Aqueous humor oxidative stress proteomic levels in primary open angle glaucoma.

The purpose of this work was to investigate the expression of glutamine synthase (GS), nitric oxide synthase (NOS) superoxide dismutase (SOD) and glutathione transferase (GST) in the aqueous humor of patients with primary open angle glaucoma and controls. Aqueous humor proteome was analyzed by antibody microarray. The expression of tested proteins was detected by protein Cy3/Cy5 labeling, column purification and hybridization on antibody-spotted glass microarray. Fluorescent signals were detected by fluorescence laser scanning. Aqueous humor levels of SOD as well as of GST were significantly lower (2.0- and 2.2-fold, p < 0.01) among patients than controls; both NOS and GS expression were significantly higher (2.2- and 2.6 fold, p < 0.01) among patients than controls. Our data showed substantial differences of GS, NOS2, SOD and GST aqueous humor levels between glaucomatous patients and controls as measured by antibody microarray technology. The overproduction of NO through inducible NOS can form toxic products and change the metabolic conditions of the TM. The GS over-expression might be related to neuronal injury or to the potential role of glutamate as a modulator in the ciliary body signaling. The reduced expression of the antioxidant enzymes SOD and GST could aggravate the unbalance between both oxygen- and nitrogen-derived free radicals production and detoxification. Based on our results, GS, NOS2, SOD and GST as measured by antibody microarray technology may be useful oxidative markers in aqueous humor of glaucomatous patients.

Increased resistance to oxidative DNA damage of trabecular meshwork cells by E. coli FPG gene transfection.

Oxidative damage plays a pathogenic role in various chronic degenerative diseases. Oxidative damage targeting trabecular meshwork (TM) cells as a consequence of mitochondrial damage is a pathogenic mechanism for glaucoma, the most common cause of irreversible blindness worldwide. Consequences of oxidative damage are attenuated by endocellular activities involved in scavenging reactive oxidative species and DNA repair. Selected bacterial genes are highly efficient at protecting cells from oxidative DNA damage. This situation occurs for Escherichia coli formamidopyrimidine DNA glycosylase (FPG), a major DNA glycosylase that repairs oxidatively damaged DNA. Accordingly, this study was aimed at transfecting human TM cells (HTMC) with Fpg in order to increase their resistance to oxidative damage. This study demonstrates that it is feasible to increase resistance of HTMC to endogenous oxidative damage by gene transfection. These findings bear relevance for primary and secondary prevention of degenerative glaucomas and other degenerative diseases where oxidative damage plays a pathogenic role.

Proteome alterations in primary open angle glaucoma aqueous humor.

As the only nourishment and scavenging source for most of the anterior and posterior chamber tissues in the eye, the aqueous humor represents one of the target for glaucoma. The aim of this study is to investigate the yet unexplored relationship between aqueous humor protein content and open-angle glaucoma (POAG) pathogenesis. Aqueous humor was collected from 10 POAG patients (cases) and 14 senile cataract patients (controls), matched for age and gender, undergoing surgery for trabeculectomy and cataract, respectively. Protein samples were cyanine-labeled and hybridized with antibody microarrays. Microarray signals were revealed by laser scanner, quantified, and compared by statistical analyses. Total protein amounts were not significantly different in patients versus controls. Conversely, a proteome cluster significantly modified in patients as compared to controls was identified as highly predictive for disease status. Selected proteins underwent dramatic variation, which was correlated to pathogenetic events characterizing POAG, including oxidative damage, mitochondrial damage, neural degeneration, and apoptosis. The results obtained indicate that proteomic analysis of aqueous humor is a new tool for POAG diagnosis in the case of otherwise uncertain disease recognition. Furthermore, this study allows a better understanding of mechanisms involved in the pathogenesis of POAG, the main cause of irreversible blindness worldwide.

Glaucoma and Helicobacter pylori infection: correlations and controversies.

A possible association between Helicobacter pylori infection and eye diseases, including Sjögren syndrome, blepharitis, central serous chorioretinopathy and uveitis, has been proposed. Glaucoma is the second leading cause of blindness in the world, after cataracts, and the leading cause of irreversible blindness, but many aspects of its pathogenesis remain unknown. H pylori infection may influence the pathophysiology of glaucoma by releasing various proinflammatory and vasoactive substances, as well as by influencing the apoptotic process, parameters that may also exert their own effects in the induction and/or progression of glaucomatous neuropathy. It is difficult to understand how H pylori infection can be linked to such varied pathologies. Systemic H pylori-induced oxidative damage may be the mechanism which links oxidative stress, H pylori infection and the damage to the trabecular meshwork and optical nerve head that results in glaucoma.

Gene-environment interactions in ocular diseases.

Degenerative ocular diseases are widespread in the population and represent a major cause of reversible and irreversible blindness. Scientific evidences have been accumulating supporting the role of genotoxic damage and gene environment interactions in the pathogenesis of these diseases mainly including glaucoma, age-related macular degeneration, and cataract. Glaucoma, in its degenerative form, is characterized by the degeneration of the trabecular meshwork, the tissue of the anterior chamber of the eye devoted to aqueous-humour outflow. Such a degenerative process results in intra-ocular pressure increase and progressive damage of optic nerve head. Oxidative stress and DNA damage play an important role in inducing the degeneration of these well differentiated target tissues in which DNA damage results in a progressive cell loss. Macular degeneration is a common age-related disease affecting the central regions of the retina inducing progressive accumulation of oxidized lipoproteins and neovascularization. Environmental genotoxic risk factors include diet, light, and cigarette smoke paralleled by individual susceptibility as determined by adverse genetic assets. Cataract is a progressive opacity of the crystalline lens resulting from molecular damages induced by various risk factors including UV-containing light. This disease has been related to a failure in antioxidant defences. Experimental study provides evidence that cataract patients possess higher basal level of DNA damage, as evaluated by Comet test, in lymphocytes than controls. This finding is paralleled by the higher susceptibility to oxidative stress observed in the same patients. These novel experimental data further support the role of DNA damage as a main factor contributing to cataract onset. In conclusion, the examined degenerative ocular diseases recognise environmental risk factors often displaying genotoxic attitudes. Whenever these factors target individuals who are susceptible due their genetic assets the results is the onset of a specific eye disease depending on the affected ocular tissue.

Oxidative stress and glaucoma: injury in the anterior segment of the eye.

The perturbation of the pro-oxidant/antioxidant balance can lead to increased oxidative damage, especially when the first line of antioxidant defense weakens with age. Chronic changes in the composition of factors present in aqueous or vitreous humor may induce alterations both in trabecular cells and in cells of the optic nerve head. Free radicals and reactive oxygen species are able to affect the cellularity of the human trabecular meshwork (HTM). These findings suggest that intraocular pressure increase, which characterizes most glaucomas, is related to oxidative and degenerative processes affecting the HTM and, more specifically, its endothelial cells. This supports the theory that glaucomatous damage is the pathophysiological consequence of oxidative stress. Glaucomatous subjects might have a genetic predisposition, rendering them more susceptible to reactive oxygen species-induced damage. It is likely that specific genetic factors contribute to both the elevation of IOP and susceptibility of the optic nerve/retinal ganglion cells (RGCs) to degeneration. Thus, oxidative stress plays a fundamental role during the arising of glaucoma-associated lesions, first in the HTM and then, when the balance between nitric oxide and endothelins is broken, in neuronal cell. Vascular damage and hypoxia, often associated with glaucoma, lead to apoptosis of RGCs and may also contribute to the induction of oxidative damage to the HTM. On the whole, these findings support the hypothesis that oxidative damage is an important step in the pathogenesis of primary open-angle glaucoma and might be a relevant target for both prevention and therapy.

Antioxidant activity of timolol on endothelial cells and its relevance for glaucoma course.

PURPOSE: A growing evidence in the scientific literature suggests that oxidative damage plays a pathogenic role in primary open-angle glaucoma. Therefore, it is of interest to test whether drugs effective against glaucoma display antioxidant activity. We test the hypothesis that the classic beta-blocker therapy for glaucoma with timolol involves the activation of antioxidant protective mechanisms towards endothelial cells. METHODS: Oxidative stress was induced in cultured human endothelial cells by iron/ascorbate with or without timolol pretreatment. Analysed parameters included cell viability (neutral red uptake and tetrazolium salt tests), lipid peroxidation (thiobarbituric reactive substances), and occurrence of molecular oxidative damage to DNA (8-hydroxy-2'-deoxyguanosine). RESULTS: Oxidative stress decreased 1.8-fold cell viability, increased 3.0-fold lipid peroxidation and 64-fold oxidative damage to DNA. In the presence of timolol, oxidative stress did not modify cell viability, whereas lipid peroxidation was increased 1.3-fold, and DNA oxidative damage 3.6-fold only. CONCLUSIONS: The obtained results indicate that timolol exerts a direct antioxidant activity protecting human endothelial cells from oxidative stress. These cells employ mechanisms similar to those observed in the vascular endothelium. It is hypothesized that this antioxidant activity is involved in the therapeutic effect of this drug against glaucoma.

A human placental polydeoxyribonucleotide (PDRN) may promote the growth of human corneal fibroblasts and iris pigment epithelial cells in primary culture.

The optimal concentration of a human placental polydeoxyribonucleotide (PDRN) preparation (100 microg/ml) enhances the growth of human corneal fibroblasts in primary culture depending upon the donor age. In particular, this effect is very consistently reproducible with donors over 60 years of age (p = 0.0028), suggesting a selective benefit of PDRN in the tissue culture of senescent cells. Moreover, this drug may promote the development of human iris pigment epithelium (IPE) cells with much lower concentrations of fetal bovine serum than those suitable for the culture of IPE. Lastly, the use of a 'gauze disk' on the pieces of the corneal explants improves the efficiency of growth of the control fibroblast primary cultures.

Evaluation of the analgesic effect of 0.1% indomethacin solution on corneal abrasions.

PURPOSE: The authors wished to verify the analgesic action of 0.1% indomethacin in a water-based solution on patients affected by traumatic corneal abrasions. METHODS: 347 patients affected by traumatic corneal abrasions, having been randomly divided into 2 groups on the basis of the administration of indomethacin, were evaluated at 30 min, 12 h and 24 h after the initial treatment of the abrasion. The level of pain experienced was evaluated on a verbal pain scale and the healing time was evaluated relative to the dimension of the abrasion. RESULTS: The pain level was initially overwhelming for both groups: p = 0.737; at successive check-ups it was possible to verify a reduction of the symptomatology, with a more pronounced decrease in pain in the group treated with indomethacin (p < 0.0001), which also demonstrated a lower sensitivity to pain in the case of larger lesions (p < 0.0001). There was no difference in the healing time between groups, and the reduction of pain is not correlated with corneal anesthesia and healing time. CONCLUSIONS: Our study highlighted the efficacy of indomethacin as a pain reducer for acute corneal pathology and suggested that the medication may act on the corneal nociceptors in a qualitative way.

Small incision nucleus capture: results of 200 cases.

PURPOSE: To compare the learning curve in a series of 200 cataract surgeries performed using small incision nucleus capture with that of phacoemulsification as reported in the literature. SETTING: Department of Ophthalmology, University of Genoa, Genoa, Italy. METHODS: Two hundred eyes of 163 consecutive patients with cataract had small incision nucleus capture, a relatively new cataract surgery technique that allows small incisions and in-the-bag intraocular lens implantation. Patients were divided into 4 groups of 50 each according to when they had surgery between August 1996 and October 1997. The incidence of intraoperative complications (capsule break with or without vitreous loss, capsulorhexis tears, Descemet's detachment, transient iris damage) and postoperative complications (raised intraocular pressure, corneal epithelial edema, Descemet's folds, and permanent iris damage) were evaluated at the different time points. Also recorded was final visual acuity. These results were compared with those obtained with phacoemulsification. RESULTS: The study comprised 92 women and 71 men with an age range of 41 to 93 years. Overall final results showed that the learning curve of nucleus capture is comparable to that of phacoemulsification. CONCLUSION: Nucleus capture cataract extraction resulted in a low incidence of complications and good visual recovery that was comparable to that obtained with phacoemulsification.

Echobiometric evaluation of the axial length of the eye and intraocular lens calculation in pseudophakic eyes: our experience.

Intraocular lens (IOL) power calculation in 46 pseudophakic eyes (extracapsular cataract extraction with IOL in posterior chamber), utilizing a Javal keratometer, a Sonomed A 2000 echobiometer (probe 10 MHz, velocity=1,548 m/s) and the SRK2 formula, although there was a statistically significant reduction of the axial length, both in normal and hyperopic eyes, demonstrated no statistically significant differences of IOL power, when compared to the power previously calculated in the phakic eye.

Effect of betaxolol on primary open-angle glaucoma and normal-tension glaucoma patients.

This study is aimed at establishing the efficacy of the therapeutic agent, betaxolol, in diurnal control of IOP (intraocular pressure). Therapy was performed on 32 eyes affected by POAG (primary open-angle glaucoma) and 16 eyes affected by NTG (normal-tension glaucoma). Two preparations of betaxolol were utilized: betaxolol hydrochloride 0.50% (Betoptic) was administered to 15 POAG and 7 NTG eyes; betaxolol hydrochloride 0.25% ophthalmic suspension (Betoptic S) was administered to 17 POAG and 9 NTG eyes. IOP measurements were taken every two hours from 8 a.m. to 8 p.m. IOP was measured before therapy and at 12 hours, 30 days, and 3 months of therapy. Betaxolol hydrochloride 0.50% was more effective at lowering IOP during the day. Diurnal pressure peaks, which are a risk factor concerning the maintenance of visual field in glaucoma patients, were also reduced using 0.50% betaxolol hydrochloride.

Role of impression cytology during hypovitaminosis A.

AIMS: Evaluation of the morphological damage to the ocular surface of patients operated for biliopancreatic diversion for pathological obesity and the correlation of impression cytology with vitamin A plasma levels, adaptometry, and other general variables. METHODS: 48 patients (15 males, 33 females, age range 21-73) and 34 normal subjects were examined with fluorescein and rose bengal, a plasma dose of vitamin A, and adaptometry. The results of the various tests were subdivided into three levels (0 = normal, 1 = moderately altered, 2 = seriously altered). The impression cytology and adaptometry results were correlated with vitamin A levels and other patient data (age, nutritional condition, time since operation, percentage weight loss). All the examinations were repeated after intramuscular therapy with vitamin A. RESULTS: Corneoconjunctival alterations visible with fluorescein and rose bengal staining were present in 67.7% of cases, impression cytology alterations in 93.7%, adaptometric alterations in 82.2%; vitamin A plasma levels were below normal in 95.8% of cases. After the therapy with vitamin A a significant reduction was found for every examination. The correlation between impression cytology and adaptometry and vitamin A plasma levels and between corneoconjunctival alterations and vitamin A plasma levels was significant. There was no significant correlation between impression cytology and nutritional condition, age time since operation, and percentage weight loss. CONCLUSION: These results show impression cytology is a specific indicator for hypovitaminosis A because it is not influenced by other factors related to the general condition of the patient. Many patients with hypovitaminosis A not demonstrating ocular symptoms of changes visible with fluorescein and rose bengal showed alterations with impression cytology.

Fluctuations of intraocular pressure during the day in open-angle glaucoma, normal-tension glaucoma and normal subjects.

PURPOSE: The aim of the present paper is to describe the variations in intraocular pressure (IOP) during the day in normals, in patients with primary open-angle glaucoma (POAG) and in patients with normal-tension glaucoma (NTG). IOP represents one of the most important risk factors for glaucoma. However the IOP value is not constant during the day and IOP fluctuation could influence the diagnostic and prognostic evaluation of the glaucomatous disease. METHODS: For this purpose IOP was evaluated every 2 h from 8 a.m. to 8 p.m. in one randomized eye of 33 normal subjects, 95 POAG and 50 NTG patients. RESULTS: The results show that the highest IOP values were detectable in the morning in all three groups. The lowest values were found in the early afternoon hours. These variations were most evident in POAG patients. The daily IOP fluctuations were directly proportional to IOP level. CONCLUSION: The study evidents that a single tonometric evaluation, especially if done in the first hours of the afternoon, is not sufficient to correctly evaluate the IOP-related risk in glaucoma patients. If the pressure peaks are important in determining the extent of glaucomatous damage in NTG patients IOP should not have an important role in optic nerve head damage. The use of diurnal curves seems to be mandatory for the assessment of IOP-related risk and of efficiency of the therapeutic approach.

Congenital glaucoma associated with an arginase deficit: a case report.

PURPOSE: Argininemia is a rare congenital disease caused by deficiency. We present here a case in which argininemia and buphthalmos are associated. CASE REPORT: This female patient, who died at eighteen, had intraocular pressure correlated with her blood levels or arginine. Hyperammoniemia causes a change in pH that could explain an increase in aqueous humor (from the ciliary body). Ammonium ions could also damage the matrix of the trabecular meshwork. CONCLUSIONS: The association of argininemia and buphthalmos have never been described. This might be because of the different expressions of the disease, or because nobody has ever looked for ocular hypertension in these patients unless they had subjective symptoms.

The mixed leukocyte reaction may promote the growth of human pericorneal fibroblasts in primary culture.

Keratitis of the cornea is believed to be the leading cause of loss of vision due to external eye disease in the United States and in Europe. At present, primary and secondary cell cultures obtained from corneal explants are the most suitable system to study the role of drug-resistant viruses in severe recurrences of infections leading to irreversible corneal scarring. In this work, the growth of human pericorneal fibroblasts in primary and secondary cultures has been obtained by cocultivation with a mixed leukocyte reaction.