Jacek P. Szaflik

Endophthalmitis prophylaxis in cataract surgery: overview of current practice patterns in 9 European countries.

Data on practice patterns for prophylaxis against infectious postoperative endophthalmitis (IPOE) during cataract surgery in 9 European countries were searched in national registers and reviews of published surveys. Summary reports assessed each nations IPOE rates, nonantibiotic prophylactic routines, topical and intracameral antibiotic use, and coherence to the European Society of Cataract & Refractive Surgeons (ESCRS) 2007 guidelines. Although the reliability and completeness of available data vary between countries, the results show that IPOE rates differ significantly. Asepsis routines with povidone-iodine and postoperative topical antibiotics are generally adopted. Use of preoperative and perioperative topical antibiotics as well as intracameral cefuroxime varies widely between and within countries. Five years after publication of the ESCRS guidelines, there is no consensus on intracameral cefuroxime use. Major obstacles include legal barriers or persisting controversy about the scientific rationale for systematic intracameral cefuroxime use in some countries and, until recently, lack of a commercially available preparation.

Oxidative Stress in the Pathogenesis of Keratoconus and Fuchs Endothelial Corneal Dystrophy

Due to its localization and function, the cornea is regularly exposed to sunlight and atmospheric oxygen, mainly dioxygen, which produce reactive oxygen species (ROS). Therefore, corneal cells are particularly susceptible to oxidative stress. The accumulation of ROS in the cornea may affect signal transduction, proliferation and may also promote cell death. The cornea has several enzymatic and non-enzymatic antioxidants involved in ROS scavenging, but in certain conditions they may not cope with oxidative stress, leading to diseases of the eye. Keratoconus (KC) and Fuchs endothelial corneal dystrophy (FECD) are multifactorial diseases of the cornea, in which pathogenesis is not fully understood. However, increased levels of oxidative stress markers detected in these disorders indicate that oxidative stress may play an important role in their development and progression. These markers are: (i) decreased levels of non-enzymatic antioxidants, and (ii) decreased expression of genes encoding antioxidative enzymes, including thioredoxin reductase, peroxiredoxins, superoxide dismutase, glutathione S-transferase, and aldehyde dehydrogenase. Moreover, the FECD endothelium displays higher levels of oxidative DNA damage, especially in mitochondrial DNA (mtDNA), whereas KC cornea shows abnormal levels of some components of oxidative phosphorylation encoded by mtDNA. In this review we present some considerations and results of experiments supporting the thesis on the important role of oxidative stress in KC and FECD pathology.

An association between vascular endothelial growth factor gene promoter polymorphisms and diabetic retinopathy

BackgroundDiabetic retinopathy is a highly prevalent cause of visual loss in Western countries. Vascular endothelial growth factor (VEGF) is a potent angiogenic factor implicated in the development of the proliferative stage of this disease. Reports have suggested that polymorphisms at positions-460 and -634 of the 5′ untranslated region of the VEGF gene increase its basal promoter activity.MethodsTo investigate whether polymorphisms are associated with diabetic retinopathy, 215 patients with type 2 diabetes mellitus (T2DM) were enrolled. Among them, 82 subjects had proliferative diabetic retinopathy (PDR), 72 had non-proliferative diabetic retinopathy (NPDR), and 61 individuals without retinopathy served as controls. Two polymorphisms of the VEGF gene, a G→C transversion at-634 (the G/C polymorphism) and a C→T transition at-460 (the C/T polymorphism), were investigated by restriction fragment length polymorphism PCR and allele-specific PCR respectively.ResultsWe did not find any association between the C/T polymorphism and diabetic retinopathy. However, the G/C polymorphism genotype distribution and the frequency of the C allele were significantly higher in the NPDR group than in control patients (OR = 1.69, 95% CI = 1.03–2.79). Analysis of the distribution of combined genotypes of the VEGF gene revealed the prevalence of the C/C-C/C genotype in NPDR patients (OR = 8.26, 95% CI = 1.79–37.99) and C/G-CC in PDR patients (OR = 3.36, 95% CI = 1.39–8.12).ConclusionsOccurrence of the -634C allele appears to be associated with increased VEGF gene promoter activity, and the G/C polymorphism might serve as a predictive factor for the development of diabetic retinopathy.

Novel mutation and three other sequence variants segregating with phenotype at keratoconus 13q32 susceptibility locus.

Keratoconus (KTCN), a non-inflammatory corneal disorder characterized by stromal thinning, represents a major cause of corneal transplantations. Genetic and environmental factors have a role in the etiology of this complex disease. Previously reported linkage analysis revealed that chromosomal region 13q32 is likely to contain causative gene(s) for familial KTCN. Consequently, we have chosen eight positional candidate genes in this region: MBNL1, IPO5, FARP1, RNF113B, STK24, DOCK9, ZIC5 and ZIC2, and sequenced all of them in 51 individuals from Ecuadorian KTCN families and 105 matching controls. The mutation screening identified one mutation and three sequence variants showing 100% segregation under a dominant model with KTCN phenotype in one large Ecuadorian family. These substitutions were found in three different genes: c.2262A>C (p.Gln754His) and c.720+43A>G in DOCK9; c.2377-132A>C in IPO5 and c.1053+29G>C in STK24. PolyPhen analyses predicted that c.2262A>C (Gln754His) is possibly damaging for the protein function and structure. Our results suggest that c.2262A>C (p.Gln754His) mutation in DOCK9 may contribute to the KTCN phenotype in the large KTCN-014 family.

Association between vascular endothelial growth factor gene polymorphisms and age-related macular degeneration in a Polish population

The pathogenesis of age-related macular degeneration (AMD) is thought to be determined by an array of environmental and genetic factors. The association of increased expression of vascular endothelial growth factor (VEGF) with AMD, especially the wet form of AMD, was reported in several studies. The VEGF gene is highly polymorphic and some of its polymorphisms may affect its expression. In our work, we searched for an association between the -460C> (rs833061) and -634G>C (rs2010963) polymorphisms of the VEGF gene and the occurrence of AMD and its dry and wet forms. We have chosen these polymorphisms because they were shown to be significant in other studies and we previously showed their association with diabetic retinopathy. A total of 401 individuals were enrolled in this study: 136 controls, and 88 patients with dry and 177 with wet AMD. The polymorphisms were determined with DNA from peripheral blood lymphocytes by allele-specific and restriction fragment length polymorphism polymerase chain reaction. The significance of the polymorphisms was assessed by multiple logistic regression, producing odds ratios (ORs) and 95% confidence intervals (CIs). We observed a weak association (OR 2.90) between AMD occurrence and the C/T genotype of the -460C>T polymorphism. An association (OR 3.77) between the C/T genotype of the -460C>T polymorphism and the occurrence of dry AMD was observed. The T/T genotype considerably lowered the risk of dry AMD (OR 0.19). Dry AMD was associated with the C/C genotype of the -634G>C polymorphism (OR 3.68). Another weak association (OR 2.63) was found between the C/T genotype of the -460C>T polymorphism and the occurrence of wet AMD. The occurrence of AMD was correlated with the presence of the combined C/T-G/G genotype of both polymorphisms (OR 2.41), whereas the T/T-G/G and T/T-G/C genotypes exerted a protective effect against the disease (OR 0.22 and 0.48, respectively). The presence of the C/T-G/G and C/T-C/C combined genotypes increased the risk of dry AMD (OR 2.08 and 3.77, respectively), whereas the presence of the T/T-G/G and T/T-G/C genotypes decreased the risk (OR 0.15 and 0.28, respectively). In the wet form of AMD, the combined genotype C/T-G/G slightly favored the disease (OR 2.61) and the T/T-G/G genotype had a protective effect (OR 0.25). Analysis of haplotypes of both polymorphisms yielded similar results for AMD in general as well as for the dry and wet forms of the disease: the CG haplotype favored both forms of AMD, whereas the TG haplotype protected against both forms of AMD. The results obtained indicate that the -460C>T and -634G>C polymorphisms of the VEGF gene may be associated with the dry and wet forms of AMD in a Polish population.

Evaluation of oxidative stress markers in pathogenesis of primary open-angle glaucoma

Primary open-angle glaucoma (POAG) is the leading cause of blindness in the industrial countries. It is reported that oxidative stress might be an important risk factor in the pathogenesis of POAG. Forty subjects including 20 patients with open-angle glaucoma (9 men and 12 women, mean age 61.8±12.1yr) and 20 controls without glaucoma symptoms (9 men and 12 women, mean age 58.1±17.7yr) were enrolled in our study. The main aim of the work was to evaluate oxidative stress markers in the pathogenesis of open-angle glaucoma. In our work the activity of antioxidant enzymes: catalase (CAT), superoxide dismutase (SOD) and glutathione peroxidase (GPX) as well as the total antioxidant status (TAS) was estimated. An alkaline comet assay was used to measure DNA damage of strand breaks (SB), oxidized purines as glicosylo-formamido-glicosylase (Fpg) sites, and oxidized pirmidines as endonuclease III (Nth) sites. We measured endogenous as well as exogenous DNA damage after 10μM hydrogen peroxide treatment (H(2)O(2)). We did not observe any statistical changes of DNA strand break lesion in examined POAG patients according to healthy subjects (P>0.05). However, either endogenous (P<0.01) or exogenous (P<0.001) levels of oxidative DNA damage in POAG patients were found to be statistically higher than controls. A significant decrease of antioxidant enzymes: CAT (P<0.001), SOD (P<0.05), and GPX (P<0.001) and a non-statistical decrease of TAS status (P>0.05) in glaucoma patients according to controls were also indicated. In conclusion our data revealed that oxidative stress had a pathogenic role in primary open-angle glaucoma. Therefore, we suggested that the modulation of a pro-oxidant/antioxidant status might be a relevant target for glaucoma prevention and therapy.

DNA damage and repair in age-related macular degeneration

Age-related macular degeneration (AMD) is a retinal degenerative disease that is the main cause of vision loss in individuals over the age of 55 in the Western world. Clinically relevant AMD results from damage to the retinal pigment epithelial (RPE) cells thought to be mainly caused by oxidative stress. The stress also affects the DNA of RPE cells, which promotes genome instability in these cells. These effects may coincide with the decrease in the efficacy of DNA repair with age. Therefore individuals with DNA repair impaired more than average for a given age may be more susceptible to AMD if oxidative stress affects their RPE cells. This may be helpful in AMD risk assessment. In the present work we determined the level of basal (measured in the alkaline comet assay) endogenous and endogenous oxidative DNA damage, the susceptibility to exogenous mutagens and the efficacy of DNA repair in lymphocytes of 100 AMD patients and 110 age-matched individuals without visual disturbances. The cells taken from AMD patients displayed a higher extent of basal endogenous DNA damage without differences between patients of dry and wet forms of the disease. DNA double-strand breaks did not contribute to the observed DNA damage as checked by the neutral comet assay and pulsed field gel electrophoresis. The extent of oxidative modification to DNA bases was greater in AMD patients than in the controls, as probed by DNA repair enzymes NTH1 and Fpg. Lymphocytes from AMD patients displayed a higher sensitivity to hydrogen peroxide and UV radiation and repaired lesions induced by these factors less effectively than the cells from the control individuals. We postulate that the impaired efficacy of DNA repair may combine with enhanced sensitivity of RPE cells to blue and UV lights, contributing to the pathogenesis of AMD.

Therapeutic use of a silicone hydrogel contact lens in selected clinical cases.

Purpose. To evaluate a silicone hydrogel contact lens as a continuous-wear bandage in selected clinical cases, a prospective, open, nonrandomized clinical study was conducted. Methods. Seventy eyes from 70 patients with anterior segment conditions for which therapeutic lenses were indicated were enrolled in the study. In most patients (47 eyes, 67%), the presenting condition was bullous keratopathy. Patients were fitted with a lotrafilcon A silicone hydrogel lens (Focus Night & Day) that was worn continuously for 7 to 30 days, and the concomitant therapies were used. The corneal condition was assessed, and subjective comfort was recorded for 7 days to 18 months after fitting (mean follow-up, 69 days). Results. Of 70 eyes, 64 (91%) showed improvement in the clinical condition of the eye, and 66 (94%) rated comfort when wearing the lens as very good or good. Conclusions. The study lens was found to be an effective and well-tolerated bandage lens in these selected patients.

Therapeutic use of a lotrafilcon A silicone hydrogel soft contact lens as a bandage after LASEK surgery.

Purpose. Soft contact lenses may be used as a bandage after corneal refractive surgery. Silicone hydrogel contact lenses offer the advantage of increased oxygen permeability over conventional hydrogel lenses for this application. To evaluate a silicone hydrogel contact lens as a continuous wear bandage applied after laser-assisted sub-epithelial keratomileusis (LASEK). Methods. We conducted a prospective, open-label, non-randomized clinical trial involving thirty patients treated with unilateral LASEK. Patients were fitted with a lotrafilcon A (Focus NIGHT & DAY; CIBA Vision, Duluth, GA) silicone hydrogel soft contact lens that was worn continuously for 3 to 4 days post-operatively. Lens movement and slitlamp evaluations of conjunctival hyperemia 1, 2, and 3 days after surgery (DAS) were recorded, as was the condition of the corneal epithelium after lens removal. Subjective comfort was rated by the patients along with the presence of symptoms or pain. Results. Post-blink lens movement was evaluated by investigators as ‘very good’ or ‘good’ in 87% of eyes at 1 DAS, in 73% at 2 DAS, and in 60% at 3 DAS. Conjunctival hyperemia was assessed as normal or trace in 96% of eyes at 1 DAS, in 76% at 2 DAS, and in 67% at 3 DAS. An average of 80% of eyes showed normal or trace conjunctival hyperemia during the trial. The condition of the corneal epithelium after contact lens removal was rated as ‘very good’ in 13% or ‘good’ in 73% of eyes. An average of 77% of subjects reported ‘very good’ or ‘good’ comfort during the trial. Symptoms or pain were rated as ‘absent’ or ‘mild’ by all subjects at 1 DAS and 2 DAS and by 96% at 3 DAS. An average of 99% rated symptoms or pain as either ‘absent’ or ‘mild’ during the trial. Conclusions. The lotrafilcon A lens was found to be an effective and well-tolerated bandage lens after LASEK.

Mutagenesis of mitochondrial DNA in Fuchs endothelial corneal dystrophy.

Fuchs endothelial corneal dystrophy (FECD) is an age-related, slowly progressive disease, which may lead to loss of vision resulting from apoptosis of corneal endothelial (CE) cells, dysfunction of Descemet membrane (DM) and corneal edema. A growing body of evidence suggests that oxidative stress may play a major role in the pathogenesis of FECD and that mitochondria of CE cells are its main target. Mitochondrial DNA (mtDNA) is particularly prone to oxidative stress and changes in mtDNA were reported in FECD patients. In the present work we studied mtDNA damage and repair, mtDNA copy number, and the 4977bp common deletion in mtDNA in DM cells and peripheral blood lymphocytes (PBLs) isolated from FECD patients. PBLs from 35 FECD patients and 32 controls were challenged for 10min with hydrogen peroxide at 20μM and then left in a fresh medium for 3h, resulting in a decrease in mtDNA copy number in both groups. Damage to mtDNA was not fully repaired after 3h and the extent of remaining lesions was significantly higher in the patients than the controls. We observed a higher copy number and an increased extent of mtDNA damage as well as a higher ratio of the common 4977bp deletion in DM cells of FECD patients than the controls. Our results confirm that mutagenesis of mtDNA may be involved in FECD pathogenesis and disturbance in mtDNA sensitivity to damaging agent as well as changes in mtDNA damage repair along with alternations in mtDNA copy number may underline this involvement.