| Objective:Diabetic nephropathy is one of the common chronic complications of diabetes,which is the one of the leading cause of end-stage renal disease. The recent researches have found that oxidative stress and its mediated apoptosis in kidneys participated the development of diabetic nephropathy. Not only mitochondria constitute the main source of intracellular reactive oxygen series, but also are the central regulator of apoptosis.As a powerful antioxidant, a-lipoic acid(a-LA)plays an important role in renal protection, but the mechanism remains unknown.In this study, we investigate the function changes of mitochondria on kidneys of diabetic rats and renal cell apoptosis mechanism induced by mitochondria, under oxidative stress resulted from high glucose.Meanwhile,we elucidate the protecting effects of a-lipoic acid on kidneys of diabetic rats and explore the related mechanism, which provide new theoretical bases for clinical treatments of diabetic nephropathy.Methods:The diabetic model was induced by intraperitoneal injection of streptozotocin(STZ) on Male SD, and then the diabetic rats were randomly divided into two groups:untreated-diabetic group (DM group), a-LA treated-diabetic group(a-LA group), and the normal rats were served as a control group (NC group).At 8 weeks after the onset of diabetes,Blood glucose(BG),Blood Urea Nitrogen(BUN),Serum Creatinine(SCr) and urinary albumin excrection rate (UAER) were detected,histomorphologic changes were observed by optics microscope and morphological changes of mitochondria were observed by electron microscope. The levels of malondiadehyde (MDA) and the activities of superoxide dismutase(SOD), in serum and renal cortex,were also evaluated.Additionally, the activities of succinate dehydrogenase(SDH) and cytochrome C oxidase of renal mitochondrial respiration chain, kidney mitochondrial membrane potential, mitochondrial swelling were measured among the different groups. Apoptosis-related protein cytochrome C (CytC) were tested by Western blotting, while(the second mitochondria-derived activat or of caspase/direct IAP binding protein, Smac/DIABLO)and apoptosis-inducing factor (AIF) were detected by Immunohistochemistry. The expression of (voltage-dependent anion channel,VDAC)on mitochondria was evaluated by both Western blotting and Immunohistochemistry.Results:1.Compared with the NC group, BG, BUN, SCr and UAER were significantly increased (P<0.01orP<0.05)in the DM group;BUN,SCr and UAER were significantly decreased in a-LA group compared with the DM group (P＜O.01 or P<0.05);no statistic differences of BG were found between the DM group and the a-LA group.2.Glomerular hypertrophy, mesangial cell proliferation, extracellular matrix accumulation were observed by optics microscope,the mitochondrial swelling and the diminution of cristae were observed by electron microscopy in the DM group.Abnormal histomorphologic changes described above were obviously improved in the a-LA group.3.Compared with the NC group,MDA levels were significantly increased in the serum and renal cortex of the DM group(all P<0.01);MDA levels were significantly decreased in the a-LA group compared with the DM group(all P<0.05);Compared with the NC group, the activities of SOD were significantly reduced in the serum and renal cortex of the DM group(all P＜0.01);the activities of SOD were significantly elevated in the a-LA group compared with the DM group(P<0.01 or P<0.05).4. Compared with the NC group, the activities of SDH, chrome C oxidase, mitochondrial membrane potential were significantly decreased (P＜0.01 or P<0.05);mitochondrial swelling trend remarkably weakened;The activities of SDH,chrome C oxidase, mitochondrial membrane potential in the a-LA group were significantly increased compared with the DM group (P<0.01or P<0.05)and mitochondrial swelling trend reforced.5.Compared with the NC group, the expression of VDAC on mitochondria were significantly reduced (P＜0.01 or P<0.05),and apoptosis-related protein CytC, Smac/DIABLO,AIF release were significantly elevated in the DM group (P＜0.01 orP<0.05);the expression of VDAC significantly were improved(all P＜0.05),and the expression of apoptosis-related protein CytC, Smac/DIABLO, AIF were significantly reduced in theα-LA group, compared with the DM group(all P<0.05).Conclusion:Our findings indicate that oxidative stress,mitochondrial dysfunction, changes in expression of VDAC on mitochondria, CytC,Smac/DIABLO, AIF released by mitochondria may implicate in the mechanism of DN. Moreover, The antioxidant a-LA exert a protective role against the development of DN,the related mechanism may include effectively preventing the generation of oxidants, protecting mitochondrial function, blocking the mitochondrial-dependent apoptosis in renal cells, which is mediated by VDAC via released Cyt C,AIF and Smac/DIABLO.
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