Reexamining the hyperglycemic pseudohypoxia hypothesis of diabetic oculopathy

PURPOSE: To test the hypothesis that diabetes alters retinal NAD+-to-NADH ratios early in the course of the disease (e.g., the hyperglycemic pseudohypoxia hypothesis). METHODS: In freshly excised age-matched control and diabetic rat retinas, measurements were made of the NAD+ and NADH content as well as a surrogate marker of NAD+-to-NADH ratios obtained from lactate and pyruvate levels. In addition, the effect of various hyperglycemic levels was assessed from measurements of retinal lactate and pyruvate concentrations and the rate of lactic acid production in vitro (isolated rat retinas, monolayer cultures of human retinal pigment epithelial cells, and rabbit lens epithelial cells). RESULTS: No significant differences (P>0.05) were found between control and diabetic tissues in their amount of total NAD+ and NADH/retina, and the ratio of NAD+ to NADH, or in their content of lactate, pyruvate, and adenosine triphosphate (ATP) or in the ratio of lactate to pyruvate. The content of lactate and pyruvate in retinas incubated for 2 hours in media containing 10 or 30 mM glucose was the same as found in fresh tissues, but the levels of these metabolites in retinas incubated in media containing 5 mM glucose declined in comparison to the fresh values. There were no significant differences in lactate content in cultured retinal and lens cells that were exposed to 5 or 30 mM glucose-containing media. DISCUSSION: The present results do not support the hyperglycemic pseudohypoxia hypothesis of diabetic retinopathy

Two autocrine pathways to regulate cyclic GMP synthesis in cultured human retinal pigment epithelial cells

AIM: To investigate the role of two separate enzymatic pathways [soluble (sGC) vs. particulate (pGC) guanylyl cyclase] in the synthesis of cyclic GMP (cGMP) in cultured human retinal pigment epithelial (RPE) cells. METHODS: cGMP accumulation was evaluated by quantitative analysis of cGMP immunoreactivity. RPE cells were also stained for inducible nitric oxide synthase (iNOS), ANP and beta(1)- and alpha(2)-subunits of sGC. RESULTS: We showed nicotinamide adenine dinucleotide phosphate (NADPH) diaphorase activity and iNOS immunoreactivity in RPE cells. Incubation of the cells in the presence of 1 mM IBMX to inhibit phosphodiesterase activity, and the simultaneous inhibition of NOS activity with L-NAME suggested the involvement of sGC in maintaining a low level of cGMP in the RPE cells. The involvement of sGC was further supported by detection of the beta(1)- and alpha(2)-subunits of sGC. Incubation of the cells in the presence of atrial natriuretic peptide (ANP) to stimulate pGC strongly increased cGMP immunoreactivity. We also demonstrated the presence of ANP in all RPE cells. CONCLUSION: Cultured human RPE cells are capable of producing cGMP after stimulation of sGC or pGC. The presence of iNOS and ANP in all cells suggests two different autocrine pathways of stimulating cGMP production in these cells. The possible role of cGMP in the regulation iNOS gene expression and in the regulation of ANP is discussed

Increased nitric oxide (NO) pathway metabolites in the vitreous fluid of patients with rhegmatogenous retinal detachment or diabetic traction retinal detachment

BACKGROUND: Nitric oxide (NO) plays a significant role in physiological and pathological processes in the retina. In the L-arginine-NO pathway, NO synthase (NOS) converts L-arginine to NO and L-citrulline. Increased NO production, mediated by inducible NOS has been implicated in the pathogenesis of various vitreoretinal diseases. In the present study it is hypothesized that in rhegmatogenous retinal detachment (RRD), the production of NO pathway metabolites might be upregulated. METHODS: Using high-pressure liquid chromatography citrulline, arginine and nitrite were measured in vitreous fluid of 93 eyes with RRD, nine eyes with a traction retinal detachment due to proliferative diabetic retinopathy (PDR), and in 49 control samples of vitreous fluid from eyes without retinal detachment. RESULTS: The mean vitreous concentrations of citrulline and arginine were significantly increased in eyes with RRD (9.6+/-4.3 and 97.3+/-29.2; respectively) or in eyes with a traction retinal detachment (25.8+/-10.3 and 130.7+/-23.7; respectively) as compared to control eyes (7.1+/-3.2 and 75.9+/-18.1; respectively). The mean level of nitrite was also higher in vitreous fluid of patients with RRD (2.24+/-1.4) or patients with a traction retinal detachment (2.21+/-0.72) than in the controls (2.01+/-0.72), although not significantly so. CONCLUSIONS: We found increased levels of NO pathway metabolites in the vitreous fluid of eyes with retinal detachment, which may reflect a possible role of NO in the pathogenesis of this disease

Scleral buckling surgery after macula-off retinal detachment: worse visual outcome after more than 6 days

PURPOSE: To determine the effect of duration of macular detachment (DMD) on visual acuity (VA) in patients with macula-off rhegmatogenous retinal detachment (RD). DESIGN: Retrospective observational case series. PARTICIPANTS: Two hundred two consecutive patients (202 eyes) with primary uncomplicated macula-off RD, preoperative VA of 10/100 or worse, a precise history of when macular function was lost, successful reattachment surgery, and a minimal follow-up of 3 months. INTERVENTION: All RDs were repaired with a primary scleral buckling procedure performed by 3 vitreoretinal surgeons. MAIN OUTCOME MEASURE: Visual acuity (best corrected and 3, 6, and 12 months postoperatively). RESULTS: Considering all eyes, the cumulative mean of the best-corrected postoperative VA (logarithm of the minimum angle of resolution [logMAR]) as a function of DMD shows a rapid worsening when DMD exceeds 6 days. Eyes were divided into 3 groups, corresponding to the DMD intervals immediate (within 10 days), delayed (11 days-6 weeks), and late (>6 weeks). Mean postoperative VAs (in logMAR) were 0.35+/-0.31 (between 20/40 and 20/50 Snellen equivalent) in eyes with DMD up to 10 days, 0.48+/-0.26 (20/60 Snellen equivalent) in the delayed group, and 0.86+/-0.30 (8/60 Snellen equivalent) in eyes with DMD longer than 6 weeks. CONCLUSIONS: The cumulative mean of the best-corrected postoperative VA (logMAR) as a function of DMD shows a rapid worsening when DMD exceeds 6 days. Our results indicate that the scleral buckling procedure should be done preferably within a 7-day DMD

Cyclic GMP synthesis by human retinal pigment epithelial cells is mainly mediated via the particulate guanylyl cyclase pathway

BACKGROUND/AIMS: Cyclic 3',5'-guanosine monophosphate (cGMP), a central molecule in the phototransduction cascade, is also involved in a number of other physiological processes in the retina, like stimulating the absorption of subretinal fluid by activating the retinal pigment epithelium (RPE) cell pump. The aim of this study was to quantify cGMP synthesis by RPE cells and to investigate the role of two separate enzymatic pathways (soluble versus particulate guanylyl cyclase) in its production. METHODS: cGMP expression was evaluated by immunochemistry and radioimmunoassay following culture of the D407 RPE cell line in the presence of a nonselective phosphodiesterase inhibitor (IBMX), in combination with the particulate guanylyl cyclase stimulator atrial natriuretic peptide (ANP) or the soluble guanylyl cyclase stimulator sodium nitroprusside (SNP). RESULTS: Stimulation of the particulate guanylyl cyclase in RPE cells with ANP resulted in high intra- and extracellular cGMP levels. Stimulation of the soluble guanylyl cyclase by SNP resulted in a slight elevation of cGMP levels compared to controls. CONCLUSIONS: These results show that cultured human RPE cells are capable of producing cGMP and that most cGMP is generated following stimulation of the particulate guanylyl cyclase pathway

Balance of vascular endothelial growth factor and pigment epithelial growth factor prior to development of proliferative vitreoretinopathy

BACKGROUND: Pigment epithelium-derived factor (PEDF) and vascular endothelial growth factor (VEGF) are imbalanced in eyes with proliferative diabetic retinopathy or proliferative vitreoretinopathy (PVR). It is not known whether such an imbalance is already present in early PVR stages. We therefore analyzed VEGF and PEDF concentrations in subretinal fluids prior to PVR development. METHODS: A large number (n = 137) of subretinal fluid samples were obtained at the time of scleral buckling surgery for rhegmatogenous retinal detachment (RRD). Thirty patients developed PVR within 6 months after surgery. One hundred and seven patients undergoing the same surgery but without complications served as controls. Furthermore, vitreous from 16 patients with macular hole or pucker (MHP) served as reference for baseline intraocular concentrations. PEDF and VEGF concentrations were measured by commercial ELISAs. RESULTS: PEDF levels were substantially higher (9.6 microg/ml) compared to MHP vitreous (0.3 microg/ml, p < 0.001). VEGF levels were also higher (RRD: 0.07 ng/ml; MHP: 0.01 ng/ml, p < 0.05). Subretinal concentrations were not significantly different between PVR and control RRD patients. CONCLUSIONS: Although both VEGF and PEDF are increased at first surgery for RRD, they do not predict PVR development later on. The high PEDF concentrations and its known antiangiogenic activity suggest a protective role against neovascularization

Incidence of redetachment 6 months after scleral buckling surgery

PURPOSE: The preoperative and intraoperative clinical variables associated with redetachment and/or a poor visual outcome following scleral buckling (SB) surgery for rhegmatogenous retinal detachment (RRD) have mainly been studied after a short follow-up. This study aimed to analyse long-term effects by following patients for at least 6 months. METHODS: In a retrospective survey we evaluated the data of 436 eyes that underwent SB surgery. Postoperative data were collected at 3-month intervals. RESULTS: After a mean follow-up period of 51 months, anatomic reattachment was achieved in 76% after one SB procedure, with a final reattachment rate of 97% after additional vitreoretinal procedures. In total, 104 eyes developed redetachment during follow-up. After more than 6 and 12 months of follow-up, 32 eyes (7%) and 20 eyes (5%), respectively, developed redetachment. Multivariate regression analysis showed that recurrent redetachment and more than 7 days of visual field loss were significant predictors for a poor postoperative visual outcome at 12 months. A cumulative size of the tear of more than three disc diameters was a significant predictor of recurrent RRD. CONCLUSION: Conventional SB surgery is a reliable procedure in a selected group of eyes with primary RRD. However, in eyes with a retinal tear with a cumulative size of more than three disc diameters, a primary vitrectomy should be considered. Taking into account that 7% of eyes developed redetachment after 6 months, a longer follow-up period seems necessary to evaluate the anatomical and visual outcomes after SB surgery