摘要Objective:To investigate the effects of astragaloside Ⅳ(AS-Ⅳ)on podocyte injury of diabetic nephropathy(DN)and reveal its potential mechanism.Methods:In in vitro experiment,podocytes were divided into 4 groups,normal,high glucose(HG),inositol-requiring enzyme 1(IRE-1)α activator(HG+thapsigargin 1 μmol/L),and IRE-1 α inhibitor(HG+STF-083010,20 μmol/L)groups.Additionally,podocytes were divided into 4 groups,including normal,HG,AS-Ⅳ(HG+AS-Ⅳ 20 μmoVL),and IRE-1 α inhibitor(HG+STF-083010,20 μ mol/L)groups,respectively.After 24 h treatment,the morphology of podocytes and endoplasmic reticulum(ER)was observed by electron microscopy.The expressions of glucose-regulated protein 78(GRP78)and IRE-1 αwere detected by cellular immunofluorescence.In in vivo experiment,DN rat model was established via a consecutive 3-day intraperitoneal streptozotocin(STZ)injections.A total of 40 rats were assigned into the normal,DN,AS-Ⅳ[AS-Ⅳ 40 mg/(kg-d)],and IRE-1 α inhibitor[STF-083010,10 mg/(kg·d)]groups(n=10),respectively.The general condition,24-h urine volume,random blood glucose,urinary protein excretion rate(UAER),blood urea nitrogen(BUN),and serum creatinine(SCr)levels of rats were measured after 8 weeks of intervention.Pathological changes in the renal tissue were observed by hematoxylin and eosin(HE)staining.Quantitative reverse transcription-polymerase chain reaction(RT-PCR)and Western blot were used to detect the expressions of GRP78,IRE-1 α,nuclear factor kappa Bp65(NF-κ Bp65),interleukin(IL)-1 β,NLR family pyrin domain containing 3(NLRP3),caspase-1,gasdermin D-N(GSDMD-N),and nephrin at the mRNA and protein levels in vivo and in vitro,respectively.Results:Cytoplasmic vacuolation and ER swelling were observed in the HG and IRE-1 α activator groups.Podocyte morphology and ER expansion were improved in AS-Ⅳ and IRE-1 α inhibitor groups compared with HG group.Cellular immunofluorescence showed that compared with the normal group,the fluorescence intensity of GRP78 and IRE-1 α in the HG and IRE-1 α activator groups were significantly increased whereas decreased in AS-Ⅳ and IRE-1 α inhibitor groups(P＜0.05).Compared with the normal group,the mRNA and protein expressions of GRP78,IRE-1 α,NF-κ Bp65,IL-1 β,NLRP3,caspase-1 and GSDMD-N in the HG group was increased(P＜0.05).Compared with HG group,the expression of above indices was decreased in the AS-Ⅳ and IRE-1 α inhibitor groups,and the expression in the IRE-1 α activator group was increased(P＜0.05).The expression of nephrin was decreased in the HG group,and increased in AS-Ⅳ and IRE-1 αinhibitor groups(P＜0.05).The in vivo experiment results revealed that compared to the normal group,the levels of blood glucose,triglyceride,total cholesterol,BUN,SCr and urinary protein in the DN group were higher(P＜0.05).Compared with DN group,the above indices in AS-Ⅳ and IRE-1 α inhibitor groups were decreased(P＜0.05).HE staining revealed glomerular hypertrophy,mesangial widening and mesangial cell proliferation in the renal tissue of the DN group.Compared with the DN group,the above pathological changes in renal tissue of AS-Ⅳ and IRE-1 αinhibitor groups were alleviated.Quantitative RT-PCR and Western blot results of GRP78,IRE-1 α,NF-κ Bp65,IL-1 β,NLRP3,caspase-1 and GSDMD-N were consistent with immunofluorescence analysis.Conclusion:AS-Ⅳ could reduce ERS and inflammation,improve podocyte pyroptosis,thus exerting a podocyte-protective effect in DN,through regulating IRE-1 α/NF-κ B/NLRP3 signaling pathway.