Metformin improves IKCa-mediated endothelial dilative dysfunction of arteriole in diabetic rats

Abstract

Methods Diabetic rat model was induced by a single intraperitoneal injection of 30 mg/kg STZ after high fat and glucose diet for 8 weeks. Animals whose blood glucose > 11.1 mmol/L were included in diabetic and metformin group. Agematched animals fed with standard chow and injected with citric acid buffer were served as control. Four weeks after STZ injection, rats in three groups were fed with normal diet for additional 8 weeks. After that, fasting blood was drawn and third-order mesenteric arteries were separated. Hemoglobin A1c (HbA1c) was measured with an automatic analyzer. The changes of Achand NS309 (opener of IKCa and small conductance Ca -activated K channel, SKCa) -induced vasodilatation mediated by IKCa in mesentery arterioles of each group and mesentery arterioles of normal rats incubated with 200 μg/mL AGE-BSA (200 μg/mL BSA as control) for 3 hours were measured by multi-myograph system. The effect of metformin on AGEBSA (200 μg/mL) and H2O2 (100 μmol/L) induced changes of IKCa mRNA and protein expression in cultured human umbilical vein endothelial cells (HUVECs) were detected by RT-PCR and Western blot. The level of malondialdehyde (MDA) and the activity of Cu-Zn superoxide dismutase (Cu-Zn SOD) in cellular supernatant were determined by colorimetric method. Results Increased HbA1c level and reduceded endotheliumdependent dilative response mediated by IKCa in mesentery arterioles were observed in diabetic rats, and metformin treatment (300 mg/kg/day by gavage) restored the adverse condition. The vasodilatation mediated by IKCa was also impaired in 200 μg/mL AGE-BSA-incubated mesentery arterioles. AGE-BSA at 200 μg/mL concentration and H2O2 (100 μmol/L) significantly decreased the mRNA and protein expression of IKCa. AGE-BSA also increased the production of MDA and inhibited Cu-Zn SOD activity in HUVECs. Metformin of 10 mol/L and 10 mol/L reversed those effects.