Presence of a soluble form of acetylcholinesterase in human ocular fluids.

Samples of ocular fluid obtained from normal persons at necropsy and during eye surgery have been assayed for the presence of acetylcholinesterase. Measurable levels could be detected in all samples examined, but levels of acetylcholinesterase in vitreous humour were consistently higher than those in aqueous humour, indicating a possible retinal origin. Polyacrylamide gel electrophoresis revealed that the enzyme of ocular fluid had the same mobility as that of acetylcholinesterase from cerebrospinal fluid. It is probable that acetylcholinesterase is secreted from neuronal structures in the retina into the ocular fluid in an analogous manner to the secretion of acetylcholinesterase from brain neurones into cerebrospinal fluid.     

Immunolocalization of transcription factor AP1 in human ocular surface epithelia.

PURPOSE: The present study examined whether normal human ocular surfcae epithelia express AP1 components. Changes in expression patterns of these components in a case of ocular surface epithelial dysplasia was also evaluated before and after topical mitomycin C treatment. METHODS: Specimens of normal corneas (n = 2) and conjunctiva (n = 4) were obtained from 4 patients during cataract surgery or post mortem, while specimens of dysplastic epithelial tissue from the limbus were obtained from one patient. Specimens were immunohistochemically studied using antibodies against components of AP1. RESULTS: The normal corneal epithelium showed no staining with antibodies against c-Fos, Fra-2, FosB, c-Jun or JunB, whereas the limbal and bulbar conjunctival epithelia were positive for c-Fos, Fra-2, and c-Jun. Anti-FosB and -JunB antibodies reacted weakly with the conjunctival epithelium. JunD was absent in normal corneal and conjunctival epithelia. The dsyplastic epithelium showed positive labelling for c-Fos, Fra-2, c-Jun, and JunD throughout its thickness. Fra-1 was present in all specimens of epithelia examined. The dysplastic epithelium treated with mitomycin C was not labeled by anti-c-Fos or -Fra-2 antibody. CONCLUSION: Individual AP1 components show specific expression patterns in normal ocular surface epithelia and a case of dysplastic epithelium before and after topical MMC treatment, implying that these factors may play important roles in modulating epithelial cell function, e.g., proliferation and differentiation.     

Rapid detection of herpes simplex virus (HSV) antigen in human ocular infections

The new HERPCHEK (Dupont, No. Billerica, MA) enzyme immunosorbent assay (EIA) was used in a double-blind clinical study for rapid and specific detection of ocular herpes simplex virus (HSV) infection. This 4-hour assay can be used to demonstrate conclusively the presence of HSV antigen without culture and thereby rapidly differentiate between HSV and other clinically similar ocular infectious diseases. Ocular samples were collected from 180 individuals including 30 patients with acute HSV, 90 with latent HSV (ie, currently asymptomatic but with a positive history), 11 with acute or latent varicella zoster virus, 30 with nonherpetic infections (due to adenovirus, Acanthamoeba or bacteria), and 19 normal controls. A clinical diagnosis was made by one of us (DPL) and duplicate tear-film samples obtained by swabbing the conjunctival cul-de-sac and cornea. Coded samples were tested by routine viral culture on Vero cell monolayers and also were run independently in the HERPCHEK test. During active HSV infection, the HERPCHEK correlated 100% with clinical diagnosis, and virus culture correlated 90% with clinical diagnosis. In all latent HSV ocular infections, other nonherpetic ocular infections and normal samples, both the HERPCHEK and culture assays were negative.     

Immunoreactive parathyroid hormone is present in subretinal fluids of the human eye

(44-68) human Parathyroid hormone (hPTH) was studied in subretinal fluids (SRF) of 20 human eyes with rhegmatogenous retinal detachments. A (44-68) hPTH carboxyl mid-regional radioimmunoassay was used. Depending on the extent and the duration of the retinal detachments, (44-68) hPTH ranged between normal serum levels (ca. 100-300 pg/ml) up to excessive amounts of 4000 pg/ml with longstanding disease and proliferative vitreoretinopathy. Aqueous humors (AH) of one patient with bilateral iridocyclitis and complicated cataract ranged at 90-132 pg/ml (44-68) hPTH, whereas a control population undergoing surgery for senile cataracts had no detectable (44-68) hPTH in AH. The AH of one diseased eye of heterochromia complicata Fuchs showed excessive amounts of more than 4000 pg/ml (44-68) hPTH.     

Identification and localization of major soluble vitreous proteins in human ocular tissue

PURPOSE: To identify the major soluble proteins from human vitreous, and to establish a baseline for comparison of vitreous samples from eyes with various diseases. DESIGN: Laboratory investigation. METHODS: Normal vitreous was obtained from eight human donor eyes and from eight eyes of patients undergoing diabetic vitrectomy. Vitreous specimens were subjected to SDS-PAGE and MALDI-TOF-MS analysis. Six specific antibodies were used to identify proteins using Western blot. Four proteins were localized within ocular tissue in a normal donor eyebank eye. RESULTS: We found eight distinct bands on SDS-PAGE in normal vitreous and two additional bands (hemoglobin) in eyes with diabetic retinopathy. Proteins were identified using MALDI-TOF-MS and confirmed by Western blot. We established a quantitative analysis of relative protein concentrations of undiluted vitreous. Immunohistochemistry localized selected proteins within the posterior segment layers. CONCLUSIONS: We present a normal human vitreous protein profile using current technologies and provide a baseline for comparison to ocular disease states. Tissue distribution of vitreous proteins may help to elucidate more specific protein function.     

Immunological study of proteins and mucosubstance in saline soluble human ocular mucus

Proteins and mucosubstance of the saline extract of human ocular mucus were studied by immunological analysis. A minor study was made with human tears for comparison. Immunoelectrophoresis of proteins from these two sources consistently revealed similar characteristic gel patterns. Proteins were found as the major constituents of both samples. However, more mucosubstance was present in the saline extract of human ocular mucus than in tears. Seventeen proteins were identified in the mucus extract. Albumin, IgA, and lactoferrin appeared to be the three major proteins, while lysozyme, lactoferrin, tear prealbumin, and ocular mucoisolate were tear and ocular mucus specific. Although saline soluble mucoisolate is complex in structure, it seemed to resist electrical dissociation, producing only one major precipitation line along with a line of IgA during immunoelectrophoresis. The ocular mucoisolate accounted for about 12% of the saline extractable proteins of human ocular mucus.     

Gradient detection and contrast transfer by the human eye

Threshold measurements were performed with stimulus patterns involving gradual changes in luminance. The results of these measurements indicate that threshold data obtained using linear luminance gradients permit quantitative prediction of the dependence of the threshold modulation on the spatial frequency as well as on the number of cycles of sinusoidal gratings. It appears that for detection purposes, a sinusoidal grating can be considered as one linear gradient with the same width. This makes it possible to predict the spatial modulation transfer function of the eye from measurements with spatial luminance gradients.     

CD44 and RHAMM hyaluronan receptors in human ocular surface inflammation

Background

CD44 and RHAMM hyaluronan (HA) receptors have been studied in several systemic diseases such as osteoarthritis and cancer. However, not too much is known about their role in ocular surface disorders. The purpose of this research was to determine if CD44 and RHAMM are implicated in human ocular surface inflammation.

Methods

Upper tarsal conjunctival epithelial samples from patients with active ocular surface inflammation (n?=?17) and healthy controls (n?=?14) were recovered by brush cytology. Patients were evaluated by an ophthalmologist and classified in different groups according to the etiology (immune atopic diseases or immune non-atopic diseases) and inflammation intensity (mild/moderate or severe). CD44, RHAMM, and p53 mRNAs were measured using real-time PCR.

Results

CD44, RHAMM, and p53 mRNAs were detected in all samples. In immune atopic diseases, higher levels of CD44 and RHAMM mRNAs were present, reaching a 300 % increase for RHAMM in severe inflammation (p?<?0.001). In contrast, in immune non-atopic diseases, the HA receptors were downregulated. CD44 tended to decrease up to 30 % in severe patients (p?=?0.06), and RHAMM decreased 40 % in severe inflammation (p?=?0.021).

Conclusions

RHAMM may be implicated in severe ocular surface inflammation affecting the upper tarsal conjunctiva.     

Role of melatonin in the eye and ocular dysfunctions

Melatonin is a ubiquitous molecule and widely distributed in nature, with functional activity occurring in unicellular organisms, plants, fungi, and animals. Several studies have indicated that melatonin synthesis occurs in the retina of most vertebrates, including mammals. The retinal biosynthesis of melatonin and the mechanisms involved in the regulation of this process have been extensively studied. Circadian clocks located in the photoreceptors and retinal neurons regulate melatonin synthesis in the eye. Photoreceptors, dopaminergic amacrine neurons, and horizontal cells of the retina, corneal epithelium, stroma endothelium, and the sclera all have melatonin receptors, indicating a widespread ocular function for melatonin. In addition, melatonin is an effective antioxidant which scavenges free radicals and up-regulates several antioxidant enzymes. It also has a strong antiapoptotic signaling function, an effect that it exerts even during ischemia. Melatonin cytoprotective properties may have practical implications in the treatment of ocular diseases, like glaucoma and age-related macular degeneration.     

Quantitative PCR for the detection of genomic DNA of Epstein-Barr virus in ocular fluids of patients with uveitis

Purpose  To measure the genomic DNA from Epstein-Barr virus (EBV) in ocular fluids and to analyze the clinical relevance of EBV in uveitis. Methods  Intraocular fluids (30 aqueous humor and 30 vitreous fluid samples) were taken from 55 patients with uveitis after informed consent was obtained. Samples were assayed for EBV DNA using qualitative multiplex polymerase chain reaction (PCR) and quantitative real-time PCR. Antibodies to EBV were examined using a complement fixation test. Results  EBV DNA was detected in 17 of 60 samples (28%) and 16 of 55 patients (29%) using multiplex PCR. However, only three of the 17 samples showed significantly high copy numbers of EBV DNA with real-time PCR. EBV DNA was not detected in the serum of all patients. EBV-specific antibodies were positive in the serum of all patients, but not in the vitreous fluid. Vitreous anti-EBV antibodies were positive only in patients displaying genomic DNA of EBV in the vitreous samples. Conclusions  EBV DNA was detected by qualitative PCR in ocular fluids of many uveitis patients, but only a small proportion of patients showed high viral loads on quantitative real-time PCR, indicating that replication of the virus takes place only in a few patients.