Eye & Ear Infirmary,
Chicago, IL; 2Ophthalmology and Neuroscience, Louisiana State
Univeristy, New Orleans, LA; 3Pharmacology, Lousiana State
University, New Orleans, LA.
Purpose: HSV-1 infection is a major cause of corneal blindness in
the USA. Associated complications include persistent inflammation
and neurotrophic keratitis. Studies using a rabbit model infected
with HSV-1 and treated with PEDF+DHA have shown increased
corneal sensitivity and reduced inflammation (ARVO 2014). Here we
investigate the inflammatory-immune response during the first two
weeks of primary HSV infection and the cornea nerve density and
presence of lymphocytes in the trigeminal ganglion (TG) during viral
latency.
Methods: Rabbits were inoculated with 100,000 pfu/eye of HSV-1
17 Syn+ strain. Twenty-four hours post inoculation (p.i.), treatment
with PEDF+DHA or vehicle was started. Animals were divided into
3 groups, and euthanized at 7, 14 days and 12 weeks. Corneas from
the first two groups were stained with antibodies against CD8+,
CD4+, CD11b, F40/8 and neutrophils. The number of cells with
positive staining was recorded for each group. In the third group,
corneal sensitivity was measured every week with a Cochet Bonet
esthesiometer. Corneas and trigeminal ganglia (TG) were removed
and corneal specimens were stained with βII-neurotubulin antibody
and TG with CD8+ antibody.
Results: At one week p.i., there was a very significant increase in
lymphocytes, macrophages and neutrophil infiltration in the rabbits
treated with PEDF+DHA that was reverted at 2 weeks. Corneal
lesions were significantly decreased in the PEDF+DHA group at
14 days. No corneal sensation was detected in rabbits treated with
vehicle for 12 weeks; however, corneal sensitivity started to recover
by week 3 in the PEDF+DHA-treated rabbits. After 12 weeks p.i.,
corneal nerve density was increased in the PEDF+DHA group, and
the TG of the rabbits contained CD8+ cells surrounding the axons.
Conclusions: The innate immune-inflammatory response is enhanced
by PEDF+DHA treatment during the early phase of viral infection,
probably to increase HSV-1 clearance from the cornea. This immune
modulation is likely to mediate, at least in part, the decrease in
corneal lesions, recovery of sensitivity and nerve regeneration
observed with treatment. The presence of CD8+ lymphocytes In
the TG has the potential to inhibit virus reactivation and suggests a
protective role of PEDF+DHA against recurrent disease.
Commercial Relationships: Maria S. Cortina, None; Jiucheng He,
None; Neumann Donna, None; Farhana Musarrat, None; Azucena
H. Kakazu, None; Haydee E. Bazan, None
Support: R01 EY 019465, and unrestricted departmental grant from
Research to Prevent Blindness
Program Number: 3076 Poster Board Number: D0272
Presentation Time: 8:30 AM–10:15 AM
Metabolic syndrome affects mouse corneal epithelium and nerve
morphology
Aubrey Hargrave1, Pooja Mehta2, Paul Landry1, Hunyh Amanda1,
Maria Dupre1, Siri Magadi1, Zhijie Li2, Clifton W. Smith2, Alan R.
Burns1, 2. 1College of Optometry, University of Houston, Houston,
TX; 2Ped-Children’s Nutrition Rsrch Ctr, Baylor College of
Medicine, Houston, TX.
Purpose: Diet-induced obesity is accompanied by a metabolic
syndrome that causes systemic inflammatory changes. The metabolic
syndrome is a cluster of disorders that include abdominal obesity,
dyslipidemia, hypertension, elevated fasting glucose levels, and
insulin resistance progressing to type 2 diabetes. Corneal nerve
damage is an early event in the progression of diabetic keratopathy,
a condition affecting approximately 70% of diabetics. This study
uses metabolic syndrome as a model to investigate early changes in
corneal anatomy that precede the development of type 2 diabetes.
Methods: Five week old C57BL/6 mice were fed a high fat milk
diet (HFD, 42% kcal) for 10 weeks and were subsequently analyzed
for systemic inflammatory and metabolic changes. Mice on a normal
chow diet served as controls. Excised corneas were labeled with
nerve-specific anti-tubulin β III antibody and nuclei were stained
with DAPI. Two central and four paracentral regions per cornea
were imaged by immunofluorescence microscopy. The effect of the
HFD on corneal nerve morphology and epithelial basal cell density
was evaluated using ImageJ and a custom MATLAB program. Data
analysis was performed using an unpaired two-tailed t-test. A p-value
of ≤ 0.05 was considered significant.
Results: Mice on a HFD had a 35% decrease in central epithelial
vertical branch density (p=0.019) and a 42% decrease in epithelial
and basal nerve density (p=0.006 and p=0.026, respectively),
compared to control mice. Paracentral nerve density was not different
©2015, Copyright by the Association for Research in Vision and Ophthalmology, Inc., all rights reserved. Go to iovs.org to access the version of record. For permission
to reproduce any abstract, contact the ARVO Office at [email protected].
ARVO 2015 Annual Meeting Abstracts
between the two groups. After 10wk on a HFD, mice had a significant
increase in central epithelial basal cell density, but not paracentral
basal cell density, compared to mice on a chow diet (p=0.0006).
Conclusions: Diet-induced metabolic syndrome is accompanied
by early anatomical changes in corneal nerves and in the corneal
epithelium. These prediabetic changes occur in the central cornea
prior to the development of hyperglycemia and insulin resistance,
raising the possibility that these changes are linked to the systemic
inflammation caused by the metabolic syndrome.
Commercial Relationships: Aubrey Hargrave, None; Pooja
Mehta, None; Paul Landry, None; Hunyh Amanda, None; Maria
Dupre, None; Siri Magadi, None; Zhijie Li, None; Clifton W.
Smith, None; Alan R. Burns, None
Support: EY07551, EY018239, USDA/ARS 6250-51000-055
Commercial Relationships: Alan R. Burns, None; Thao Do, None;
Paul Landry, None; Ali Behzad, None; Samuel D. Hanlon, None
Support: NIH Grants EY007551
Program Number: 3077 Poster Board Number: D0273
Presentation Time: 8:30 AM–10:15 AM
Ultrastructural details of corneal nerve mitochondria revealed by
serial block face imaging and 3D reconstruction
Alan R. Burns1, 2, Thao Do1, Paul Landry1, Ali Behzad3, Samuel D.
Hanlon1. 1College of Optometry, University of Houston, Houston,
TX; 2Pediatrics, Baylor College of Medicine, Houston, TX; 3Imaging
and Characterization Core Lab, King Abdullah University of Science
and Technology, Thuwal, Saudi Arabia.
Purpose: The corneal epithelium is richly supplied with nerve fibers
which are not only essential for sensation but also for maintaining
the health/integrity of the epithelium and cornea as a whole. The
plexus of nerves is richly invested with mitochondria. Mitochondrial
dysfunction/degeneration has been described in neuropathies such as
diabetic peripheral neuropathy (DPN). Standard transmission electron
microcopy, when coupled with stereology, can be used to estimate
mitochondrial number, volume, density, and distribution within nerve
fibers, but is limited by the difficulty encountered in obtaining large
numbers of serial sections. The purpose of this study was to explore
serial block face imaging as a novel ultrastructural method for 3D
reconstruction and analysis of corneal nerves.
Methods: Corneas from C57BL/6 mice between 8-12 weeks of age
were fixed and embedded in resin blocks for histological sectioning.
Serial block-face images (100 nm intervals with 500-1000 images/zstack) in perfect registration were obtained using an ultramicrotome
(Gatan 3View) system mounted within the base of a scanning
electron microscope. From the z-stacks generated, stromal, basal,
and epithelial corneal nerves were manually segmented, as were the
mitochondria within the nerves. Amira 5.2 software was used for
3D reconstruction and to evaluate potential means for characterizing
nerves/mitochondria and obtaining quantifiable data.
Results: Large stromal nerves were easily recognized whereas small
epithelial fibers were often difficult to discern and did not become
evident until scrolling through the stack of images. Small basal
nerve fibers from the horizontal plexus just anterior to the epithelial
basement membrane made occasional connections through the
basement membrane to stromal fibers. Vertical extensions of fibers
from the basal plexus within the epithelial cells were relatively more
common. Quantification of nerve length, mitochondrial surface
area, volume density, and number within nerve fibers was easily
accomplished with the Amira 5.2 software.
Conclusions: Reconstruction of serial block-face images not only
provided, for the first time, the means to observe 3D arrangements
of mitochondria within corneal nerves but also to obtain quantitative
data. Serial block face imaging has the potential for documenting
mitochondrial changes that accompany a variety of corneal
pathologies.
©2015, Copyright by the Association for Research in Vision and Ophthalmology, Inc., all rights reserved. Go to iovs.org to access the version of record. For permission
to reproduce any abstract, contact the ARVO Office at [email protected].