Analysis The Relationship Between The Expression Of P66shc And Clinical Phenotypes Of Diabetic Nephropathy

Backgroud:Diabetic nephropathy is a well known and most serious microvascular complication in patients with diabetes mellitus,which is the leading cause of death in patients with diabetes. In the past10years,the annual incidence of this disease has been doubled, and at present it accounts for almost50%of all end-stage renal diseases. Although the pathogenetic mechanisms of diabetic nephropathy is still unclear,more and more studies have shown that overproduction of ROS induced by hyperglycemia play a central role in the development of diabetic nephropathy. In the past,studies on the pathogenesis of diabetic nephropathy have focused on the renal glomerulus, until recently,a few reports found that tubulointerstitial lesions is not secondary to glomerular damage,but the early and original features of the DN,which has become an important indicator to predict the deterioration of renal function. p66Shc localizing within the mitochondrial inter-membrane space, is the key protein in the regulation of cellular oxidative stress responses and life span. Under various proapoptotic signals, overexpression of p66Shc oxidizes cytc, leading to the excessive production of ROS, aggravating cell oxidative damage and apoptosis. There is growing evidence that the proapoptotic activity of p66Shc is dependent on oxidative stress-mediated phosphorylation at serine36residue. Although several studies in vitro and in vivo have shown that expression of p66Shc and phosphorylated p66Shc (at Ser36residue)was increased in mice with STZ-induced diabetes. Deletion of p66Shc longevity gene protects against STZ-induced experimental diabetic glomerulopathy by reducing Oxidative Stress and streptozotocin-induced diabetic p66Shc knockout (KO) mice showed less marked changes in renal function and structure, as indicated by the significantly lower levels of advanced glycationend products、 nuclear factor-κβ(NF-κβ)and proteinuria. Based on these studies, We hypothesize that the p66shc expression level in renal tissue is upregulated and It may be closely related to oxidative damage and apoptosis in tubular cells in patients with DN. To verify our hypothesis, we carried out the experiments as follow.Chapter:1Analysis of the relationship between the expression of p66shc in peripheral blood mononuclear cell (PBMC) and clinical phenotypes of diabetic nephropathyObjective:To observe the expression of p66shc in the PBMC and analyse the ralationship among clinical index of patients with diabetie kidney disease.Methods:60patients were enrolled into the study, they were diagnosed as DN(n=30)、 non-DN(primary kidney disease,n=15)and Normal control (n=15) based on clinical manifestations and renal pathology. The expression of p66ShcmRNA in PBMC was examined by Real-time PCR. Results:Real-time PCR showed that expression of p66Shc mRNA in PBMC of non diabetic nephropathy increased remarkably than in Normal control group(p<0.05);Compare with the normal control and non diabetic nephropathy group, there was a significant increase in the expression of p66shc in diabetic nephropathy(p<0.05). The correlation analysis indicated that the expression p66shc in PBMC is closely related to diabetes duration, triglycerides and blood glucose levels.Conclusion:The expression of p66shc is significantly up-regulated in PBMC of DN patients and p66shc is positively related with diabetes duration, triglycerides and blood glucose. Objective:To observe the expression of p66shc in the kidney in order to investigate the relationship of p66shc and high gluose ambience in diabetic nephropathy patients.Methods:22patients were enrolled into the study, they were diagnosed as DN(n=12)or minimal change disease (primary kidney disease,n=10) based on clinical manifestations and renal pathology. Renal pathological changes were observed by Masson、 PASM and PAS staining.Interstitial damage score was evaluated.The expression of p66shc、p-p66shc and ROS was tested by immunohistochemistry or immunofluorescence.Results:Masson,PASM and PAS stainin showed that there were many lesions in the kidneys of DN patients such as moderate to severe proliferation of mesangial matrix and diffuse or segmental thickening of the glomerular basement membrane (GBM),which eventually leads to nodular glomerulosclerosis, also known as Kimmelstiel-Wilson lesions, tubular hypertrophy followed by thickening of the tubular basement membrane (TBM) compensatory expansion and local interstitial fibrosis with inflammatory cells infiltration.Meanwhile, the lesions in the kidneys of patients with minimal change disease were less severe. There were mild mesangial cell and matrix proliferation, basement membrane is not thick, mild granular degeneration and vascuar degeneration in renal tubular epithelial cells and no significant abnomalities in interstitial and renal vascular. Interstitial damage score in the kidneys of DN patients were higher than that in control group. Immnunohistochemistry and immunofluorescence showed that the expression of p66shc and p-p66shc was significant up-regulated in DN patients compared with that in control group.Conclusion:The expression of p66shc、p-p66shc is significantly up-regulated in the kidney of DN patients and p66shc is positively related to production of cellular ROS.