The Analysis Of Inhibiting Glycogen Synthase Kinase-3β In The Role Of Diabetic Nephropathy In Rats

Background:Diabetic nephropathy (DN) is the major cause of the end-stage renalfailure (ESRD). The main pathological change of diabetic nephropathy isglomerular basement membrane thickening and extracellular matrixaccumulation. In the early of DN, glomerular filtration rate increased is themainly change demonstrating increased blood flow in the kidney. The increasedglomerular size, thickened glomerular basement membrane and increasedextracellular matrix were the main demonstration in Pathology. A recent studyshows that, activity of GSK3β in liver was abnormally increased in the type2diabetes. It weakened the conduction of insulin signals, inhibited the use of thesugar and glycogen synthesis process, and then insulin resistance appeared.Therefore, GSK3β inhibitor can relieve insulin resistance, and treat type2diabetes hopefully. Application of GSK3β related of inhibitors in thetreatment of DN have been reported, the GSK3β as a member of theimportant in insulin signaling pathway, its specific mechanism is not clear inglomerular lesions, therefore, to explore the role of GSK3β in DN andmechanisms in the pathogenesis of DN and find new targets for therapy.Objective:Observing the DN rat proteinuria, blood glucose, serum creatinine andkidney pathology, kidney GSK3β, Akt1mRNA expression level and theexpression of GSK3β and p-GSK3β protein levels after the experient modelbuilt，thus clear GSK3β inhibitor effect and its mechanism of action of DN.Methods:The male Wistar rats were divided into four groups: normal group (NC group, n=10), simple inhibitor group (NY, n=10), diabetic nephropathy group(DN group, n=10), diabetic nephropathy group after application of inhibitor(DY group, n=10). This study diabetes group and diabetes applicationinhibitors all applicate on a high-fat diet in rats， and inject low-dose STZ byintraperitoneal, diabetic nephropathy rat model was established. Stay after thesuccess of the rat model of diabetic nephropathy, DY group celiac injection ofGSK3β inhibitor (lithium chloride). Observation group rats general condition,body weight, blood sugar, urine protein, serum creatinine and other indicators,observe kidney pathological changes. Real-time PCR method is applied todetect glomerular GSK3β, silk/threonine kinase (Akt1) mRNA expression.Theprotein expression of GSK3β and p-GSK3β were examined by Westernblotting.Results:Successful build the rat model of diabetic nephropathy. Compared withNC group, DN group increased significantly, the renal pathological features forthe increasing of glomerular volume, mesangial matrix, diffuse thickening ofbasement membrane, etc. Application of diabetic nephropathy rats GSK3βinhibitor lithium chloride24h urine protein quantitative declined obviously,PAS staining observed DY group kidney pathological changes significantlyreduce, increased mesangial cell proliferation and matrix tend to be better,protein tubular and interstitial inflammatory cell infiltration.Real-time PCRmethod is applied to research shows that: compared with NC group, DN groupGSK3β expression capacity significantly increased (P <0.05), while after DYinhibitor treatment group rats kidney GSK3β expression quantity reduced (P <0.05)，the change of Akt1have not statistical significance; Western-blotmethod is applied to research shows that: compared with NC group, DN groupp-GSK3β expression level is low, DY group p-GSK3β expression increased(p <0.05), two groups of total GSK3β protein expression level changes arenot obvious.Conclusion:By lithium chloride inhibiting GSK3β of diabetic nephropathy ratskidney,we found that lithium chloride can make p-GSK3β expression quantityincreases, and the total GSK3β did not change significantly. We can deduce thatlithium chloride can control the non-phosphorylated GSK3β expression, reduceproteinuria and relieve renal pathological changes for protect kidney function.