Transmembrane Mucin Response in Conjunctival Epithelial Cells Exposed to Wall Shear Stresses.

Human conjunctival epithelium cells (HCEC) line the inner surface of the eyelid and cover the sclera and are continuously subjected to wall shear stresses (WSS). The effects of external forces on the conjunctival epithelium are not fully known. The conjunctival epithelium contains stratified squamous cells that synthesize the membrane-spanning mucins MUC1 and MUC16, which play important roles in protecting the ocular surface. Alterations in both gel-forming and membrane-tethered mucins occur in drying ocular surface diseases. The aim of this study was to explore the mechanobiological characteristics of transmembrane mucin secretion and cellular alterations of primary HCEC exposed to airflow-induced WSS perturbations. We exposed the HCEC to a steady WSS of 0.5 dyne/cm2 for durations of 15 and 30 min. Cytoskeletal alterations and MUC1 secretions were studied using immunohistochemically fluorescent staining with specific antibodies. We investigated for the first time an in vitro model of membrane-tethered mucin secretion by HCEC in response to WSS. The exposure of HCEC to WSS increased the polymerization of F-actin, altered the cytoskeletal shape and reduced the secretion of membrane-tethered MUC1.

Mechanobiology of conjunctival epithelial cells exposed to wall shear stresses.

The human conjunctival epithelial cells (HCEC) line the inner sides of the eyelids and the anterior part of the sclera. They include goblet cells that secret mucus into the tear film that protects the ocular surface. The conjunctival epithelium is subjected to mechano-physical stimuli due to eyelid movement during blinking, during wiping and rubbing the eyes, and when exposed to wind and air currents. We cultured primary HCEC under air-liquid interface (ALI) conditions in custom-designed wells that can be disassembled for installation of the in vitro model in a flow chamber. We exposed the HCEC after ALI culture of 8-10 days to steady and oscillatory airflows. The in vitro model of HCEC was exposed to steady wall shear stresses (sWSS) of 0.5 and 1.0 dyne/cm2 for lengths of 30 and 60 min and to oscillatory wall shear stresses (oWSS) of 0.5 and 0.77 dyne/cm2 amplitudes for a length of 10 min. Cytoskeletal alterations and MUC5AC mucin secretion in response to WSS were investigated using immunohistochemically fluorescent staining and enzyme-linked lectin assay (ELLA), respectively. The results revealed that both exposure times and sWSS values increased the polymerization of F-actin filaments while mucin secretion decreased. However, after a recovery of 24 h in the incubator we observed a decrease of F-actin fibers and mucin secretion only for exposure of 30 min. The length of exposure was more influential on cytoskeletal alterations than the level of sWSS. The very small effect of sWSS on mucin secretion is most likely related to the much smaller amount of goblet cell than in other mucus-secreting tissue. The results for both oWSS amplitudes revealed similar trends regarding F-actin and mucin secretion. Immediately post-exposure we observed an increase in polymerization of F-actin filaments while mucin secretion decreased. However, after 24-h recovery we observed that both F-actin and mucin secretion returned to the same values as for unexposed cultures. The results of this study suggest that WSS should be considered while exploring the physiological characteristics of HCEC.

Kelly Punch Punctoplasty and Simple Punctal Dilatation Are Equally Effective for Punctal Stenosis: A Prospective Study.

PURPOSE: To examine the clinical, imaging, and histopathologic differences between Kelly punch punctoplasty and punctal dilatation, both with mini Monoka stent insertion. METHODS: This is a prospective, comparative clinical study. Patients with bilateral punctal stenosis and for whom surgery was advised participated in the study. The right eye underwent Kelly punch punctoplasty and the left eye simple punctal dilatation, both with mini Monoka stents were participated. Data were collected and analyzed before and 3 months following the procedure and included the degree of epiphora (Munk score) and punctal size as measured by anterior segment optical coherence tomography (AC- OCT). All specimens removed by the Kelly punch punctoplasty were sent for histopathologic evaluation, including Masson's trichome muscle staining. RESULTS: The study included 46 eyes of 23 patients (4 males and 19 females) whose mean age at presentation was 60.43 ± 14.81 years (range 19-84 years). There was a significant decrease in the Munk score after both punctoplasties compared with baseline values (P < 0.01, matched pairs). There were no significant differences between groups in the delta Munk score. There was a significant increase in punctal size after both procedures compared with baseline values as measured by AS-OCT. Horner-Duverney's muscle was visible in only 3 of the 23 (13.04%) specimens from patients who underwent the Kelly punch punctoplasty. CONCLUSIONS: The Kelly punch punctoplasty and the simple dilator punctoplasty, both with stents, are equally effective treatments for epiphora due to acquired punctal stenosis. Only a few specimens removed during the Kelly punch method show muscle fibers.

The distribution of relaxin receptors in the anterior segment of primary open-angle glaucoma patients.

Purpose: Relaxin is a polypeptide hormone produced by the corpus luteum and the decidua in females and by the prostate in males. It has vasodilatory and antifibrotic effects. In the past, it has been reported that injection of relaxin hormone has caused a reduction in the intraocular pressure although its mechanism has not yet been fully understood and the expression of relaxin receptors has not yet been evident in the human eye. Therefore, the purpose of this current study was to examine the presence of relaxin receptors in the human eye anterior segment. Methods: This is an interventional non-randomized study. Patients with primary open-angle glaucoma who underwent trabeculectomy were recruited. During the surgery, a punch biopsy (including the cornea, sclera, trabecular meshwork, and Schlemm's canal) was taken and stained with hematoxylin-eosin, and immunohistochemistry staining for relaxin/insulin-like family peptide receptor 1 (RXFP1). Results: A positive staining was noted for RXFP1 in the inner uveal, corneoscleral, and cribriform meshwork and Schlemm's canal's endothelium. Negative staining for RXFP1 was noted in the cornea and sclera. Conclusion: RXFP1 is present in the anterior segment drainage system of the human eye. Therefore, this receptor may have an active role in regulating the outflow facility and in the reduction of intraocular pressure.

Effect of Rho-associated kinase inhibitor on human corneal endothelial cell apoptosis.

PURPOSE: To evaluate whether exposure to Rho-associated protein kinase (ROCK) inhibitor will promote human-cultured corneal endothelial cells (CECs) survival in a commercial storage medium. SETTING: Edith Wolfson Medical Center, Holon, and Sheba Medical Center, Tel Hashomer, Israel. DESIGN: Experimental study. METHODS: Fragments of human donor corneolimbal rings were stored in commercial storage media for 1 week, half with the addition of 10 µM ROCK inhibitor (Y-27632). Evaluation of CECs for early and late apoptosis\necrosis rates was performed using anti-human CD166 antibody and flow cytometric double staining analysis of propidium iodide and Annexin V. RESULTS: CECs of 6 corneolimbal rings demonstrated a reduced early apoptosis rate (4.35% ± 1.07% vs 12.18% ± 5.5%, P = .026) and a reduced late apoptosis\necrosis rate (5.5% ± 2.39% vs 9.43% ± 2.61%, P = .004) compared with control. Subsequently, the rate of apoptotic CECs expressing ROCK was significantly lower in cells exposed to ROCK inhibitor compared with cells that were not (19.01% ± 4.17 vs 30.42% ± 4.27, P < .001). CONCLUSIONS: ROCK inhibitor reduced endothelial cell loss in vitro and might be used to limit or slow CEC loss in donor corneal tissue during eye banking. This might be a promising new method for promoting future graft survival.

The clinical and histopathological characteristics of Kelly punch punctoplasty.

PURPOSE: To examine the clinical presentation, histopathological characteristics and surgical success of patients that underwent Kelly punch punctoplasty. METHODS: A retrospective consecutive case series of all patients diagnosed with punctal stenosis and referred to Kelly punch punctoplasty and mini-monoka insertion between January 2017 and December 2017. The demographic, clinical and prognosis parameters were retrieved and analyzed. Moreover, a histopathological examination was performed on the tissue retrieved from the Kelly punch at the end of the procedure in order to evaluate the presence of muscle in the tissue, as well as chronic inflammation. RESULTS: The series was composed of 44 eyes of 28 patients. All patients' complaints prior to surgery were epiphora: 18 patients had bilateral epiphora, 5 presented with right-eye epiphora and 5 with left-eye epiphora. The average Munk degree of epiphora before surgery was 3 and at least 3 months after surgery 1.04 (matched pairs, p < 0.0001). Two patients underwent the second punctoplasty. In haematoxylin and eosin staining, the Riolan muscle was not visible in the mucosal layer in 40 specimens. In four specimens, the muscle was observed. A Masson's trichrome staining ensured that the Riolan muscle existed only in those four specimens. DISCUSSION: Kelly punch punctoplasty is a simple and minimally invasive procedure for punctal dilation with high functional success rate. Moreover, in histopathological specimens, in all but four of the cases there was no muscle indicated. Intact Riolan muscle enables the punctum to stay in the right circular shape and position, and the lacrimal pump to maintain its pressure gradient.

Strong bonding of corneal incisions using a noncontact fiber-optic laser soldering method.

Suturing of corneal incisions requires significant skill. We demonstrate a noncontact method that will simplify the bonding process. 5-mm-long penetrating vertical and slanted incisions were made in corneas of eyes, extracted from dead piglets. A fiber-optic laser system was used for laser soldering of the incisions, under close temperature control, using albumin solder. The burst-pressure PB immediately after the soldering was found to be PB ≈ 92 and 875 mmHg, for vertical and slanted incisions, respectively. PB = 875 mmHg is an exceptionally high figure, ≈10 times the clinically acceptable value for sutured incisions. Laser soldering was then performed on penetrating incisions made in the corneas of live healthy piglets, of weight ≈10 Kg. After a healing period, the eyes were extracted, and the corneas were examined by histopathology and by optical coherence tomography. Our method immediately generated watertight and strong bonding without noticeable corneal shape distortion. These results would be beneficial for cataract surgery and for corneal transplantations. The fiber-optic system makes it much easier to bond corneal incisions. In the future, laser soldering could be automated and efficiently used by less experienced surgeons, thereby reducing the workload on the experienced ones.

Yellow subconjunctival deposits.

In this case report, we present a healthy man who was referred for removal of subconjunctival yellow lesions found during a routine eye examination. In histopathological examination, an amyloidosis was found. There were no remnants or new lesions during 1-year follow-up. There was no systemic involvement. Conjunctival amyloidosis is a rare diagnosis that often is overlooked. Failure to recognise conjunctival amyloidosis might lead to late diagnosis of systemic amyloidosis. This case may rise the awareness to this rare diagnosis.

Spatial visual function in anomalous trichromats: Is less more?

Color deficiency is a common inherited disorder affecting 8% of Caucasian males with anomalous trichromacy (AT); it is the most common type of inherited color vision deficiency. Anomalous trichromacy is caused by alteration of one of the three cone-opsins' spectral sensitivity; it is usually considered to impose marked limitations for daily life as well as for choice of occupation. Nevertheless, we show here that anomalous trichromat subjects have superior basic visual functions such as visual acuity (VA), contrast sensitivity (CS), and stereo acuity, compared with participants with normal color vision. Both contrast sensitivity and stereo acuity performance were correlated with the severity of color deficiency. We further show that subjects with anomalous trichromacy exhibit a better ability to detect objects camouflaged in natural gray scale figures. The advantages of color-deficient subjects in spatial vision performance could explain the relatively high prevalence of color-vision polymorphism in humans.

Prophylactic exposure of human corneal endothelial cells to Rho-associated kinase inhibitor reduced apoptosis rate after phacoemulsification: Ex vivo study.

PURPOSE: To evaluate whether prophylactic exposure of corneal endothelial cells (CECs) to a selective Rho-associated kinase (ROCK) inhibitor will inhibit CEC apoptosis after phacoemulsification. SETTING: Laboratory evaluations at the Edith Wolfson Medical Center, Holon, Israel and the Chaim Sheba Medical Center, Tel-Hashomer, Ramat-Gan, Israel and the Chaim Sheba Medical Center, Tel-Hashomer, Ramat-Gan, Israel. DESIGN: Experimental study. METHOD: Human donor corneolimbal rings were divided into fragments that were stored in commercial storage media with or without the addition of 10 mM ROCK inhibitor for 1 week and were then exposed to phacoemulsification energy. Samples were dissociated into single cells by trypsin digestion and CECs were targeted using the antihuman CD166 antibody, a new biomarker. The CEC survival was evaluated for early and late apoptosis rate with flow cytometric analysis of annexin-V and propidium iodide (PI) double staining. RESULTS: Six corneoscleral rings from 4 donors were studied. After phacoemulsification, CEC exposed to ROCK inhibitor demonstrated a 37.06% reduction in early apoptosis rate (29.36% ± 4.33% [SD] versus 46.65% ± 1.51%, P = .006) and 45.27% reduction in late apoptosis rate (17.6% ± 16.81% versus 32.16% ± 26.30%, P = .007), compared with controls. Subsequently, ROCK levels in apoptotic CECs were significantly lower in cells incubated with ROCK inhibitor than the control medium. CONCLUSIONS: In this ex vivo study, ROCK inhibitor reduced endothelial loss and thus, could be used to limit or slow down CEC loss. Rho-associated kinase inhibitor might be used before cataract surgery, especially in high risk patients. This might be a promising new method for preventing pseudophakic bullous keratopathy.

Periocular molluscum contagiosum: six different clinical presentations.

PURPOSE: To describe the different clinical presentations of periocular molluscum contagiosum (MC) lesions and their epidemiological, clinical and histopathological features. METHODS: Medical records and histopathological sections of all cases of periocular MC treated at the oculoplastic clinic of the Goldschleger Eye Institute, Sheba Medical Center, Israel, between 1995 and 2016 were retrospectively reviewed. The following data were extracted: gender, age at the time of MC diagnosis, immune competency, location of the periocular lesions, number of lesions, dimensions of the lesions, clinical presentation, histopathological features, suspected clinical diagnosis before histopathological diagnosis and treatment. RESULTS: The series was composed of 41 patients (19 males, 22 females) whose mean age at presentation was 20.41 â€Š± â€Š21.10 years (range 1-71 years). Only one patient was immunosuppressed. The cases were classified into six proposed clinical presentations: 'umbilicated nodular', 'big/giant', 'conglomerated', 'erythematous', 'inflamed' and 'pedunculated'. CONCLUSION: This is the first time that different clinical types of MC lesions are labelled. The current evidence also indicates that MC lesions should be suspected not only in children and in immunosuppressed adult patients but also in immunocompetent patients of all ages.

The Epidemiological, Clinical, and Histopathological Characteristics of Lacrimal Gland Biopsies in a Tertiary Care Center in Israel.

BACKGROUND: The distribution of pathology and clinical characteristics of lacrimal gland diseases are different in different areas of the world. OBJECTIVES: To evaluate the incidence rate, patient characteristics, and indications for surgical intervention of lacrimal gland lesions in a tertiary care center in Israel. METHODS: All biopsied or surgically removed lacrimal gland lesions at the Goldschleger Eye Institute from 2009 to 2015 were identified. The following data were collected: age, gender, indications for surgical intervention, diagnosis, treatment, and prognosis. RESULTS: We evaluated 28 lacrimal gland biopsies from 26 patients (11 men, 15 women). Mean age at biopsy was 47.5 years old. The most common presenting symptoms were: eyelid swollenness (57.14%), ptosis (32.14%), and proptosis (10.71%). All patients underwent computed tomography and magnetic resonance imaging. In 28 cases, infiltrations of the lacrimal gland were found. In nine cases infiltration of muscles or orbital extension were found. The most common pathologies were non-specified inflammation (44.82%), lymphoma (20.68%), and immunoglobulin G4-related disease (10.34%). The treatment was diverse according to the patient diagnosis. Prognosis of lacrimal gland disease was good; however, in five patients the systemic disease progressed. CONCLUSIONS: Lesions of the lacrimal gland comprise a wide variety of pathological findings that require different treatment strategies. Lacrimal gland biopsies enable physicians to precisely recognize the pathology; therefore, it is important to consider this surgical method in any patient with lesions in the lacrimal gland.

An Unusual Presentation of Conjunctival Dermolipoma With a Skin Tag.

Dermolipoma is considered the second most common benign conjunctival tumor in children after nevus. Usually, dermolipoma located just deep in the superotemporal bulbar and forniceal conjunctiva. The authors describe an uncommon presentation of dermolipoma that arise in the conjunctiva as well as in the skin of the lateral canthus. A 5-month-old baby with lateral limbal lesion involving the conjunctiva and the skin underwent excision biopsy. The pathological examination found a dermolipoma in the conjunctiva as well as in the dermis. The outcomes of the excision were satisfactorily functional and cosmetic.This is only the second case of dermolipoma in the dermis in normal baby that described those far in the literature. This case should raise the awareness that dermolipoma should be considered in differential diagnosis of lesions of the lateral cantus and the periocular skins near it.

Feasibility of using experimental high viscosity silicone oils: a pilot study.

BACKGROUND: Polydimethyl silicones (silicone oils) are used in complex retinal surgeries, including difficult or recurrent retinal detachments, severe eye trauma, and other indications for long term tamponade. Two major problems with currently available silicone oils are emulsification and recurrent retinal detachment. The primary endpoint of this study was to evaluate the toxicity and safety of high viscosity silicone oils and second, the feasibility of using them with currently available vitrectomy systems. METHODS: In this experimental study, 8 eyes of 8 New Zealand White rabbits underwent vitrectomy. The vitreous cavities of 2 eyes were filled with medical grade 5500 cS silicone oil, 4 eyes with 12,500 cS oil, and 2 eyes with 30,000 cS oil for 3 months, after which the animals were sacrificed and the eyes sent for histopathological examination. The duration required to inject 5 cc each of 1300, 12,500 and 30,000 cS oils, using a commercially available system (Alcon VFC Pac) were also evaluated. RESULTS: Retinal histopathology was comparable in all eyes, with no excess toxic effect or damage seen in eyes with experimental oils. All oils were readily injectable with the VFC Pac system. CONCLUSIONS: High viscosity experimental silicone oils have similar chemical and physical properties to lower viscosity oils currently used. Therefore, toxicities are expected to be similar. In a small pilot sample of 8 rabbit eyes filled with silicone oil for 3 months, histopathology in eyes with 12,500, 30,000 cS or medical grade 5500 cS silicone oil were similar. Injecting these oils using available vitrectomy equipment is feasible. New, high viscosity silicone oils may offer several advantages over currently available oils for some patients.

Long-Term Safety of Transplanting Human Bone Marrow Stromal Cells into the Extravascular Spaces of the Choroid of Rabbits.

Incurable neuroretinal degeneration diseases cause severe vision loss and blindness in millions of patients worldwide. In previous studies, we demonstrated that transplanting human bone marrow stromal cells (hBMSCs) in the extravascular spaces of the choroid (EVSC) of the Royal College of Surgeon rats ameliorated retinal degeneration for up to 5 months. Assessing the safety of hBMSC treatment and graft survival in a large animal is a crucial step before initiating clinical trials. Here, we transplanted hBMSCs into the EVSC compartment of New Zealand White rabbits. No immunosuppressants were used. Transplanted cells were spread across the EVSC covering over 80 percent of the subretinal surface. No cells were detected in the sclera. Cells were retained in the EVSC compartment 10 weeks following transplantation. Spectral domain optical coherence tomography (SD-OCT) and histopathology analysis demonstrated no choroidal hemorrhages, retinal detachment, inflammation, or any untoward pathological reactions in any of transplanted eyes or in the control noninjected contralateral eyes. No reduction in retinal function was recorded by electroretinogram up to 10 weeks following transplantation. This study demonstrates the feasibility and safety of transplanting hBMSCs in the EVSC compartment in a large eye model of rabbits.

Very long term success of pterygium surgery with conjunctival graft.

PURPOSE: To examine to the very long term prognosis (16 years in average) of pterygium surgery with superior or inferior conjunctival graft. METHODS: Clinical data for patients that underwent pterygium surgery with conjunctival graft by two surgeons at the Goldschleger Eye Institute, Sheba Medical Center, Israel, between the years 1997-2001 were retrieved from medical records. For all the included patients, data about their eye examinations was summarized and a questionnaire about their eye history was done. RESULTS: Twenty four patients were included in the study. Eleven with superior conjunctival flap (Group 1), and thirteen with inferior conjunctival graft (Group 2). In group 1, there was one surgery for recurrent pterygium, and two in group 2. In all cases the grafts were sutured and Mitomycin C was not used. No recurrence of pterygium was found in either group. No eye complications or side effects were found after sixteen years in average. CONCLUSIONS: After a very long follow-up of almost two decades there are no recurrences of pterygium or complications when using upper conjunctival flap or lower conjunctival graft during the surgery. It seems that lower graft is as good as upper flap for preventing recurrence of pterygium. Larger studies are required in order to confirm these results.

An Unusual Location of Fibrous Hamartoma of Infancy in the Eyelid.

AIM: To describe the clinical, imaging, and histopathological features of fibrous hamartoma of infancy (FHI) in the eyelid, a rare differential diagnosis for eyelid lesions. METHODS: We describe the case of a 7-month-old patient with a FHI in the eyelid that was diagnosed after surgical removal. The unique histopathological appearance of the triphasic histologic components provided the diagnosis. CONCLUSIONS: FHI is included in the long list of differential diagnosis for eyelid lesions in infancy. Therefore, it is important for the ophthalmologists to be familiar with this entity in order to avoid misdiagnosis of other fibromatosis and malignant tumor as well as unnecessary aggressive treatment.

A minimally invasive adjustable-depth blunt injector for delivery of pharmaceuticals into the posterior pole.

PURPOSE: To investigate the feasibility and safety of a novel minimally invasive adjustable-depth blunt injector for pharmaceuticals delivery into the posterior segment. METHODS: Indocyanine green (ICG), sodium fluorescein and iron oxide nanoparticles (IONPs) were injected using the new injector into the extravascular spaces of the choroid (EVSC) compartment of rabbits and cadaver pig eyes. Spectral domain optical coherence tomography (SD-OCT), fundus imaging and histology analysis were performed for assessment of injection safety and efficacy. RESULTS: Indocyanine green, fluorescein and IONPs were detected across the EVSC in rabbit eyes, covering over 80 per cent of the posterior eye surface. Injected IONPs were retained in the EVSC for at least 2 weeks following injection. No retinal detachment, choroidal haemorrhage or inflammation was detected in any of the injected eyes. In cadaver pig eyes, ICG was detected across the EVSC. CONCLUSIONS: This novel minimally invasive delivery system may be used to safely deliver large volumes of pharmaceuticals into a new treatment reservoir compartment - the EVSC which can serve as a depot, in close proximity to the retina, covering most of the surface of the back of the eye without insertion of surgical instruments under the central retina. This system is predicted to enhance the therapeutic effect of treatments for posterior eye disorders.

Low thyroid hormone levels improve survival in murine model for ocular melanoma.

Uveal melanoma is highly metastatic, prognosis is poor and there are no effective treatments to extend survival. Accumulating evidence suggests that thyroid hormones have a mitogenic effect via binding to αvß3 integrin. We aimed to examine the impact of thyroid status on survival in a murine B16F10 model for ocular melanoma, highly expressing the integrin. In two independent experiments oral propylthiouracil (PTU) was used to induce hypothyroidism (n=9), thyroxine to induce hyperthyroidism (n=11) and mice given plain water served as control (n=8). At day 21, the subretinal space was inoculated with 10(2) B16F10 cells. In non-inoculated mice (n=6 of each group) serum free T4 (FT4) levels were measured and additional non-inoculated mice (3 given PTU and 4 given thyroxine or water) served as internal control to demonstrate the impact of the dissolved substance. The PTU-inoculated mice showed clinical evidence of intraocular tumor growth significantly later than the thyroxine mice (P=0.003) and survival time was significantly longer (P<0.001). FT4 levels differed significantly between groups (P<0.001) and with no signs of illness in the internal control group. Our findings suggest that hyperthyroidism shortens survival, whereas relative hypothyroidism may have a protective role in metastatic ocular melanoma.