Huangqi Decoction Inhibits Oxidative Stress Via AMPK/Nrf2 Signaling Pathway To Improve Diabetic Nephropathy Podocyte Injury In C57 Mice

Objective:Diabetic nephropathy(DN)is one of the important causes of end-stage renal disease(ESRD),accounting for 16.4% of the primary cause,which is a serious threat to human health.The exact mechanism of the development of DN is still unclear,and the clinical treatment measures are limited.The main measures are to control blood sugar,blood pressure,improve circulation and symptomatic treatment.The curative effect is not significant,which seriously endangers human health and causes enormous medical and social economic burden.Huangqi Decoction(HQD)is from the Seventh Anniversary of Renzhai Zhifang,which is composed of Astragalus,Astragalus,Melon Root,Maimendong,Schisandra,and Glycyrrhiza uralensis.Huangqi Decoction is widely used in the prevention and treatment of diabetes and its related complications,but its role in DN and its underlying mechanisms are not yet clear.This experiment mainly explores the protective effect of HQD on DN podocyte injury by establishing a diabetes model in mice,and studies whether HQD can inhibit oxidative stress and improve podocyte injury through the regulation of AMPK/Nrf2 signaling pathway,thereby improving the occurrence and development of DN.Methods:After 48 SPF C57/6J mice were given adaptive feeding for one week,8mice were randomly selected as the normal group(NC),8 as NC+HQD1.08g/kg group,and the rest as the diabetes model group.Mice were injected intraperitoneally with 100mg/kg streptozotocin(STZ)for two consecutive days,and fasting blood glucose was measured one week later.Blood glucose higher than 350mg/d L was considered as the success of the diabetes model.The mice were randomly divided into 4 groups.There was no significant difference in blood glucose between the groups(P>0.05).The group was divided into model group(DN),HQD group,Compound Cgroup,HQD+Compound C group.Groups were given oral administration for 8 weeks and intraperitoneal injection of Compound C for the last2 weeks.After 8 weeks,the mice were sacrificed.the normal indexes and renal function were detected.PAS staining was used to observe the renal pathology of mice.WT1 immunohistochemistry and TUNEL staining were used to observe the apoptosis of podocytes.Immunohistochemistry and Western blot were used to detect the expression of Nephrin and Podocin.The level of oxidative stress in mouse kidney tissue was detected by fluorescent probe DHE;the expression of AMPK/Nrf2 signaling pathway-related protein was detected by immunohistochemistry,immunofluorescence and Western blot,and the intervention target of HQD to improve podocyte injury was explored.Resultes:Huangqi Decoction can significantly improve the general state of mental state and activity of DN mice.When HQD is given,the body weight is increasing,but it is not statistically significant.Blood glucose,24 h drinking water,24 h urine output are significantly reduced;when HQD treatment is given The levels of ACR and BUN in post-DN mice were decreased,and there was no improvement in serum creatinine.The renal pathology of mice with diabetic nephropathy was improved,the glomerular volume was significantly reduced,the glycogen content was significantly reduced,and the mesangial matrix expansion and glomerular sclerosis were improved.Pathological injury;WT-1 immunohistochemical staining,TUNEL staining can be observed to significantly reduce the apoptosis of glomerular podocytes in DN group after HQD intervention;MCP-1,TNF-α cell inflammation was detected by immunohistochemistry Factors,observed HQD intervention significantly reduced renal inflammation;Western blot analysis found that HQD intervention can up-regulate the expression of p-AMPK,n-Nrf2 protein in renal tissue AMPK / Nrf2 signaling pathway,up-regulation of downstream proteins NQO1,GCLC,GCLM,SOD1,SOD2,HO-1 protein expression;using fluorescent probe DHE detection found that HQD intervention reduced renal tissue oxidative stress;Western blot analysis found that HQD intervention can down-regulate mitochondria apoptotic pathway associated protein Bax,cleaved Caspase3 expression,and Effective prevention and treatment of DN.Conclusion:The experiment have confirmed that the protective effect of HQD on kidney and podocyte injury in DN mice may be related to the regulation of AMPK/Nrf2 signaling pathway.