Gökhan Sadi

Effect of vitamin C and lipoic acid on streptozotocin-induced diabetes gene expression: mRNA and protein expressions of Cu–Zn SOD and catalase

The involvement of oxidative stress in the pathogenesis of diabetes mellitus has been confirmed by numerous studies. In this study, the expression of two antioxidant enzymes, superoxide dismutase (SOD), and catalase which are involved in the detoxification of reactive oxygen species was studied in the streptozotocin-induced diabetic rat liver tissues. The enzyme assays showed a significant decrease in both enzymes activities compared to control animals. The RT-PCR and Western-blot analysis results demonstrated that this decrease in activity is regulated at the level of gene expression, as both catalase and Cu–Zn SOD mRNA and protein expressions were also suppressed. Supplementing the animals with vitamin C, a powerful antioxidant increased both SOD and catalase activities with no change in both mRNA and protein expressions suggesting a role of post-translational modification. However, even though mRNA expressions of both catalase and Cu–Zn SOD were not changed, the protein levels increased in parallel to activities in the case of another antioxidant, α-lipoic acid. An increase in the rate of translation, without changing the rate of transcription indicates a translational effect of lipoic acid in changing the activities of antioxidant enzymes to prevent the oxidative damage in diabetes.

Gene expressions of Mn-SOD and GPx-1 in streptozotocin-induced diabetes: effect of antioxidants

Increased oxidative stress and impaired antioxidant defense mechanisms are believed to be the important factors contributing to the pathogenesis and progression of diabetes mellitus. In this study, we have reported the effects of the streptozotocin-induced diabetes on the gene expression and the activities of two antioxidant enzymes, manganese superoxide dismutase (MnSOD) and glutathione peroxidase (GPx). We also studied the effects of two antioxidants, vitamin C and DL-α-lipoic acid (LA), on the system. Our results showed no significant change in both enzymes activities in diabetic animals compared to controls. Similarly, mRNA and protein profiles of MnSOD showed no change. Though the mRNA expression of GPx did not show any change, Western-blot analysis results demonstrated that protein expression is increased. LA, which is a water- and lipid-soluble antioxidant, decreased the protein expression of MnSOD, though mRNA levels and activities remained unchanged. LA treatment increased the GPx activities in diabetic tissues, significantly, and RT-PCR and Western-blot analysis results demonstrated that this increase in activity is not regulated at the gene level, as both mRNA and protein levels did not change. Supplementing the animals with vitamin C, a powerful water-soluble antioxidant, increased the mRNA expression of MnSOD, though the protein expression and the activity did not change statistically. On the other hand GPx activity increased significantly through post-translational modifications, as both mRNA and protein expressions did not change. These results together with our previous findings about the gene expressions of catalase and Cu–Zn SOD indicate the presence of very intricate control mechanisms regulating the activities of antioxidant enzymes in order to prevent the damaging effects of oxidative stress.

Redox regulation of antioxidant enzymes: post-translational modulation of catalase and glutathione peroxidase activity by resveratrol in diabetic rat liver

Resveratrol is a strong antioxidant that exhibits blood glucose-lowering effects, which might contribute to its usefulness in preventing complications associated with diabetes. The present study aimed to investigate resveratrol effects on catalase (CAT) and glutathione peroxidase (GPx) gene and protein expression, their phosphorylation states and activities in rat liver of STZ-induced diabetes. Diabetes increased the levels of total protein phosphorylation and p-CAT, while mRNA expression, protein levels, and activity were reduced. Although diabetes induced transcriptional repression over GPx, it did not affect the protein levels and activity. When resveratrol was administered to diabetic rats, an increase in activity was associated with an increase in p-GPx levels. Decrease in Sirtuin1 (SIRT1) and nuclear factor erythroid 2-related factor (Nrf2) and increase in nuclear factor kappa B (NFκB) gene expression in diabetes were associated with a decrease in CAT and GPx mRNA expression. A possible compensatory mechanism for reduced gene expression of antioxidant enzymes is proved to be nuclear translocation of redox-sensitive Nrf2 and NFκB in diabetes which is confirmed by the increase in nuclear and decrease in cytoplasmic protein levels of Nrf2 and NFκB. Taken together, these findings revealed that an increase in the oxidized state in diabetes intricately modified the cellular phosphorylation status and regulation of antioxidant enzymes. Gene regulation of antioxidant enzymes was accompanied by nuclear translocation of Nrf2 and NFκB. Resveratrol administration also activated a coordinated cytoprotective response against diabetes-induced changes in liver tissues.

Resveratrol prevents high-fructose corn syrup-induced vascular insulin resistance and dysfunction in rats

Dietary intake of fructose and sucrose can cause development of metabolic and cardiovascular disorders. The consequences of high-fructose corn syrup (HFCS), a commonly consumed form of fructose and glucose, have poorly been examined. Therefore, in this study, we investigated whether HFCS intake (10% and 20% beverages for 12 weeks) impacts vascular reactivity to insulin and endothelin-1 in conjunction with insulin receptor substrate-1(IRS-1), endothelial nitric oxide synthase (eNOS) and inducible NOS (iNOS) mRNA/proteins levels in aorta of rats. At challenge, we tested the effectiveness of resveratrol (28-30 mg/kg body weight/day) on outcomes of HFCS feeding. HFCS (20%) diet feeding increased plasma triglyceride, VLDL, cholesterol, insulin and glucose levels, but not body weights of rats. Impaired nitric oxide-mediated relaxation to insulin (10⁻⁹ to 3×10⁻⁶ M), and enhanced contraction to endothelin-1 (10⁻¹¹ to 10⁻⁸ M) were associated with decreased expression of IRS-1 and eNOS mRNA and protein, but increased expression of iNOS, in aortas of rats fed with HFCS. Resveratrol supplementation restored many features of HFCS-induced disturbances, probably by regulating eNOS and iNOS production. In conclusion, dietary HFCS causes vascular insulin resistance and endothelial dysfunction through attenuating IRS-1 and eNOS expressions as well as increasing iNOS in rats. Resveratrol has capability to recover HFCS-induced disturbances.

Changes in expression profiles of antioxidant enzymes in diabetic rat kidneys

In diabetes mellitus, increased formation of reactive oxygen species due to high level of glucose in both blood plasma and tissues creates oxidative stress and damages the tissues. Antioxidants together with the antioxidant enzymes are very important in order to protect the cells against oxidative damage.

Resveratrol improves hepatic insulin signaling and reduces the inflammatory response in streptozotocin-induced diabetes

Diabetes mellitus is a heterogeneous metabolic disorder essentially characterized by deficiency of insulin secretion, insulin receptor or post-receptor events. This study aims to investigate the effects of resveratrol administration on the metabolic characteristics, hepatic functions, histopathological features and insulin signaling pathway components in streptozotocin induced diabetes. Male Wistar rats were randomly divided into four groups: (1) control/vehicle; (2) control/20mg/kg resveratrol; (3) diabetic/vehicle; and (4) diabetic/20mg/kg resveratrol. Histopathological examinations were carried out to reveal hepatic tissue damage and inflammation. In addition to hepatic glucose, lipid, insulin, ALT, AST, resistin and XOD contents, gene and protein expressions of insulin signaling pathway components such as insulin Rβ, IRS-1, IRS-2, eNOS, PI3K, Akt, and FOXO3a were analyzed by qRT-PCR and Western blot. The rats in the diabetes group had significantly lower terminal body weight and hepatic insulin level, but significantly higher hepatic glucose, total cholesterol, triglyceride and resistin concentrations. Diabetes triggered the inflammatory process in the liver tissues that was evidenced by histopathological deformations and increase in the hepatic ALT and AST levels. Hepatic inflammation was considerably associated with insulin signaling pathway ever since a significant down-regulation of insulin signaling components; IRS-1, IRS-2, PI3K, Akt and mTOR have been identified in the diabetic group. To some extent, resveratrol treatment reversed the diabetes-induced changes in the liver tissues. Taken together, resveratrol partly improved hepatic dysfunction induced by diabetes. This may be due to the healing activity of resveratrol on insulin signaling pathway, resistin levels and hepatic glucose-lipid contents.

Differential Gene Expression in Liver Tissues of Streptozotocin-Induced Diabetic Rats in Response to Resveratrol Treatment

This study was conducted to elucidate the genome-wide gene expression profile in streptozotocin induced diabetic rat liver tissues in response to resveratrol treatment and to establish differentially expressed transcription regulation networks with microarray technology. In addition to measure the expression levels of several antioxidant and detoxification genes, real-time quantitative polymerase chain reaction (qRT-PCR) was also used to verify the microarray results. Moreover, gene and protein expressions as well as enzymatic activities of main antioxidant enzymes; superoxide dismutase (SOD-1 and SOD-2) and glutathione S-transferase (GST-Mu) were analyzed. Diabetes altered 273 genes significantly and 90 of which were categorized functionally which suggested that genes in cellular catalytic activities, oxidation-reduction reactions, co-enzyme binding and terpenoid biosynthesis were dominated by up-regulated expression in diabetes. Whereas; genes responsible from cellular carbohydrate metabolism, regulation of transcription, cell signal transduction, calcium independent cell-to-cell adhesion and lipid catabolism were down-regulated. Resveratrol increased the expression of 186 and decreased the expression of 494 genes in control groups. While cellular and extracellular components, positive regulation of biological processes, biological response to stress and biotic stimulants, and immune response genes were up-regulated, genes responsible from proteins present in nucleus and nucleolus were mainly down-regulated. The enzyme assays showed a significant decrease in diabetic SOD-1 and GST-Mu activities. The qRT-PCR and Western-blot results demonstrated that decrease in activity is regulated at gene expression level as both mRNA and protein expressions were also suppressed. Resveratrol treatment normalized the GST activities towards the control values reflecting a post-translational effect. As a conclusion, global gene expression in the liver tissues is affected by streptozotocin induced diabetes in several specific pathways. The present data suggest the presence of several processes which contribute and possibly interact to impair liver functions in type 1 diabetes, several of which are potentially amenable to therapeutic interventions with resveratrol.

Resveratrol regulates oxidative biomarkers and antioxidant enzymes in the brain of streptozotocin-induced diabetic rats.

Abstract Context: Oxidative stress has been implicated in the progression of pathogenesis in diabetes mellitus and leads to a variety of deformations in the central nervous system. Recent studies have provided several insights on therapeutic uses of resveratrol in diabetic complications. Objective: The present study determines if resveratrol ameliorates oxidative stress and molecular changes in the brain frontal cortex of streptozotocin-induced diabetic rats. Materials and methods: Rats were divided into four groups: control, diabetic, resveratrol-treated control, and resveratrol-treated diabetic. After diabetes induction, resveratrol (20 mg/kg) was given intraperitoneally once daily for 4 weeks. In addition to enzymatic activities, gene and protein expression of brain antioxidant enzymes were utilized by qRT-PCR and Western blot, respectively. Results: The results indicated a significant elevation in total oxidant species (1.22-fold) and malonedialdehyde (1.38-fold) contents in diabetic rat brain cortex tissues. In addition, significant augmentation in the activities of catalase (1.38-fold) and superoxide dismutase (3-fold) was witnessed with the gene and protein expression levels reflecting a transcriptional regulation. Resveratrol treatment significantly normalized diabetic malonedialdehyde and oxidized glutathione levels and strengthens the action of all antioxidant enzymes. Recovery of the diabetes-associated changes reflects the reduction of oxidative conditions by resveratrol and reveals the decrease in the requirement for the activation of antioxidant defense systems in the brain tissues of diabetic rats. Discussion and conclusion: Potent antioxidant and neuroprotective properties of resveratrol against diabetes-induced oxidative damage were demonstrated and the results support the conduct of new studies searching for the molecular mechanism of diabetes-induced changes in brain tissues.

Long-Term Dietary Fructose Causes Gender-Different Metabolic and Vascular Dysfunction in Rats: Modulatory Effects of Resveratrol

Background/Aims: There is limited knowledge on the gender differences in the effects of dietary fructose. In the current study, we investigated whether long-term fructose intake impacts metabolic parameters and vascular reactivity differently between male and female rats. Moreover, we tested whether resveratrol has a gender-specific effectiveness on the alterations. Methods: Male and female rats were divided into four groups as control; resveratrol; fructose and resveratrol plus fructose. Fructose was given to the rats as 10% solution in drinking water for 24 weeks. All rats were fed with the standard diet with or without resveratrol. Results: High-fructose diet increased plasma insulin, triglyceride and VLDL levels as well as omental weights in both genders. Long-term dietary fructose causes marked increase in body weight of males, but not females. Dietary fructose impaired endothelial relaxation to acetylcholine and intensified contraction to phenylephrine in the aortas of male and female rats, but differently it also reduced insulin-induced vasodilation in aortas of female rats. These changes were associated with decreased expression levels of endothelial nitric oxide synthase (eNOS) mRNA and protein, but increased in inducible NOS (iNOS), in aortas of male and female rats. Dietary fructose suppressed expression levels of sirtuin 1 (SIRT1) and insulin receptor substrate-2 (IRS-2) mRNA in aortas from female rats. Resveratrol supplementation efficiently restored fructose-induced metabolic and vascular dysfunction in both genders probably by regulating eNOS and iNOS production. Moreover, the augmentations in SIRT1 and IRS-2 mRNA in females and IRS-1 mRNA in males may possibly contribute to the beneficial effects of resveratrol as well. Conclusion: Long-term fructose intake may differently affect metabolic and vascular function between male and female rats, which are modified by resveratrol.

Dietary Fructose Activates Insulin Signaling and Inflammation in Adipose Tissue: Modulatory Role of Resveratrol

The effects of high-fructose diet on adipose tissue insulin signaling and inflammatory process have been poorly documented. In this study, we examined the influences of long-term fructose intake and resveratrol supplementation on the expression of genes involved in insulin signaling and the levels of inflammatory cytokines and sex hormones in the white adipose tissues of male and female rats. Consumption of high-fructose diet for 24 weeks increased the expression of genes involved in insulin signaling including IR, IRS-1, IRS-2, Akt, PI3K, eNOS, mTOR, and PPARγ, despite induction of proinflammatory markers, iNOS, TNFα, IL-1β, IL-18, MDA, and ALT, as well as anti-inflammatory factors, IL-10 and Nrf2 in adipose tissues from males and females. Total and free testosterone concentrations of adipose tissues were impaired in males but increased in females, although there were no changes in their blood levels. Resveratrol supplementation markedly restored the levels of MDA, IL6, IL-10, and IL-18, as well as iNOS, Nrf2, and PI3K mRNA, in adipose tissues of both genders. Dietary fructose activates both insulin signaling and inflammatory pathway in the adipose tissues of male and female rats proposing no correlation between the tissue insulin signaling and inflammation. Resveratrol has partly modulatory effects on fructose-induced changes.