肉桂醛调控糖酵解改善糖尿病肾病小鼠炎症反应及肾纤维化的作用机制OA

Objective To investigate the mechanism by which cinnamaldehyde regulates glycolysis to ameliorate inflammatory response and renal fibrosis in mice with diabetic kidney disease(DKD).Methods(1)Animal experiment:A DKD mouse model was established by feeding a high-fat diet for 6 weeks followed by intraperitoneal injection of streptozotocin(50 mg·kg-1)for 5 consecutive days.C57BL/6J mice were randomly divided into a normal group,a model group,a cinnamaldehyde group(10 mg·kg-1),and a dapagliflozin group(1 mg·kg-1),with 8 mice per group.Drugs were administered by gavage once daily for 4 weeks.Body weight and fasting blood glucose levels were measured weekly during the treatment period.The kidney index was calculated.Serum levels of renal function markers[creatinine(Scr)and blood urea nitrogen(BUN)]and lipid profiles[triglycerides(TG)and total cholesterol(TC)]were detected.Urine was collected for measurement of urinary microalbumin(mALB).Kidney tissue supernatant was prepared to determine the levels of lactate and adenosine triphosphate(ATP).Renal histopathological changes were observed by HE,Masson,and periodic acid-Schiff(PAS)staining.The mRNA expression levels of TNF-α,IL-6,IL-1β,Col1a1,and FN in kidney tissue were detected by RT-qPCR.The protein expression levels of E-cadherin,N-cadherin,TGF-β,α-SMA,PKM2,and LDHA in kidney tissue were detected by Western Blot.(2)Cell experiment:mouse renal tubular epithelial cells(mRTECs)were injured by high glucose(30 mmol·L-1)plus 10 μg·mL-1 LPS to establish the model group.The cells were treated with 10,20,or 40 μmol·L-1 cinnamaldehyde and the glycolysis inhibitor 2-deoxy-D-glucose(2-DG,5 mmol·L-1)for 24 hours.Cell viability was assessed by CCK-8 assay.The mRNA expression levels of TNF-α,IL-6,IL-1β,Col1a1,and FN were detected by RT-qPCR,and the protein expression levels of PKM2,HK2,and LDHA were detected by Western Blot.Results(1)Compared with the normal group,the model group exhibited significantly increased kidney index,fasting blood glucose,and mALB levels(P＜0.05,P＜0.01);significantly elevated serum levels of Scr,BUN,TG,and TC(P＜0.01);mesangial proliferation,disordered arrangement of tubular epithelial cells,vacuolar degeneration,inflammatory cell infiltration,marked mesangial matrix expansion,thickened glomerular basement membrane,and obviously increased glycogen and collagen fiber deposition;significantly upregulated mRNA expression of TNF-α,IL-6,IL-1β,Col1a1,and FN,and protein expression of N-cadherin,α-SMA,and TGF-β(P＜0.05,P＜0.01),whereas E-cadherin protein expression was significantly decreased(P＜0.05);markedly increased lactate level(P＜0.01),significantly decreased ATP level(P＜0.01),and significantly upregulated LDHA protein expression(P＜0.05)in kidney tissue.Compared with the model group,both the cinnamaldehyde group and the dapagliflozin group showed significantly reduced kidney index and mALB level(P＜0.01);significantly decreased serum levels of Scr,BUN,TG,and TC(P＜0.05,P＜0.01);obviously ameliorated renal pathological injury with attenuated inflammatory cell infiltration,glycogen deposition,and fibrosis;significantly downregulated mRNA expression of TNF-α,IL-6,IL-1β,Col1a1,and FN,and protein expression of N-cadherin and TGF-β(P＜0.05,P＜0.01),whereas E-cadherin protein expression was significantly increased(P＜0.05);remarkably reduced lactate level(P＜0.05),elevated ATP level(P＜0.05),and significantly downregulated LDHA protein expression(P＜0.05,P＜0.01)in kidney tissue.(2)Compared with the low-glucose control group,the model group showed significantly decreased mRTECs viability(P＜0.01),significantly increased mRNA expression of TNF-α,IL-6,IL-1β,Col1a1,and FN(P＜0.01),and significantly upregulated HK2 protein expression(P＜0.05).Compared with the model group,10,20,and 40 μmol·L-1 cinnamaldehyde significantly increased mRTECs viability(P＜0.05,P＜0.01);the 40 μmol·L-1 cinnamaldehyde group and the 2-DG group exhibited significantly reduced mRNA expression of TNF-α,IL-6,IL-1β,Col1a1,and FN(P＜0.05,P＜0.01),and significantly downregulated PKM2 and HK2 protein expression(P＜0.05,P＜0.01).Conclusion Cinnamaldehyde can ameliorate renal pathological injury,and alleviate inflammatory response and renal fibrosis in DKD mice,the mechanism of which may be associated with the inhibition of glycolysis-related pathways.