B.P. Srinivasan

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-1β), 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-1β) 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.

Triple verses glimepiride plus metformin therapy on cardiovascular risk biomarkers and diabetic cardiomyopathy in insulin resistance type 2 diabetes mellitus rats

IRtype2DM patients are often treated with a combination of antidiabetic agents. Drugs with different complementary mechanisms of action frequently used in daily clinical practice but glycemic control with monotherapeutic attempts fail in the long run. To date, biomarkers for cardiovascular risk and insulin sensitivity with combination of triple oral hypoglycemic therapies are not fully revelled in view of additional cardiovascular risk reduction. In the present study, IRtype2DM induced by administering streptozotocin (90 mg/kg, i.p.) in neonatal rat model. IRtype2DM rats were selected by determining FPI [>60 pmol/l]; HOMA-IR & Hyperinsulinemic euglycemic clamp technique at 6 weeks and then treated for 8 weeks with (i) Metformin (120 mg/kg, o.d.)+Glimepiride (1mg/kg, o.d.), (ii) Metformin (265 mg/kg, o.d.)+Rosiglitazone (1mg/kg, o.d.)+Glimepiride (0.7 mg/kg, o.d.). At the end cardiovascular risk parameters evaluated by ELISA kits and insulin sensitivity were determined by HOMA-IR. In conclusion, triple oral hypoglycemic therapy improves glycemic control, insulin sensitivity, retards diabetic cardiomyopathy and does not increased body weight; decrease more detrimental inflammatory markers, increase interleukin-10 and adiponectin in neonatal streptozotocin-induced IRtype2DM Wistar Albino Rats. Triple therapy showed a synergistic effect and was promising in insulin resistance, better in additional cardiovascular risk reduction and those nonresponders to metformin add on glimepiride therapy.

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-β), 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-β 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-β. 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.

Sitagliptin, sitagliptin and metformin, or sitagliptin and amitriptyline attenuate streptozotocin-nicotinamide induced diabetic neuropathy in rats

Diabetic neuropathies are a family of nerve disorders caused by diabetes. Symptoms of the disease include nerve palsy, mononeuropathy, mononeuropathy multiplex, diabetic amyotrophy, painful polyneuropathy, autonomic neuropathy, and thoracoabdominal neuropathy. In this study, type 2 diabetes in rats was induced with nicotinamide-streptozotocin. Drug treatment was initiated on the d 15, with the combination regimen of metformin, pioglitazone and glimipiride or metformin and sitagliptin or sitagliptin, amitriptyline and sitagliptin and led to significantly improved glycemic control, increased grip strength and paw jumping response on d 21, 28 and 35 (P < 0.001). Significant increases in blood protein levels and decreases in urinary protein levels were observed in the animals treated with the different regimens on d 21, 28 and 35 (P < 0.001). Combined treatment of streptozotocin and nicotinamide caused marked degeneration of nerve cells, while administration of metformin and sitagliptin showed tissue regeneration and no body weight gain. In conclusion, treatment with sitagliptin and sitagliptin combined with metformin or amitriptyline results in no body weight gain, but causes an increase in grip strength and pain sensitivity, exhibits neural protection, and reverses the alteration of biochemical parameters in rats with streptozotocin-nicotinamide induced type 2 diabetes.

Novel diallyldisulfide analogs ameliorate cardiovascular remodeling in rats with L-NAME-induced hypertension.

Diallyldisulfide (DADS), an active principle of garlic (Allium sativum) is known for its antihypertensive properties. The present study was designed to evaluate the effect of novel DADS analogs, against L-NAME induced hypertension in Wistar rats. The daily administration of L-NAME (50mg/kg) for six weeks along with DADS analogs (20 mg/kg) significantly decreased the elevated systolic blood pressure (SBP) and the activity of angiotensin converting enzyme (ACE) and also inhibited the decline in nitrite/nitrate (NO(x)) concentrations and cyclic guanosine monophosphate (cGMP) levels. Adverse changes such as lipid peroxidation, protein damage and a decrease in the levels of antioxidant enzymes, were rectified after the administration of DADS analogs. Oral administration of DADS analogs preserved the expression of endothelial nitric oxide synthase (eNOS). The ability of the DADS analogs to inhibit L-NAME induced hypertension was compared with Enalapril (15 mg/kg), which was taken as a standard. The DADS analogs prevented L-NAME-induced cardio toxicity, which was also reflected at the microscopic level indicative of its cardio protective effects. DADS analogs induced vasorelaxation was completely abolished by the removal of the endothelium or by pre-treatment with L-NAME, an inhibitor of nitric oxide synthase. DADS analogs inhibited the calcium influx induced by phenylephrine (0.3 μM) and high K(+) (60mM) and this effect was completely abolished by pretreatment of L-NAME. Taken together, our results show that the DADS analogs induce vasorelaxation and have antihypertensive properties, which may be mediated through activation of eNOS.

Vildagliptin selectively ameliorates GLP-1, GLUT4, SREBP-1c mRNA levels and stimulates β-cell proliferation resulting in improved glucose homeostasis in rats with streptozotocin-induced diabetes.

OBJECTIVE Inhibitors of dipeptidyl peptidase-4 (DPP-4), a key regulator of the actions of incretin hormones, exert antihyperglycemic effects in type 2 diabetic patients. A major unanswered question concerns the potential ability of DPP-4 inhibition to have beneficial disease-modifying effects, specifically to attenuate loss of pancreatic β-cell mass and function due to oxidative stress induced inflammation. Here, we investigated the effects of a potent and selective DPP-4 inhibitor vildagliptin on glycemic control, pancreatic β-cell mass and function, genes and proteins expressions, tumor necrosis factor-alpha, and nitric oxide in an n2-STZ diabetic model of rat with defects in insulin sensitivity and secretion. METHOD To induce NIDDM, STZ (sigma chemicals, USA) (90 mg/kg) was administered i.p. to a group of 2 days old pups. Another group of pups received only saline. The pups were weaned for 21 days, and 6 weeks after the injection of STZ, the animals were checked for fasting glucose level (FPG) ≥160 mg/dl were considered as diabetic. RESULTS Significant and dose-dependent correction of postprandial and fasting hyperglycemia was observed in diabetic rats following 8 weeks of chronic therapy. Treatment with vildagliptin showed increase in the number of insulin-positive β-cells in islets and improved the expressions of genes and proteins are responsible for insulin secretions. In addition, treatment of rats with vildagliptin significantly increased insulin content; and decreased the nitric oxide and TNF-alpha concentration. CONCLUSION These findings suggest that DPP-4 inhibitors may offer long-lasting efficacy in the treatment of diabetes mellitus by modifying the courses of the disease.

A novel long acting DPP-IV inhibitor PKF-275-055 stimulates β-cell proliferation resulting in improved glucose homeostasis in diabetic rats

The enzyme dipeptidyl peptidase-IV (DPP-4) inactivates the incretin hormone glucagon-like peptide-1 (GLP-1). GLP-1 has therapeutic effects in patients with type 2 diabetes, but its potential is limited by a short half-life, DPP-4 inhibition is a promising approach to diabetes treatment. This study examined chronic (once-a-day dosing for 8 weeks) effects of the DPP-4 inhibitor PKF-275-055 (1, 3, and 10mg/kg) on β-cell regeneration and plasma DPP-IV activity, intact GLP-1, glucose, and insulin after an oral glucose load in neonatal wistar rats injected with streptozotocin (STZ) (n2-STZ model), a recognized model of type 2 diabetes. In streptozotocin induced diabetic rats, PKF-275-055 (3, and 10mg/kg) significantly reduced glucose excursion during the oral glucose tolerance test conducted 2h and 10h after administration, with increases in plasma insulin and active glucagon-like peptide-1 (GLP-1) levels and significantly inhibited (> 50% inhibition) plasma DPP-IV activity during both the 1st and 2nd OGTT in diabetic rats. In contrast, PKF-275-055 (1-10mg/kg) did not cause hypoglycemia in fasted normal rats. Furthermore, PKF-275-055 significantly inhibited advance glycation end product (HbA1c), HOMA-Index, gastric emptying and small intestinal transit rates, with significance at doses of 1mg/kg or higher. Immunological staining showed PKF-275-055 stimulates β-cell regeneration and reduces pancreatic cell apoptosis in diabetic treated rats. The present preclinical studies indicate that PKF-275-055 is a novel selective DPP-IV inhibitor with long-acting antidiabetic effect that might be a potential agent for type 2 diabetes.

Chronic DPP-IV inhibition with PKF-275-055 attenuates inflammation and improves gene expressions responsible for insulin secretion in streptozotocin induced diabetic rats.

Inhibitors of dipeptidyl peptidase-4 (DPP-IV), a key regulator of the actions of incretin hormones, exert antihyperglycemic effects in type 2 diabetic patients. A major question concerns the potential ability of long term DPP-IV inhibition to have beneficial disease-modifying effects, specifically to attenuate loss of pancreatic β-cell mass due to oxidative stress induced inflammation. Here, we investigated the effects of a potent and selective DPP-4 inhibitor, an analog of vildagliptin (PKF-275-055), on glycemic control, pancreatic β-cell mass, genes and proteins expressions, tumor necrosis factor-alpha, and nitric oxide in an n2-STZ diabetic model of rat with defects in insulin sensitivity and secretion. To induce NIDDM, streptozotocin (STZ) 90 mg/kg was administered i.p. to a group of 2 days old pups. Diabetic rats were administered orally with vildagliptin analog PKF-275-055. Saline treated animals served as diabetic control. Significant and dose-dependent correction of postprandial hyperglycemia was observed in diabetic rats following 8 weeks of chronic therapy. Treatment with PKF-275-055 showed increased the number of insulin-positive β-cells in islets and improved the expressions of genes and proteins are responsible for insulin secretions. In addition, treatment of rats with PKF-275-055 significantly increased insulin content, glycogen content and total proteins content; and decreased the inflammatory markers i.e. nitric oxide and TNF-alpha. The present studies indicate that PKF-275-055 is a novel selective DPP-IV inhibitor having potential to reduce inflammation that might be a potential agent for type 2 diabetes.

Aliskiren improves insulin resistance and ameliorates diabetic renal vascular complications in STZ-induced diabetic rats.

Aliskiren, a direct renin inhibitor (DRI), has therapeutic effects in patients with hypertension and associated complications, but its potential mechanism in diabetic nephropathy is lacking. The effects of aliskiren in Streptozotocin (STZ)-induced renal complication in diabetic rats were investigated. Aliskiren treatment for eight weeks at the dose of 10 mg/kg/day, via osmotic mini-pump, induced improvement in blood glucose levels, systolic blood pressure (BP) and serum creatinine. Improvement of insulin resistance by aliskiren was confirmed by increased glucose translocation in liver and muscle and hence insulin levels. The treated group also showed improvement in glomerulosclerosis and tubulointerstitial injury. Aliskiren treatment also improved albumin levels in plasma, suppressed profibrotic and proinflammatory cytokine synthesis viz TNF-α and TGF-β and angiogenesis by a decrease in VEGF. In addition, the level of total proteins and GFR via cystatin c and beta-2microglobulin along with adiponectin and erythropoietin were also improved. These results suggest that the beneficial organ protective effect of aliskiren is mediated by improvement in insulin resistance as well as a direct anti-fibrotic effect in the target organ in STZ-induced diabetic rats with a slight effect on blood pressure. Aliskiren may be a useful therapeutic agent in the treatment of type 2 diabetes and diabetic nephropathy.

Aqueous extract of tamarind seeds selectively increases glucose transporter-2, glucose transporter-4, and islets' intracellular calcium levels and stimulates β-cell proliferation resulting in improved glucose homeostasis in rats with streptozotocin-induced diabetes mellitus

Tamarindus indica Linn. has been in use for a long time in Asian food and traditional medicine for different diseases including diabetes and obesity. However, the molecular mechanisms of these effects have not been fully understood. In view of the multidimensional activity of tamarind seeds due to their having high levels of polyphenols and flavonoids, we hypothesized that the insulin mimetic effect of aqueous tamarind seed extract (TSE) might increase glucose uptake through improvement in the expression of genes of the glucose transporter (GLUT) family and sterol regulatory element-binding proteins (SREBP) 1c messenger RNA (mRNA) in the liver. Daily oral administration of TSE to streptozotocin (STZ)-induced (90 mg/kg intraperitoneally) type 2 diabetic male Wistar rats at different doses (120 and 240 mg/kg body weight) for 4 weeks showed positive correlation with intracellular calcium and insulin release in isolated islets of Langerhans. Tamarind seed extract supplementation significantly improved the GLUT-2 protein and SREBP-1c mRNA expression in the liver and GLUT-4 protein and mRNA expression in the skeletal muscles of diabetic rats. The elevated levels of serum nitric oxide (NO), glycosylated hemoglobin level (hemoglobin (A1c)) and tumor necrosis factor α (TNF-α) decreased after TSE administration. Immunohistochemical findings revealed that TSE abrogated STZ-induced apoptosis and increased β-cell neogenesis, indicating its effect on islets and β-cell mass. In conclusion, it was found that the antidiabetic effect of TSE on STZ-induced diabetes resulted from complex mechanisms of β-cell neogenesis, calcium handling, GLUT-2, GLUT-4, and SREBP-1c. These findings show the scope for formulating a new herbal drug for diabetes therapy.