Expression of JAM-A in the human corneal endothelium and retinal pigment epithelium: localization and evidence for role in barrier function

PURPOSE: Junctional adhesion molecules (JAMs) are a family of adhesion proteins found in intercellular junctions. Evidence suggests that JAM-A is important for the regulation of tight junction assembly and epithelial barrier function. The authors recently reported that JAM-A is expressed in rabbit corneal endothelium and that antibody to JAM-A produces corneal swelling. In the present study, they investigate JAM-A expression in the human corneal endothelium and retinal pigment epithelium (RPE) and examine the effect of a function-blocking antibody to JAM-A on the permeability of cultured RPE cell monolayers. METHODS: Expression of JAM-A in human corneal endothelium, human RPE tissue, and cultured ARPE-19 monolayers was assessed by immunofluorescence confocal microscopy. Localization of JAM-A was compared with the tight junction-associated protein zonula occludens-1 (ZO-1). To investigate JAM-A function in ARPE-19 cells, ARPE-19 monolayers were subjected to a calcium switch protocol to disrupt cell junctions and treated with a function-blocking antibody to JAM-A or an isotype-matched control. Dextran flux assays were performed to assess the effect of JAM-A antibody on ARPE-19 monolayer permeability. RESULTS: Expression of JAM-A was observed in human corneal endothelium, and its distribution correlated with the tight junction-associated protein ZO-1. In addition, expression of JAM-A was observed in human RPE and in intercellular junctions of ARPE-19 monolayers. The localization pattern of JAM-A in the RPE and ARPE-19 monolayers was similar to that of ZO-1. ARPE-19 monolayers treated with antibody to JAM-A demonstrated a 33% increase in permeability to 10,000 MWt dextran compared with monolayers treated with control antibody. CONCLUSIONS: Results of this study provide new information about JAM-A expression in tight junctions of the human corneal endothelium and human RPE. The observation that antibodies to JAM-A increase ARPE-19 monolayer permeability is consistent with previous findings of JAM-A function in epithelial tight junctions and suggests JAM-A may have a role in the regulation of RPE barrier function.     

Gap junctions containing alpha8-connexin (MP70) in the adult mammalian lens epithelium suggests a re-evaluation of its role in the lens.

A missense mutation in one of the three lens connexins, alpha8-connexin, has been recently shown to be the genetic basis of the zonular pulverant lens cataract. This connexin had been considered to be expressed only in lens fibre cells. The present studies show that alpha8-connexin is also expressed in the lens epithelial cell layer.For this study, the distribution of gap junctions in the adult bovine lens has been investigated by confocal immunofluorescence microscopy using antibodies against alpha8-connexin (MP70) and alpha1-connexin (Cx43). In addition to the anticipated localisation of alpha8-connexin to the broad faces of lens fibre cells as reported in other species, alpha8-connexin was also found colocalized with alpha1-connexin at plaques in the lateral epithelial-epithelial plasma membranes of the bovine lens. These data suggest that mixed alpha8-connexin/alpha1-connexin plaques are between epithelial cells at their apico-lateral plasma membranes, rather than between epithelial and fibre cells. Indeed, freeze fracture analyses of the epithelial-fibre cell interface failed to reveal gap junctions connecting the epithelium and the underlying fibre cells. Importantly, microdissection and subsequent immunoblotting of lens epithelium samples confirmed the immunolocalisation results. The data suggest mature mammalian lens epithelial cells could form either heteromeric, heterotypic and/or mixed homomeric-homotypic gap junctional complexes with unique physiological properties, an important point when considering the role of epithelial cell connexins in cataractogenesis.     

Effect of xyloglucan (tamarind seed polysaccharide) on conjunctival cell adhesion to laminin and on corneal epithelium wound healing

PURPOSE: To explore the role of a natural polysaccharide extracted from tamarind seed (xyloglucan, or tamarind seed polysaccharide, TSP) on the integrin-substrate recognition system and on repair of corneal wounds. METHODS: a) Cultured human conjunctival cells were labeled by addition of a tritiated amino acid mixture. Their adhesion to laminin-coated culture wells in the absence or presence of TSP was checked by radioactivity count. b) The corneal epithelium of albino rabbits was damaged by applying a paper disc soaked with n-heptanol. The eyes were then treated with TSP, with a hyaluronate reference formulation and with normal saline solution (controls). The diameter of corneal wounds was measured daily, after fluorescein staining. RESULTS: Compared to hyaluronate, TSP slightly but significantly increased the wound healing rate. TSP 1.0% exerted a positive influence on cell adhesion to laminin, up to a certain laminin concentration. CONCLUSIONS: The ability of the polysaccharide to promote corneal wound healing might depend on its influence on the integrin recognition system.     

Lipid metabolism in retinal pigment epithelium (RPE): a possible role of LDL receptors

PURPOSE: The retinal pigment epithelium (RPE) regulates the lipid metabolism of the photoreceptors by catalysis of membrane outer segments and via choriocapillary perfusion is also exposed to the regulation of blood lipid levels. Since the uptake the metabolism of cholesterol are mediated by specific low-density lipoprotein (LDL) receptors, expression and regulation of this receptor-type were studied in RPE cultures. METHODS: In vitro experiments were carried out in transformed (SV40) human RPE cells. Human fibroblasts were used as a comparative cell line with known receptor expression. LDL was coupled to a fluorescent marker (Dil); receptor binding was quantified by flow cytometry. Expression and saturation characteristics were determined. LDL metabolism was examined by variation of the temperature (4 and 37 degrees C). LDL and Dil-LDL showed competition at the receptor. RESULTS: RPE cells demonstrated a higher uptake of Dil-LDL than fibroblasts. Expression could be further stimulated by culture conditions. Uptake kinetics were saturable with complete saturation at 50 micrograms/ml Dil-LDL. LDL uptake was shown to be temperature-dependent, indicating an energy-dependent pathway. CONCLUSIONS: RPE cells exhibit significant expression of receptors for native LDL, possibly mediating the lipid metabolism of the RPE-photoreceptor complex, as well as the uptake of blood lipids. Lack of regulation of the receptor for LDL may lead to intracellular accumulation of lipids, which may play a role in the pathogenesis of AMD.     

Combination of adult inclusion conjunctivitis and mucosa-associated lymphoid tissue (MALT) lymphoma in a young adult

PURPOSE: To report a patient who was diagnosed with combined adult inclusion conjunctivitis (AIC) and mucosa-associated lymphoid tissue (MALT) lymphoma. METHODS: This is a case report. RESULTS: An 18-year-old male patient presented with chronic conjunctivitis and giant follicles. Evaluation by chlamydial antigen assay was positive. Conjunctival biopsy for the immunohistochemical stain and polymerase chain reaction of the left eye showed MALT lymphoma. CONCLUSIONS: MALT lymphoma can masquerade as other ocular surface diseases. Chlamydial infection causes chronic inflammation of the conjunctiva. Both of these diseases should be considered as a differential diagnosis of refractory follicular conjunctivitis. It is worthy of further study to determine whether chronic inflammation resulting from chlamydial infection increases the risk of MALT lymphoma or it is coincidental.     

Cell adhesion molecules in subretinal fluid: Soluble forms of VCAM-1 (vascular cell adhesion molecule-1) and L-selectin

Purpose: In this study we investigated the presence of soluble VCAM-1 and soluble L-selectin-1 in subretinal fluids (SRF) of patients suffering from rhegmatogenous retinal detachment . Method: Subretinal fluids were collected from drainage sclerotomies during surgery from 27 patients with rhegmatogenous retinal detachment complicated by proliferative vitreoretinopathy (PVR) or uncomplicated retinal detachment. Levels of sVCAM-1 and sL-selectin-1 were quantified with ELISA. Results: The mean ± SEM values of sVCAM-1 and sL-selectin-1 were 222.2 ± 81 ng/ml and 171.7 ± 42.1 ng/ml, respectively. The concentrations of sVCAM-1 in patients with Grade C PVR (498.2 ± 170 3 ng/ml) were significantly different from those with Grade B PVR (45.6 ± 16.5 ng/ml) and uncomplicated retinal detachments (19.4 ± 12.3 ng/ml). SVCAM-1 concentration in detachments which had been present for more than 8 weeks was 738.8 ± 431 ng/ml, significantly higher than the levels in detachments of shorter duration (132.4 ± 47.7 ng/ml). sL-selectin-1 level in Grade C PVR (291.6 ± 92.8 ng/ml) was higher than in uncomplicated retinal detachments (72.8 ±13.5 ng/ml). Significantly elevated levels of sL-selectin-l were observed in detachments lasting more than 8 weeks (605 ± 151.1 ng/ml) compared to those of shorter duration (96.3 ± 13.1 ng/ml). Conclusion: The present study supports growing evidence that these cell adhesion molecules are involved in the inflammatory process during the development and progression of PVR. This revised version was published online in July 2006 with corrections to the Cover Date.     

Hyperosmolarity and electroretinogram (ERG) potentials in isolated rat retinas: possible implications in diabetic models

In this report, the effects of increases in the osmolarity of media superfusing isolated rat retinas on the a-wave, b-wave and oscillatory potentials of the electroretinogram (ERG) were examined. Osmolarity of the media was raised from 310 milliosmoles (control) to 340 and 370 milliosmoles by addition of NaCl or sucrose. Increases in osmolarity led to rapid decreases in the amplitudes of the b-wave and oscillatory potentials with little change in the amplitude of the a-wave. Excellent recovery of the ERG potentials was observed when control conditions were restored. The implications of these effects of an hyperosomotic load on ERG potentials in vitro are discussed with regard to a possible role of this load in models of experimental diabetes.     

Raising the suborbicularis oculi fat (SOOF): its role in chronic facial palsy

AIMS: To determine the adjuvant role of unilateral suborbicularis oculi fat (SOOF) lift in the periorbital rehabilitation of patients with chronic facial palsy. METHODS: In a non-comparative prospective case series nine adult patients (seven male, two female) aged 34-90 years (mean 60.5) with chronic unrecovered facial palsy (over 1 year), who had not had any previous rehabilitative periorbital surgery, were studied. Lateral tarsal strip and adjuvant transconjunctival approach subperiosteal SOOF lift under local or general anaesthesia were performed; medial canthoplasty was performed where indicated. There was clinical observation of the long term (over 1 year) effect on the ptotic palpebral-malar sulcus and lower eyelid retraction. RESULTS: The patients were followed up for 12-24 months (mean 16). Seven patients (77%) had sustained clinical reduction of palpebral-malar sulcus ptosis. All patients had sustained reduction of lagophthalmos. Early postoperative complications included conjunctival cheimosis in 77%. Three patients with persistent keratitis required further surgical procedures on their upper eyelid to reduce the palpebral aperture. There were no cases of infraorbital nerve anaesthesia or recurrent lower eyelid retraction. CONCLUSIONS: The SOOF lift has an adjuvant role in chronic facial palsy with lower eyelid retraction and ptotic-palpebral malar sulcus. It supports the lower eyelid elevation and tightening achieved with the lateral tarsal strip. The best results were obtained in congenital facial palsy.     

mRNA for interphotoreceptor retinoid-binding protein (IRBP): distribution and size diversity in vertebrate species

Northern blots of total retinal RNA from a number of different vertebrate species were probed with a cDNA fragment corresponding to translated portions of bovine IRBP mRNA. A hybridizing band was detected in normal human retina (4.6 kb), Y-79 human retinoblastoma cells (4.4 kb), and in retinas of monkey (4.6 kb), guinea-pig (4.9 kb), mouse (6.1 kb), rat (two bands at 5.4 kb and 6.6 kb), rabbit (6.3 kb), cow (6.5 kb), and hamster (7.6 kb). Thus, the IRBP gene is expressed in the retinas of a wide variety of mammalian species. The mRNAs could be readily detected in about 20 micrograms of total RNA, suggesting that, in these species, IRBP message is relatively abundant. In contrast, only very weak hybridization was detected on northern blots of the three bird species examined (chicken, duck, quail). IRBP mRNA is thus relatively well expressed in mammals, but not in birds.     

Antiviral agents and corneal epithelium cicatrisation: study in rabbits (author's transl)

Five clinically-used antiviral drugs (3% adenine arabinoside ointment; 3% acycloguanosine ointment; 0.24% idoxuridine ointment; 1% trifluorothymidine drops) were compared with a control (petrolatum base) to determine their toxic effects on rabbit corneal epithelium after injury by iodine vapors: --Only trifluorothymidine significantly retarded healing of epithelial erosions. --Histopathologic examination after seven-day treatment showed that all five drugs, except vidarabine and to a lesser degree acycloguanosine, caused toxic changes in the regenerating epithelium.     

Expression of intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) on proliferating vascular endothelial cells in diabetic epiretinal membranes.

The present study demonstrated the expression of intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1), the proliferating status of the neovascular endothelial cells, and the activation of vascular endothelial cells bearing the two cell adhesion molecules in diabetic epiretinal membranes by using double immunofluorescence and APAAP techniques. The results showed that ICAM-1 was detected in 36 out of 40 (90%) proliferative diabetic retinopathy epiretinal membranes, VCAM-1 was found in 32 out of 40 cases (80%); in 21 out of 26 (81%) vascularised membranes the endothelial cells of the new vessels in the membranes were still in a proliferative stage (positive for proliferating endothelial cell marker EN 7/44) and, further, in 20 out of 26 cases (77%) ICAM-1 was detected on the proliferating endothelial cells and VCAM-1 was found in 21 cases (81%). The expression of cell adhesion molecules, especially ICAM-1 and VCAM-1 in diabetic epiretinal membranes suggests that cell to cell interactions may play a significant role in the development of PDR membranes. In particular, the expression of ICAM-1 and VCAM-1 on proliferating endothelial cells indicates the activation of these cells, which is the first critical step for lymphocyte/endothelial cell interactions and the initiation of immune responses. The significance of proliferating status of the neovascularisation in the membranes may be related to the clinical course and prognosis of the disease.     

Plasminogen activator inhibitor-1 (PAI-1) stimulates human corneal epithelial cell adhesion and migration in vitro

In addition to its role as an inhibitor of urokinase plasminogen activator (uPA), plasminogen activator inhibitor-1 (PAI-1) is hypothesized to regulate epithelial cell adhesion and migration. We have previously reported that PAI-1 may be an important regulatory factor of the uPA system in cornea. The purpose of this study was to extend those observations by determining the effect of exogenous PAI-1 on the migration and adhesion of human corneal epithelial cells (HCEC) in vitro. The expression of PAI-1 in non-transformed early passage HCEC was confirmed by immunofluorescence microscopy and Western blot analysis. Colorimetric assays coupled with function-inhibiting antibody studies using the matrix assembled in situ by cultured cells demonstrate that immobilized PAI-1 serves as an efficient substrate for HCEC adhesion. HCEC attachment to PAI-1 is comparable to that of laminin-10, a known strong adhesion protein for epithelial cells. In addition to serving as an adhesion substrate, PAI-1 also functions as a chemotactic agent for corneal epithelium. Additionally it promotes the random migration of HCEC, from an initial cell cluster, along a culture substrate. Our results in corneal epithelium are consistent with reports from other investigators showing that PAI-1 facilitates both epithelial adhesion and migration. From our studies we conclude that PAI-1 may play a dual role in corneal wound healing. Initially PAI-1 may function to stimulate migration and facilitate the reepithelialization of the wound bed. Post-reepithelization, PAI-1 may ensure corneal epithelial cell adhesion to matrix to promote successful wound healing.     

Inflammatory cell adhesion and surface defects on heparin-surface-modified poly(methyl methacrylate) intraocular lenses in diabetic patients

PURPOSE: To evaluate the incidence of surface scratches on heparin-surface-modified (HSM) poly(methyl methacrylate) (PMMA) intraocular lenses (IOLs) and the possible influence of these alterations on the biocompatibility of HSM PMMA. SETTING: University Eye Clinic of Trieste, Trieste, Italy. METHODS: Twenty-six diabetic patients had phacoemulsification and implantation of an HSM PMMA IOL (809C, Pharmacia & Upjohn). Patients with proliferative diabetic retinopathy or iridopathy were excluded from the study. On postoperative days 7, 30, 90, and 180, specular microscopy was performed to study and photograph the anterior IOL surface. The presence of scratches on the anterior IOL surface was assessed and the inflammatory cell reaction noted and graded using a semiquantitative scale. Finally, the location of the inflammatory cells in relation to the surface scratches was established. RESULTS: Scratches and other surface defects were found in 88.4% of cases. All patients had small cells on the IOL surface 7 days after surgery. At 30 days, small cells were observed in 88.4% of cases. The inflammatory cells were mainly located inside the scratches rather than throughout the IOL surface. CONCLUSIONS: This in vivo cytology study provides further evidence of the effectiveness of heparin surface modification in improving the biocompatibility of PMMA. In diabetic patients, inflammatory cells adhered to the exposed PMMA surface more than to the HSM surface, suggesting that the use of HSM PMMA in patients with conditions predisposing them to increased postoperative blood-aqueous barrier breakdown is beneficial.     

Activation and role of MAP kinase-dependent pathways in retinal pigment epithelium cells: JNK1, P38 kinase,and cell death

PURPOSE: Retinal pigment epithelial (RPE) cell death is an important step in the pathogenesis of ocular diseases. JNK1 and P38 kinase, two stress-activated kinases, play key roles relaying stress signals leading to cell death through cyclin D1 and c-Myc. Recently, stress-activated kinases have been shown to regulate cell proliferation. In the current study, the involvement of the JNK1 and P38 kinase signaling pathways in RPE cell proliferation and death was investigated. METHODS: RPE cell proliferation was stimulated with 10% fetal calf serum (FCS). Activation of the JNK1 and P38 kinase cascades and their potential targets was detected by Western blot analysis. Pharmacologic inhibitors and activators, and antisense oligodeoxynucleotides (ODN) directed against the stress kinases were used to analyze the signaling involved in RPE cell death. RESULTS: P38 and JNK1 and their respective upstream activating kinases, MKK3/6 and -4, were all transiently activated in FCS-stimulated RPE cell cultures. Ras controlled only the activation of JNK1, whereas Rho transmitted the activation of both JNK1 and P38, suggesting parallel signaling pathways and cross talk between the two kinases. Pharmacologic inhibition of JNK1 did not affect cell proliferation in FCS-stimulated cells. Inactivation of P38 kinase and antisense ODN-induced downregulation of P38 kinase also had no affect on cell proliferation. Long-term, high-level activation of JNK1 and P38 kinase occurred during serum depletion-induced RPE cell death. Overactivation of JNK1 and P38 kinase was also observed during pharmacologically induced cell death, suggesting that this process is common to RPE cell-death-signaling pathways induced by various stress stimuli. Cell death mediated by the overactivation of JNK1 and P38 kinase was cyclin D1- and c-Myc-independent. CONCLUSIONS: The inhibition of JNK1 or P38 kinase had no effect on FCS-stimulated proliferation of RPE cells, whereas the overactivation of these two enzymes was involved in RPE cell death in FCS-depleted cultures. Parallel upstream signaling pathways and cross talk between the two kinases suggest that the regulation of signaling in RPE cell death is complex.     

Optometry and its expanded role in health care delivery systems (1600-1987)

Today's optometry students and practitioners alike are often unaware of the richness of optometric history. The use of ophthalmic pharmaceutical agents and the designation of primary care practitioners are often taken for granted. This article utilizes some of the early research conducted by Harrington, Hofsteter, Prentice, Hirsch, Wick, Fitch, and Gregg, and describes the advent of drug legislation and the expanded role of optometry within the health care system.     

Amacrine and horizontal cell dysfunction in adult ceroid-lipofuscinosis (Kufs disease) and anatomical correlates in the ovine model

Psychophysical examination of the visual performance of a patient with Kufs disease suggests abnormal functioning of the amacrine and horizontal cell systems of lateral inhibition. The sheep model offers insight into early patho-physiology of these cell types.