Effects Of Liraglutide On Oxidative Stress In Kidney Of Insulin Resistance Rats And Relative Mechanism Investigation

Objective:To study the damage of oxidative stress to kidney and explore the protective effect of liraglutide on kidney and provide an effective way to prevent chronic kidney disease in clinical.Method:Class SPF male Wistar rats(n=54),4 month old(230-260g),were randomly divided into normal control group(n=16)and model group(n=38),fed with normal diet and high-fat diet for 8 weeks.Selective 5 rats in each group and glucose infusion rate(GIR)was measured by euglycemic clamp test in each group,and the IR model was successfully established.Then the model rats were randomly divided into four groups:normal control group,insulin resistance(IR)group,intervention group 1,intervention group 2,intervention group 1 and intervention group 2 were given subcutaneous injection of liraglutide,concentration were 100μg/(kg·d)and 200μg/(kg·d).After 2 weeks of drug intervention,5 rats in each group were taken,and then the clamp test was performed to calculate the GIR;the ultrastructural changes of renal tissue were observed under transmission electron microscope;separating serum,the fasting blood glucose(FBG),fasting plasma insulin(FINS),free fatty acids(FFA),total cholesterol(TC),triglyceride(TG),serum creatinine(Scr),blood urea nitrogen(BUN),24 hours urine protein and renal oxidative stress index(MDA,SOD,8-OHdG)were detected;and detect the expression of renal diacylglycerol(DAG),Protein kinase C-β(PKC-β),NADPH oxidase subunit P22 phox and P47 phox in kidney.Results:1.High-fat diet induced IR model in ratsAfter fed 8 weeks,compared to the control group,GIR decreased obviously(20.4±3.6 vs 30.2±2.5,P<0.05),but FBG,FINS increased obviously in the HF group(P<0.05).2.Comparison of the general situationFBG,FINS,FFA,TCH,TG and DAG in kidney were increased in IR group than control group(P<0.05);the TG,FFA in intervention group 1 were decreased than IR group(P<0.05);while the targets in intervention group 2 were decreased remarkablely than IR group(P<0.05);while the FFA,DAG in intervention group 2 were significantly decreased(P<0.05).3.Comparison of glucose infusion rate(GIR)Before the liraglutide intervention:the GIR was decreased in IR group than the contol group(20.4±3.6 vs 30.2±2.5,P<0.05).After tervention two weeks,the GIR was decreased in IR group while compared with contol group(18.8±2.0 vs 30.8±2.4,P<0.05);while the GIR in intervention groups was deeply increased(25.8±1.4,21.6±2.5 vs 18.8±2.0,P<0.05);Compared with intervention group 1,the intervention group 2 was increased still more(P<0.05).4.Ultrastructural changes of kidney tissue in four groups of ratsCompared with contol group,the change of glomerular basement membrane thickening and some foot process fusion,lack of such could be seen in electron microscopy of IR.Treatment by liraglutide,the abnormal changes were alleviated in liraglutide groups,although not normalized.5.Comparison of the Scr,BUN,24 hours urine protein in four groups of ratsCompared with contol group,Scr,BUN,24 hours urine protein were increased in IR group(P<0.05);Compared with IR group,the 24 hours urine protein was decreased in intervention group 1(P<0.05)and the Scr,BUN,24 hours urine protein were decreased in intervention group 2(P<0.05);Compared with intervention group 1,the Scr,BUN,24 hours urine proteinin high dose group were significantly decreased(P<0.05).6.Effect of liraglutide on renal oxidative stress parameter and antioxidant markersThe content of MDA in renal of IR was increased,the activity of SOD and 8-OHdG were significantly lower compared with the control group(P<0.05);treatment by liraglutide reducing damage of oxidative stress to the kidneys: lower the content of MDA,increased the activity of SOD and 8-OHdG(P<0.05);while the high dose group was vary significantly(P<0.05).7.Effect of liraglutide on the expression of mRNA in PKC-β,P22 phox,P47phoxThe expression of mRNA in PKC-β,P22 phox,P47phox were increased in IR group than contol group(PKC-β:6.54±0.41 vs 1.00±0.00;P22phox: 4.89±0.38 vs 1.00±0.00;P47phox:4.77±0.76 vs 1.00±0.00;P<0.05);Compared with IR group,the expression of mRNA in PKC-β,P22 phox,P47phox were decreased in intervention groups(PKC-β:2.35±0.51 vs 5.82±0.60 vs 6.54±0.41;P22phox:2.30±0.42 vs 4.47±0.41 vs 4.89±0.38;P47phox:2.21±0.30 vs 3.63±0.44 vs 4.67±0.76,P<0.05).Compared with intervention group 1,the expression of mRNA in PKC-β,P22 phox,P47phox were decreased inintervention group 2(P<0.05).8.Effect of liraglutide on the expression of protein in PKC-β,p-PKC-βThe protein expression of p-PKC-β was increased(2.71±0.29 vs 1.55±0.16,P<0.05),while the PKC-β was found no obvious change in IR group(3.39±0.14 vs 3.60±0.28,P<0.05);treatment by liraglutide,the expression of p-PKC-β was decreased(1.91±0.10 vs2.32±1.19 vs 2.71±0.29,P<0.05),and the expression of p-PKC-β in intervention group 2 was decreased more obvious(P<0.05).9.Effect of liraglutide on the expression of protein in P22 phox,P47phox in kidney of ratsThe protein expression of P22 phox,P47phox were increased in IR group(P22phox:2.72±0.20 vs 1.27±0.24;P47phox:2.17±0.39 vs 1.13±0.15,P<0.05);Compared with IR group,the expression of P22 phox,P47phox were decreased in intervention groups(P22phox:1.75±0.13 vs 2.32±0.21 vs 2.72±0.20;P47phox:1.30±0.18 vs 1.66±0.15 vs 2.17±0.39,P<0.05),while the expression of P22 phox,P47phox were decreased further in intervention group 2(P<0.05).Conclusion:1.High fat diet can cause IR in rats,and kidney damage has been found in IR.2.Liraglutide can downregulate the PKC-NADPH oxidase pathway,thereby improving the renal oxidative stress,reduce kidney damage caused by IR,and then play a protective effect on the kidney.