中国被毛孢菌粉水提物对链脲佐菌素诱导2型糖尿病并发肾病小鼠的保护作用及机制研究OA

Objective:To investigate the mechanism by which Hirsutella sinensis mycelium(HSM)protects glomerular function and alleviates streptozotocin-induced type 2 diabetic nephropathy via regulating apoptosis and epithelial-mesenchymal transition(EMT)signaling pathways.Methods:A total of 45 male C57BL/6 mice were enrolled.Thirty-five mice were fed a high-fat diet for 3 weeks,followed by intraperitoneal injec-tion of streptozotocin(50 mg/kg)for 5 consecutive days to establish a type 2 diabetes model.Thirty mice with successful model establishment,diabetic mice were randomly divided into model group,low-dose HSM group(2 g/kg)and high-dose HSM group(4 g/kg),and administered continuously for 28 days.Fasting blood glucose(FBG)of each group was measured at baseline(day 0)and after 28 days of administration.In-traperitoneal glucose tolerance test was performed to evaluate islet secretory function,and the area under the curve(AUC)was calculated.After 28 days of treatment,serum samples were collected for detection of bio-chemical indicators including triglyceride(TG),alkaline phosphatase(ALP)and alanine transaminase(ALT).Renal pathological lesions were observed via hematoxylin-eosin(HE),Masson's trichrome and periodic acid-Schiff(PAS)staining.The mRNA expression levels of Caspase-3,Caspase-9,TGF-β1,Vimentin and Slug in renal tissues were determined by quantitative real-time reverse transcription polymerase chain reac-tion(qRT-PCR).The xCELLigence RTCA DP system was used to dynamically monitor the protective effect of HSM water extract on H2O2-injured NRK-52E renal tubular epithelial cells,and the results were further verified by CCK-8 assay.Results:After 4 weeks of HSM intervention,FBG level in the model group was markedly higher than that in the normal group(P＜0.001),while FBG was significantly decreased in both HSM treatment groups compared with the model group(P＜0.01).In addition,HSM markedly reversed the excessive water intake,food intake and urine output in diabetic nephropathy mice.The AUC value of glu-cose tolerance test was significantly elevated in the model group(P＜0.001),whereas obvious reduction of AUC was observed in HSM-treated groups(P＜0.05).Biochemical analysis revealed that serum TG,ALP and ALT levels were remarkably reduced after HSM treatment(P＜0.05,P＜0.001,P＜0.05).The renal in-dex of the model group was significantly higher than that of the normal group.Renal histopathological stain-ing showed disorganized renal structure,glomerular atrophy,vacuolation and partial exfoliation of renal tubu-lar epithelial cells,massive collagen deposition in renal interstitium and expanded glomerular mesangial ma-trix in model mice;all these pathological abnormalities were alleviated to varying degrees after HSM treat-ment.qRT-PCR results demonstrated that the renal mRNA expression of Caspase-3,Caspase-9,TGF-β1,Vimentin and Slug was significantly downregulated in HSM-treated groups relative to the model group(P＜0.05,P＜0.05,P＜0.01,P＜0.001).Real-time monitoring by xCELLigence RTCA DP revealed that HSM water extract(1.2 mg/mL,2.4 mg/mL)exerted persistent protective effects against H2O2-induced injury in NRK-52E cells.This result was also confirmed in the viability detection of NRK-52E renal cells after 24h of H2O2 stimulation.Conclusion:Hirsutella sinensis mycelium ameliorates diabetic nephropathy through sup-pressing renal apoptosis and EMT by modulating the TGF-β1 signaling pathway.