The aim of this study was to investigate the protective effects and mechanisms of hesperidin, a plant based active flavanone found in citrus fruits, under the oxidative stress and apoptosis induced by high levels of glucose in retinal ganglial cells (RGCs). RGC-5 cells were pretreated with hesperidin (12.5, 25, or 50 μmol/L) for 6 h followed by exposure to high (33.3 mmol/L) d-glucose for 48 h. The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay was adopted to evaluate cell viability. Mitochondrial function was estimated by measuring the mitochondrial membrane potential (ΔΨm). A fluorescent probe was employed to evaluate the intercellular production of reactive oxygen species (ROS). Colorimetric assay kits were used to evaluate lipid peroxidation, antioxidant enzyme activities, and protein carbonyls formation. The expression of apoptosis-related proteins and mitogen-activated protein kinase (MAPK) were measured with Western blotting. Hesperidin inhibited high glucose-mediated cell loss and restored mitochondrial function including a reversion of ΔΨm loss and cytochrome c release. Treated with hesperidin, high glucose-induced increase in ROS, malondialdehyde, and protein carbonyl levels were blocked in RGC-5 cells. Hesperidin was found to elevate the activities of superoxide dismutase, catalase, glutathione peroxidase, and to recover glutathione levels. Hesperidin inhibited high glucose-induced cell apoptosis by attenuating the downregulation of caspase-9, caspase-3, and Bax/Bcl-2. Furthermore, the phosphorylation of c-Jun N-terminal kinases (JNK) and p38 MAPK triggered by high glucose were attenuated in RGC-5 cells after their incubation with hesperdin. We concluded that hesperidin may protect RGC-5 cells from high glucose-induced injury since it owns the properties of antioxidant action and blocks mitochondria-mediated apoptosis.

Plantaginis semen, the dried ripe seed of Plantago asiatica L. or Plantago depressa Willd. (Plantaginaceae), has been traditionally used to treat blurred vision in Asia. The aim of this work was to investigate the effect of plantaginis semen ethanol extract (PSEE) on the amelioration of diabetic retinopathy (DR) in streptozotocin (STZ)-diabetic rats. PSEE has abundant polyphenols with strong antioxidant activity. PSEE (100, 200 or 300 mg/kg) was oral administrated to the diabetic rats once daily consecutively for 8 weeks. Oral administration of PSEE resulted in significant reduction of hyperglycemia, the diameter of the retinal vessels, and retinal vascular permeability and leukostasis in diabetic rats. In addition, PSEE administration increased the activities of superoxidase dismutase (SOD) and catalase (CAT), and glutathione peroxidase (GSH) level in diabetic retinae. PSEE treatment inhibited the expression of vascular endothelial growth factor (VEGF) and hypoxia-inducible factor-1α (HIF-1α) and the phosphorylation of Akt without altering the Akt protein expression in diabetic retinae. PSEE not only down-regulated the gene expression of hypoxia-inducible factor-1α (TNF-α) and interleukin-1β (IL-1β), but also reduced ICAM-1 and VCAM-1 expression in diabetic retinae. Moreover, PSEE reduced the nuclear factor-κB (NF-κB) activation and corrected imbalance between histone deacetylases (HDAC) and histone acetyltransferases (HAT) activities in diabetic retinae. In conclusion, phenolic antioxidants extract from plantaginis semen has potential benefits in the prevention and/or progression of DR.

Gigantol is a bibenzyl compound derived from several medicinal orchids. This biologically active compound has shown promising therapeutic potential against diabetic cataracts, but whether this compound exerts beneficial effects on the other diabetic microvascular complications remains unclear. This study was carried out to examine effects of gigantol on high glucose-induced renal cell injury in cultured mouse kidney mesangial cells (MES-13). MES-13 cells were pretreated with gigantol (1, 5, 10 or 20 μmol/L) for 1 h followed by further exposure to high (33.3 mmol/L) glucose for 48 h. Gigantol concentration dependently enhanced cell viability followed by high glucose treatment in MES-13 cells. High glucose induced reactive oxygen species (ROS) generation, malondialdehyde production and glutathione deficiency were recoved in MES-13 cells pretreated with gigantol. High glucose triggered cell apoptosis via the the loss of mitochondrial membrane potential, depletion of adenosine triphosphate, upregulation of caspases 9 and 3, enhancement of cytochrome c release, and subsequent interruption of the Bax/Bcl-2 balance. These detrimental effects were ameliorated by gigantol. High glucose also induced activation of JNK, p38 mitogen-activated protein kinase (MAPK) and nuclear factor-κB (NF-κB) in MES-13 cells, which were blocked by gigantol. The results suggest that treatment MES-13 cells with gigantol halts high glucose-induced renal dysfunction through the suppression of the ROS/MAPK/NF-κB signaling pathways. Our data are of value to the understanding the mechanism for gigantol, and would benefit the study of drug development or food supplement for diabetes and nephropathy.

We investigate diosmin for its effect on the ARPE-19 human retinal pigment epithelial cells exposed to high glucose, a model of diabetic retinopathy (DR). After incubation for 4 days with a normal (5 mmol/L) concentration of D-glucose, ARPE-19 cells were exposed separately to normal or high concentrations of D-glucose (30 mmol/L) with or without diosmin at different concentrations (0.1, 1, 10 μg/mL) for another 48 h. Next, we assessed cell viability, reactive oxygen species (ROS) generation and antioxidant enzyme activities. In order to examine the underlying molecular mechanisms, we meanwhile analyzed the expressions of Bax, Bcl-2, total and phosphorylated JNK and p38 mitogen-activated protein kinase (MAPK). Diosmin dose dependently enhanced cell viability following high glucose treatment in ARPE-19 cells. The activities of superoxide dismutase and glutathione peroxidase, as well as the levels of reduced glutathione were decreased, while it was observed that levels of ROS in high glucose cultured ARPE-19 cells increased. High glucose also disturbed Bax and Bcl-2 expression, interrupted Bcl-2/Bax balance, and triggered subsequent cytochrome c release into the cytosol and activation of caspase-3. These detrimental effects were ameliorated dose dependently by diosmin. Furthermore, diosmin could abrogate high glucose-induced apoptosis as well as JNK and P38 MAPK phosphorylation in ARPE-19 cells. Our results suggest that treatment ARPE-19 cells with diosmin halts hyperglycemia-mediated oxidative damage and thus this compound may be a candidate for preventing the visual impairment caused by DR.

The present study investigates the amelioration of diabetic retinopathy (DR) by Zingiber zerumbet rhizome ethanol extracts (ZZRext) in streptozotocin-induced diabetic rats (STZ-diabetic rats). ZZRext contains high phenolic and flavonoid contents. STZ-diabetic rats were treated orally with ZZRext (200, 300 mg/kg per day) for three months. Blood-retinal barrier (BRB) breakdown and increased vascular permeability were found in diabetic rats, with downregulation of occludin, and claudin-5. ZZRext treatment effectively preserved the expression of occludin, and claudin-5, leading to less BRB breakdown and less vascular permeability. Retinal histopathological observation showed that the disarrangement and reduction in thickness of retinal layers were reversed in ZZRext-treated diabetic rats. Retinal gene expression of tumor necrosis factor-α, interleukin (IL)-1β, IL-6, vascular endothelial growth factor, intercellular adhesion molecule-1 and vascular cell adhesion molecule-1 were all decreased in ZZRext-treated diabetic rats. Moreover, ZZRext treatment not only inhibited the nuclear factor κB (NF-κB) activation, but also downregulated the protein expression of p38 mitogen-activated protein kinase (MAPK) in diabetic retina. In conclusion, the results suggest that the retinal protective effects of ZZRext occur through improved retinal structural change and inhibiting retinal inflammation. The antiretinopathy property of ZZRext might be related to the downregulation of p38 MAPK and NF-κB signal transduction induced by diabetes.

The excessive activation of glutamate in the brain is a factor in the development of vascular dementia. γ-Oryzanol is a natural compound that has been shown to enhance brain function, but more research is needed to determine its potential as a treatment for vascular dementia. This study investigated if γ-oryzanol can delay or improve glutamate neurotoxicity in an in vitro model of differentiated HT-22 cells and explored its neuroprotective mechanisms. The differentiated HT-22 cells were treated with 0.1 mmol/L glutamate for 24 h then given γ-oryzanol at appropriate concentrations or memantine (10 µmol/L) for another 24 h. Glutamate produced reactive oxygen species and depleted glutathione in the cells, which reduced their viability. Mitochondrial dysfunction was also observed, including the inhibition of mitochondrial respiratory chain complex I activity, the collapse of mitochondrial transmembrane potential, and the reduction of intracellular ATP levels in the HT-22 cells. Calcium influx triggered by glutamate subsequently activated type II calcium/calmodulin-dependent protein kinase (CaMKII) in the HT-22 cells. The activation of CaMKII-ASK1-JNK MAP kinase cascade, decreased Bcl-2/Bax ratio, and increased Apaf-1-dependent caspase-9 activation were also observed due to glutamate induction, which were associated with increased DNA fragmentation. These events were attenuated when the cells were treated with γ-oryzanol (0.4 mmol/L) or the N-methyl-D-aspartate receptor antagonist memantine. The results suggest that γ-oryzanol has potent neuroprotective properties against glutamate excitotoxicity in differentiated HT-22 cells. Therefore, γ-oryzanol could be a promising candidate for the development of therapies for glutamate excitotoxicity-associated neurodegenerative diseases, including vascular dementia.