Quantitation of Pax-6 protein in ocular impression cytology samples using an electrochemiluminescence immunoassay.

Paired box protein Pax-6 (oculothrombin) is a transcription factor that plays an important regulatory role in ocular, brain, and pancreatic development. Mutations of the PAX6 gene cause aniridia and Peters anomaly. Reduction in Pax-6 protein is also associated with ocular diseases such as dry eye. An electrochemiluminescence immunoassay method using the Meso Scale Discovery platform was developed to measure Pax-6 protein levels in corneal epithelial cells obtained by impression cytology. Impression cytology involves harvesting ocular epithelial cells by applying a polyethersulfone membrane patch briefly to the ocular surface using a commercially available EYEPRIM™ device. The epithelial cells that adhere to the membrane patch of the EYEPRIM™ device provide a biological sample which can be assayed for Pax-6 protein levels. Assay development identified an antibody pair capable of detecting purified recombinant Pax-6 protein produced in mammalian cells. The optimized assay has a dynamic range of 24 pg mL-1 to 100,000 pg mL-1 and a lower limit of quantification of 24 pg mL-1. Assay selectivity was demonstrated using either HeLa or HEK293 cells transfected with inhibitory RNA. Finally, the method was validated by measuring Pax-6 protein levels in impression cytology acquired samples obtained using the EYEPRIM™ device from rabbit cornea.

Complement-Mediated Activation of the NLRP3 Inflammasome and Its Inhibition by AAV-Mediated Delivery of CD59 in a Model of Uveitis.

Uveitis is an inflammatory disorder of the eye responsible for approximately 10%-15% of blindness in the US. In this study, we examined the role of the complement membrane attack complex (MAC) and the NLRP3 inflammasome in the pathogenesis of experimental autoimmune uveitis (EAU) in normal and C9-/- mice that are incapable of assembling the MAC. We discovered that the MAC and the NLRP3 inflammasome and associated production of IL-1ß are elevated in EAU mice and that MAC may be involved in regulation of Th1 and Th17 cell differentiation. In contrast, MAC and the NLRP3 inflammasome were not elevated in C9-/- mice. However, EAU-associated pathophysiology including retinal structure and function were not rescued in C9-/- mice. Unexpectedly, AAV-mediated delivery of sCD59, an inhibitor of C9 incorporation into the MAC, successfully attenuated activation of the NLRP3 inflammasome and EAU pathology as well as MAC. Our studies provide an improved understanding of the role of the MAC and the NLRP3 inflammasome in EAU as well as suggest a novel approach for the treatment of uveitis.

G-quartet oligonucleotide mediated delivery of functional X-linked inhibitor of apoptosis protein into retinal cells following intravitreal injection.

There is currently no efficient method available for the delivery of full length functional proteins into the cytoplasm of retinal cells in vivo. Historically, the most successful approach for the treatment of retinal diseases has been intravitreal injection of antibodies or recombinant proteins, but this approach is not yet utilized for the delivery of proteins that require intracellular access for a therapeutic effect. Here we describe a platform for the delivery of functional proteins into ganglion cells, photoreceptors and retinal pigment epithelium via intravitreal injection. A nucleolin binding aptamer, AS1411, was biotinylated and complexed with traptavidin and utilized as a platform for the delivery of GFP or X-linked inhibitor of apoptosis (XIAP) proteins by intravitreal injection in BALB/c mice. Retinal sections were analyzed for uptake of proteins in the retina. Apoptosis was induced by intravitreal injection of N-methyl-D-aspartate (NMDA). Retinas were harvested for analysis of TUNEL and caspase 3/7 activity. Intravitreal injection of AS1411-directed GFP or XIAP complexes enabled delivery of these proteins into ganglion cells, photoreceptors and retinal pigment epithelium in vivo. AS1411-XIAP complexes conferred significant protection to cells in the outer and inner nuclear layers following NMDA induced apoptosis. A concomitant decrease in activity of Caspase 3/7 was observed in eyes injected with the AS1411-XIAP complex. In conclusion, AS1411 can be used as a platform for the delivery of therapeutic proteins into retinal cells. This approach can potentially be utilized to introduce a large variety of therapeutically relevant proteins that are previously well characterized to maintain the structural integrity and function of retina, thus, preventing vision loss due to ocular trauma or inherited retinal degeneration.

Evaluation of Supportive Care Management Outcomes in Cancer Chemotherapy: A Prospective Observational Study in a Tertiary Care Teaching Hospital in South India.

AIMS: Evaluation of supportive care management of cancer patients experiencing drug-related problems (DRPs) is a challenge because it might increase the cost due to additional therapy. The main objectives of this study were to estimate chemotherapy-associated drug-related hospital admissions in the department of medical oncology and to estimate the cost of managing chemotherapy-associated DRPs. SETTINGS AND DESIGN: This study is a prospective observational study. SUBJECTS AND METHODS: Patients with chemotherapy-related DRPs were prospectively identified from the patient's medical records. The contribution of DRPs and cost incurred due to each hospitalization was assessed. STATISTICAL ANALYSIS USED: Data were analyzed using SPSS® 20.0 version. RESULTS: Out of 55 patients analyzed for DRPs, 25 (45.5%) patients in the age group of 51-60 years experienced DRPs most frequently. Most commonly occurring DRP was adverse drug reactions 42 (76.4%), which were more frequent in females. DRPs were maximum with alkylating agents 15 (27.3%) and the least with hormonal agents 1 (1.8%). The mean length of hospitalization was 9.6 ± 6.5 days. The total direct medical cost was Rs. 31,540 ± 42,476, of which medicine cost accounted for Rs. 16,550 ± 25,404, constituting a major share of the total medical costs. CONCLUSIONS: Pharmacists can provide better patient care by identifying and preventing DRPs and reducing drug-related morbidity and mortality.

Diabetic retinopathy and transcriptional regulation of a small molecular weight G-Protein, Rac1.

In diabetic retinopathy, increased cytosolic reactive oxygen species, produced by NADPH oxidase (Nox), damage mitochondria, and this accelerates apoptosis of retinal capillary cells, resulting in the histopathology. Activation of Nox2 is mediated by a small molecular weight GTPase, Rac1, and retinal Rac1 is activated in diabetes. Our goal is to investigate the molecular mechanism responsible for transcriptional activation of Rac1 in the development of diabetic retinopathy. Using retinal microvessels, the site of histopathology associated with diabetic retinopathy, from streptozotocin-induced diabetic rats, we investigated the binding of the nuclear transcriptional factor-kB (NF-kB) at Rac1 promoter. Since activation of NF-kB is regulated by its acetylation-deacetylation, the role of acetylation in Rac1 transcription was confirmed in the retina from diabetic mice overexpressing a deacetylase, Sirtuin 1. Diabetes increased the binding of p65 subunit of NF-kB at the Rac1 promoter. Overexpression of Sirtuin 1 prevented hyper-acetylation of p65, decreased its binding at the Rac1 promoter and ameliorated Rac1-Nox2 mediated mitochondrial damage. Thus, in diabetes Rac1 transcriptional activation in the retina is mediated by acetylation of NF-kB, and modulation of acetylation during the early stages of diabetic retinopathy has potential to inhibit/retard its development.

Cardioprotective effects of rutin via alteration in TNF-α, CRP, and BNP levels coupled with antioxidant effect in STZ-induced diabetic rats.

Diabetic cardiomyopathy (DCM) is a dreadful complication of diabetes responsible for 80 % mortality in diabetic patients, but unfortunately its pharmacotherapy is still incomplete. Rutin is a naturally occurring flavonoid having a long history of use in nutritional supplements for its action against oxidative stress, inflammation, and hyperglycemia, the key players involved in the progression of DCM, but remains unexplored for its role in DCM. This study was conducted to address this lacuna. It was performed in 4-week-old Streptozotocin-induced (45 mg/kg) diabetic rats for a period of 24 weeks to mimic the cardiotoxic effect of chronic hyperglycemia in diabetic patient's heart and to investigate the effect of rutin (50 mg/kg/day) in ameliorating these effects. Heart of the diabetic rats showed altered ECG parameters, reduced total antioxidant capacity, increased inflammatory assault, and degenerative changes. Interestingly, rutin treatment significantly ameliorated these changes with decrease in blood glucose level (p > 0.001), % HbA1c (p > 0.001) and reduced expression of TNF-α (p < 0.001), CRP (p < 0.001), and BNP (p < 0.01) compared to diabetic control rats. In addition, rutin provided significant protection against diabetes associated oxidative stress (p < 0.05), prevented degenerative changes in heart, and improved ECG parameters compared to diabetic control rats. The heart-to-body weight ratio was significantly reduced in rutin treatment group compared to diabetic control rats (p < 0.001). In conclusion, this study implicates that oxidative stress and inflammation are the central players involved in the progression of DCM and rutin ameliorates DCM through its antioxidant and anti-inflammatory actions on heart.

Tiam1-Rac1 Axis Promotes Activation of p38 MAP Kinase in the Development of Diabetic Retinopathy: Evidence for a Requisite Role for Protein Palmitoylation.

BACKGROUND/AIMS: Evidence in multiple tissues, including retina, suggests generation of reactive oxygen species (ROS) and the ensuing oxidative stress as triggers for mitochondrial defects and cell apoptosis. We recently reported novel roles for Tiam1-Rac1-Nox2 axis in retinal mitochondrial dysfunction and cell death leading to the development of diabetic retinopathy. Herein, we tested the hypothesis that activation of p38 MAP kinase, a stress kinase, represents the downstream signaling event to Rac1-Nox2 activation in diabetes-induced metabolic stress leading to capillary cell apoptosis. METHODS: Activation of p38 MAP kinase was quantified by Western blotting in retinal endothelial cells incubated with high glucose (20 mM) for up to 96 hours, a duration where mitochondrial dysfunction and capillary cell apoptosis can be observed. NSC23766 and 2-bromopalmitate (2-BP) were used to assess the roles of Tiam1-Rac1 and palmitoylation pathways, respectively. RESULTS: Activation of p38 MAP kinase was observed as early as 3 hours after high glucose exposure, and continued until 96 hours. Consistent with this, p38 MAP kinase activation was significantly higher in the retina from diabetic mice compared to age-matched normal mice. NSC23766 markedly attenuated hyperglycemia-induced activation of p38 MAP kinase. Lastly, 2-BP inhibited glucose-induced Rac1, Nox2 and p38 MAP kinase activation in endothelial cells. CONCLUSIONS: Tiam1-Rac1-mediated activation of Nox2 and p38 MAP kinase constitutes early signaling events leading to mitochondrial dysfunction and the development of diabetic retinopathy. Our findings also provide the first evidence to implicate novel roles for protein palmitoylation in this signaling cascade.

Differential Expression of AQP1 and AQP4 in Avascular Chick Retina Exposed to Moderate Light of Variable Photoperiods.

Aquaporins (AQPs) are integral membrane proteins which maintain cellular water and ion homeostasis. Alterations in AQP expression have been reported in rod-dominated rodent retinas exposed to light. In rodents and also in birds, light of moderate intensities (700-2000 lux) damages the retina, though detailed changes were not examined in birds. The aim of our study was to see if light affects cone dominated retinas, which would be reflected in expression levels of AQPs. We examined AQP1 and AQP4 expressions in chick retina exposed to 2000 lux under 12 h light:12 h dark (12L:12D; normal photoperiod), 18L:6D (prolonged photoperiod) and 24L:0D (constant light). Additionally, morphological changes, apoptosis (by TUNEL) and levels of glutamate and GFAP (a marker of injury) in the retina were examined to correlate these with AQP expressions. Constant light caused damage in outer and inner nuclear layer (ONL, INL) and ganglion cell layer (GCL). Also, there were associated increases in GFAP and glutamate levels in retinal extracts. In normal photoperiod, AQP1 was expressed in GCL, outer part of INL and photoreceptor inner segments of. AQP4 was additionally expressed in nerve fiber layer. Immunohistochemistry and Western blotting revealed over all decreased AQP1 and AQP4 expression in constant light condition compared to those in other two groups. The elevated GFAP and glutamate levels might be involved in the reduction of AQPs in constant light group. Such decreases in AQP expressions are perhaps linked with retinal cell damage seen in constant light condition, while their relatively enhanced expression in two other conditions may help in maintaining a normal retinal architecture, indicating their neuroprotective potential.

Retinal neuroprotective effects of quercetin in streptozotocin-induced diabetic rats.

The aim of the present study was to evaluate the effects of Quercetin (Qctn), a plant based flavonol, on retinal oxidative stress, neuroinflammation and apoptosis in streptozotocin-induced diabetic rats. Qctn treatment (25- and 50 mg/kg body weight) was given orally for six months in diabetic rats. Retinal glutathione (GSH) and antioxidant enzymes [superoxide dismutase (SOD) and catalase (CAT)] were estimated using commercially available assays, and inflammatory cytokines levels [tumor necrosis factor-α (TNF-α), Interleukin-1ß (IL-1ß)] were estimated by ELISA method. Immunofluorescence and western blot studies were performed for nuclear factor kappa B (NF-kB), caspase-3, glial fibrillary acidic protein (GFAP) and aquaporin-4 (AQP4) expressions. Structural changes were evaluated by light microscopy. In the present study, retinal GSH levels and antioxidant enzyme (SOD and CAT) activities were significantly decreased in diabetic group as compared to normal group. However, in Qctn-treated rats, retinal GSH levels were restored close to normal levels and positive modulation of antioxidant enzyme activities was observed. Diabetic retinas showed significantly increased expression of pro-inflammatory cytokines (TNF-α and IL-1ß) as compared to that in normal retinas, while Qctn-treated retinas showed significantly lower levels of cytokines as compared to diabetic retinas. Light microscopy showed significantly increased number of ganglion cell death and decreased retinal thickness in diabetic group compared to those in normal retina; however, protective effect of Qctn was seen. Increased apoptosis in diabetic retina is proposed to be mediated by overexpression of NF-kB and caspase-3. However, Qctn showed inhibitory effects on NF-kB and caspase-3 expression. Microglia showed upregulated GFAP expression, and inflammation of Müller cells resulted in edema in their endfeet and around perivascular space in nerve fiber layer in diabetic retina, as observed through AQP4 expression. However, Qctn treatments inhibited diabetes-induced increases in GFAP and AQP4 expression. Based on these findings, it can be concluded that bioflavonoids, such as Qctn can be effective for protection of diabetes induced retinal neurodegeneration and oxidative stress.

Effects of Trigonella foenum-graecum (L.) on retinal oxidative stress, and proinflammatory and angiogenic molecular biomarkers in streptozotocin-induced diabetic rats.

The aim of the present study was to investigate the protective effects of Trigonella foenum-graecum Linn. (fenugreek) in Streptozotocin-induced diabetic rat retina. Fenugreek (100 and 200 mg/kg body weights) treatment was carried out for 24 weeks and evaluated for inflammatory [tumor necrosis factor (TNF)-α and interleukin (IL)-1ß] and angiogenic [vascular endothelial growth factor (VEGF) and protein kinase C (PKC)-ß] molecular biomarkers. Retinal oxidative stress was evaluated by estimating antioxidant (Glutathione, Superoxide dismutase, and Catalase) parameters. Fluorescein angiography was performed to detect retinal vascular leakage. Electron microscopy was performed to determine basement membrane thickness. In the present study, significant rises in the expressions of retinal inflammatory (TNF-α and IL-1ß) and angiogenic (VEGF and PKC-ß) molecular biomarkers were observed in diabetic retinae compared with normal retinae. However, fenugreek-treated retinae showed marked inhibition in the expression of inflammatory and angiogenic molecular biomarkers. Moreover, results from the present study showed positive modulatory effects of fenugreek on retinal oxidative stress. Fluorescein angiograms and fundus photographs obtained from diabetic retinae showed retinal vascular leakage. On the other hand, fenugreek-treated retinae did not show vascular leakage. Further, thickened BM was recorded in diabetic retina compared with normal retinae. However, fenugreek-treated retinae showed relatively lesser thickening of capillary BM. In conclusion, it may be postulated that fenugreek has great potential in preventing diabetes-induced retinal degeneration in humans after regular consumption in the specified dosage.

Hesperetin rescues retinal oxidative stress, neuroinflammation and apoptosis in diabetic rats.

The purpose of the study was to evaluate the effects of hesperetin (Hsp) on diabetes-induced retinal oxidative stress, neuroinflammation and apoptosis in rats. The Hsp treatment (100 mg/kg body weight) was carried for twenty four weeks in STZ-induced diabetic rats and evaluated for antioxidant (Superoxide dismutase; SOD, Catalase; CAT and glutathione; GSH) enzymes, inflammatory cytokines (TNF-α, IL-1ß), caspase-3, glial fibrillary acidic protein (GFAP) and aquaporin-4(AQP4) expression. Histological changes were evaluated by light and transmission electron microscopic (LM and TEM) studies. Retinal GSH levels and anti-oxidant enzymes (SOD and CAT) activity were significantly decreased in diabetic group as compared to normal group. However, in Hsp-treated rats, retinal GSH levels were restored close to normal levels and positive modulation of anti-oxidant enzyme activity was observed. Diabetic retinae showed significantly increased expression of Pro-inflammatory cytokines (TNF-α and IL-1ß) as compared to normal retinae. While Hsp-treated retinae showed significantly lower levels of cytokines as compared to diabetic retinae. Diabetic retinae showed increased caspase-3, GFAP and AQP4 expression. However, Hsp-treated retinae showed inhibitory effect on caspase-3, GFAP and AQP4 expression. LM images showed edematous Müller cell endfeet, and also degenerated photoreceptor layer; however, protective effect of Hsp was seen on Müller cell processes and photoreceptors. TEM study showed increased basement membrane (BM) thickness in diabetic retina, while relatively thin BM was recorded in Hsp-treated retina. It can be postulated that dietary flavanoids, like Hsp, can be effective for the prevention of diabetes induced neurovascular complications such as diabetic retinopathy.

Green tea prevents hyperglycemia-induced retinal oxidative stress and inflammation in streptozotocin-induced diabetic rats.

PURPOSE: Our objective was to investigate the effect of green tea (GT) on diabetes-induced retinal oxidative stress and proinflammatory parameters in rats. METHODS: Treatment (200 mg/kg body weight) was carried out for a period of 16 weeks in streptozotocin-induced diabetic rats and was evaluated for hypoglycemic, antioxidant [reduced glutathione (GSH), superoxide dismutase (SOD) and catalase (CAT)] and anti-inflammatory [tumor necrosis factor (TNF) α, vascular endothelial growth factor (VEGF)] activity. Histological changes were evaluated by transmission electron microscopy. RESULTS: Retinal GSH levels were 1.5-fold lower in diabetic rats as compared to normal rats (p < 0.05). However, in GT-treated rats, retinal GSH levels were restored close to those of the normal group. The antioxidant enzymes SOD and CAT showed a more than 2-fold decrease in activity in diabetic retinae as compared to normal retinae (p < 0.05). Both SOD and CAT enzymatic activities were restored close to normal in the GT-treated group. Expression of proinflammatory parameters (TNF-α and VEGF) was significantly inhibited in GT-treated retinae as compared to diabetic retinae (p < 0.05). Moreover, GT treatment prevented retinal capillary basement membrane thickness. CONCLUSION: The beneficial effects of GT suggest its potential role in the prevention and treatment of diabetic retinopathy in human subjects.

Hyperlipidemia and the development of diabetic retinopathy: Comparison between type 1 and type 2 animal models.

AIM: In the pathogenesis of diabetic retinopathy, reactive oxygen species (ROS) are elevated in the retina and the mitochondria are damaged, resulting in accelerated apoptosis. Dyslipidemia is also considered as one of the major factors in its development, and our aim is to investigate the compounding effect of hyperlipidemia in retinopathy. METHODS: Retinal ROS, mitochondrial damage and vascular pathology were investigated in Zucker diabetic fatty rats (ZDF, type 2 diabetes model), during the age that spans from hyperlipidemia/pre-hyperglycemia (6weeks), to severe hyperglycemia/moderate hyperlipidemia (~12weeks), and ultimately to severe hyperglycemia/hyperlipidemia (20-40weeks). For comparison, retina from streptozotocin-induced Wistar rats (type 1 diabetic for 10-40weeks) was analyzed. RESULTS: Compared to age-matched lean rats, despite increased retinal cytosolic ROS in 6-week-old ZDF rats, mitochondrial dysfunction and DNA damage were not detected, and in 12-week-old ZDF rats, retinal mitochondria were dysfunctional, but mtDNA damage and vascular pathology (cell apoptosis and degenerative capillaries) were not detectable. Retina from 20-week-old ZDF rats (hyperglycemic for 14weeks or less) had significant mitochondrial dysfunction, mtDNA damage and vascular pathology, and similar abnormalities were observed in 40-week-old ZDF rats. Although retinal mitochondrial dysfunction was observed in Wistar rats diabetic for 20weeks, mtDNA damage and vascular pathology were not detectable till the duration of diabetes was further extended. CONCLUSIONS: Hyperlipidemia, in a hyperglycemic milieu, potentiates mitochondrial damage and augments the development of retinopathy. Control of dyslipidemia in pre-diabetic patients may prevent/delay the development and the progression of this devastating disease.

Lipotoxicity augments glucotoxicity-induced mitochondrial damage in the development of diabetic retinopathy.

PURPOSE: Although hyperglycemia is the main instigator in the development of diabetic retinopathy, dyslipidemia is also considered to play an important role. In the pathogenesis of diabetic retinopathy, cytosolic NADPH oxidase 2 (Nox2) is activated before retinal mitochondria are damaged. Our aim was to investigate the effect of lipids in the development of diabetic retinopathy. METHODS: Reactive oxygen species (ROS, by 2',7'-dichlorofluorescein diacetate) and activities of Nox2 (by a lucigenin-based method) and Rac1 (by G-LISA) were quantified in retinal endothelial cells incubated with 50 µM palmitate in 5 mM glucose (lipotoxicity) or 20 mM glucose (glucolipotoxicity) for 6 to 96 hours. Mitochondrial DNA (mtDNA) damage was evaluated by extended-length PCR and its transcription by quantifying cytochrome b transcripts. RESULTS: Within 6 hours of exposure of endothelial cells to lipotoxicity, or glucotoxicity (20 mM glucose, without palmitate), significant increase in ROS, Nox2, and Rac1 was observed, which was exacerbated by glucolipotoxic insult. At 48 hours, neither lipotoxicity nor glucotoxicity had any effect on mtDNA and its transcription, but glucolipotoxicity significantly damaged mtDNA and decreased cytochrome b transcripts, and at 96 hours, glucotoxicity and glucolipotoxicity produced similar detrimental effects on mitochondrial damage. CONCLUSIONS: Although during initial exposure, lipotoxic or glucotoxic insult produces similar increase in ROS, addition of lipotoxicity in a glucotoxic environment further exacerbates ROS production, and also accelerates their damaging effects on mitochondrial homeostasis. Thus, modulation of Nox2 by pharmacological agents in prediabetic patients with dyslipidemia could retard the development of retinopathy before their hyperglycemia is observable.

Oxidative stress and epigenetic modifications in the pathogenesis of diabetic retinopathy.

Diabetic retinopathy remains the major cause of blindness among working age adults. Although a number of metabolic abnormalities have been associated with its development, due to complex nature of this multi-factorial disease, a link between any specific abnormality and diabetic retinopathy remains largely speculative. Diabetes increases oxidative stress in the retina and its capillary cells, and overwhelming evidence suggests a bidirectional relationship between oxidative stress and other major metabolic abnormalities implicated in the development of diabetic retinopathy. Due to increased production of cytosolic reactive oxygen species, mitochondrial membranes are damaged and their membrane potentials are impaired, and complex III of the electron transport system is compromised. Suboptimal enzymatic and nonenzymatic antioxidant defense system further aids in the accumulation of free radicals. As the duration of the disease progresses, mitochondrial DNA (mtDNA) is damaged and the DNA repair system is compromised, and due to impaired transcription of mtDNA-encoded proteins, the integrity of the electron transport system is encumbered. Due to decreased mtDNA biogenesis and impaired transcription, superoxide accumulation is further increased, and the vicious cycle of free radicals continues to self-propagate. Diabetic milieu also alters enzymes responsible for DNA and histone modifications, and various genes important for mitochondrial homeostasis, including mitochondrial biosynthesis, damage and antioxidant defense, undergo epigenetic modifications. Although antioxidant administration in animal models has yielded encouraging results in preventing diabetic retinopathy, controlled longitudinal human studies remain to be conducted. Furthermore, the role of epigenetic in mitochondrial homeostasis suggests that regulation of such modifications also has potential to inhibit/retard the development of diabetic retinopathy.

Retinoprotective effects of Moringa oleifera via antioxidant, anti-inflammatory, and anti-angiogenic mechanisms in streptozotocin-induced diabetic rats.

PURPOSE: The present study was aimed to evaluate the retinoprotective effects of Moringa oleifera (MO) in Streptozotocin-induced diabetic rats. METHODS: The study was continued for 24 weeks and evaluated for inflammatory (tumor necrosis factor [TNF]-α and interleukin [IL]-1ß, angiogenic (vascular endothelial growth factor [VEGF] and protein kinase C [PKC]-ß) and antioxidant (Glutathione, Superoxide dismutase, and Catalase) parameters. Retinal leakage was checked by Fluorescein angiography (FA) and fundus photographs were evaluated for retinal vessel caliber (arteriolar and venular). Transmission electron microscopy was done to determine basement membrane (BM) thickness. RESULTS: The results of the present study showed potential hypoglycemic and retinal antioxidant effects of MO. In the present study, a significant rise in the expression of retinal inflammatory (TNF-α and IL-1ß) and angiogenic (VEGF and PKC-ß) parameters was observed in diabetic retinae as compared to normal retinae. However, MO-treated retinae showed marked inhibition in the expression of inflammatory and angiogenic parameters. Further, in the present study, diabetic retinae showed dilated retinal vessels as compared to normal. However, MO-treated retinae showed marked prevention in the dilatation of retinal vessels. Fluorescein angiograms obtained from diabetic retinae showed leaky and diffused retinal vasculature. On the other hand, MO-treated retinae showed intact retinal vasculature. Further, results of the transmission electron microscopy study showed thickened capillary BM in the diabetic retina as compared to normal retinae. However, treatment with MO prevented thickening of capillary BM. CONCLUSION: Our result suggests that MO may be useful in preventing diabetes induced retinal dysfunction.

Hesperetin ameliorates hyperglycemia induced retinal vasculopathy via anti-angiogenic effects in experimental diabetic rats.

The purpose of the study was to evaluate vasculoprotective effects of Hesperetin (Hsp) in Streptozotocin induced diabetic rats. The study was carried out for a period of 24weeks and evaluated for angiogenic parameters (VEGF and PKC-ß), retinal vascular leakage by fluorescein angiography and, vessel (arteriolar and venular) diameters and any morphological abnormality through fundus photographs. Apart from this, transmission electron microscopy (TEM) was done to determine capillary basement membrane (BM) thickness. The results of the present study showed a significant increase in the expression of VEGF and PKC-ß in diabetic retinae as compared to normal retinae. On the other hand, Hsp-treated retinae showed marked inhibition in the expression of VEGF and PKC-ß. In the present study, diabetic retinae showed increase vascular permeability and leakage as compared to normal retinae. However, Hsp-treated retinae have not shown any such vascular dysfunctions. Moreover, there was significant increase in vessel caliber recorded in diabetic retinae compared to normal retinae, on the contrary Hsp-treated retinae showed lesser dilated vessels. Further, TEM study showed thickened BM in diabetic group as compared to normal group. However, Hsp-treated retinae showed marked prevention in BM thickness. In conclusion, it can be sated that Hsp has potential vasoprotective effects and can be useful in preventing diabetes induced vasculopathy.

Curcumin prevents experimental diabetic retinopathy in rats through its hypoglycemic, antioxidant, and anti-inflammatory mechanisms.

PURPOSE: The purpose of this study was to evaluate the therapeutic potential of oral curcumin (1 g/kg body weight of rat) in the prevention and treatment of streptozotocin-induced diabetic retinopathy in Wistar albino rats. METHODS: The treatment was carried out for a period of 16 weeks in diabetic rats and evaluated for hyperglycemic, antioxidant (superoxide dismutase, catalase, and glutathione), and inflammatory parameters (tumor necrosis factor-α, vascular endothelial growth factor). Rat fundus was observed weekly to see any visible changes in the retina, such as tortuosity and dilation of retinal vessels. Histological changes were evaluated by transmission electron microscopy. RESULTS: Treatment with curcumin showed significant hypoglycemic activity compared with the diabetic group. Retinal glutathione levels were decreased by 1.5-fold, and antioxidant enzymes, superoxide dismutase and catalase, showed >2-fold decrease in activity in the diabetic group; on the other hand, curcumin positively modulated the antioxidant system. Proinflammatory cytokines, tumor necrosis factor-α and vascular endothelial growth factor, were elevated >2-fold in the diabetic retinae, but prevented by curcumin. Transmission electron microscopy showed degeneration of endothelial cell organelles and increase in capillary basement membrane thickness in diabetic retina, but curcumin prevented the structural degeneration and increase in capillary basement membrane thickness in the diabetic rat retinae. CONCLUSION: Based on the above results, it may be concluded that curcumin may have potential benefits in the prevention of retinopathy in diabetic patients.