The Research Of Dapagliflozin Improving Vascular Dysfunction By Activating SIRT1/eNOS Signaling Pathway

Objective:This study aims to figure out whether dapagliflozin attenuates endothelial senescence and improves diabetic endothelium-dependent vasorelaxation function,and explore the possible pathway affecting vasodilation.Methods:This study was divided into in vivo and in vitro experiments.In vivo experiment,the diabetic mice（db/db mice）and heterozygous littermates（db/m mice）were randomized four group.Dapagliflozin treatment groups were given a daily oral dose of dapagliflozin（1 mg/kg/day）for 8 week.The placebo groups were given a daily oral dose of vehicle（saline）as was consumed by the dapagliflozin-treated group.Blood glucose,body weight and food intake were measured weekly.In vitro experiment,high glucose or H₂O₂ induced human umbilical vein endothelial cells（HUVECs）senescence.We used a specific and potent pharmacological inhibitor of SIRT1,Nicotinamide（NAM）,to inhibit the activity of SIRT1.It was evaluated that SIRT1 is the key protein of dapagliflozin for its anti-aging effect.Endothelial dysfunction and senescence was assessed by senescence?associated beta?galactosidase（SA‐β‐gal）activity,senescence marker protein（p21,p53）,oxidative stress,nitric oxide（NO）and inflammatory cytokines.Assessment of chronic vaso-protective effect of dapagliflozin was performed in organ bath studies.The interaction between SIRT1 and e NOS was detected by co-immunoprecipitation.Results:In vivo experiment,dapagliflozin treatment decreased the level of blood glucose and insulin resistance index,while increased serum NO concentration.As for chronic vaso-protective effect of dapagliflozin,dapaglifozin treatment improved endothelium-dependent vasorelaxation such that dilation was modestly and significantly increased.Dapagliflozin treatment also decreased the senescence of tunica intima and endothelial cells senescence of the aorta,as well as inflammatory cytokines.In vitro experiment,HG or H₂O₂ treatment induced cellular senescence in HUVECs,increased p21,p53 and ROS formation,and decreased the expression of SIRT1 and p-e NOS,but exposure to dapagliflozin reversed the effect.e NOS acetylation levels as well as levels of SIRT1 associated with e NOS following co-immunoprecipitation.The e NOS acetylation was higher in HG-or H₂O₂-treated cells and was restored by dapagliflozin treatment.SIRT1 levels associated with e NOS were decreased in H₂O₂-treated cells and were restored by dapagliflozin treatment.NAM reduced the anti-senescence effects of dapagliflozin in HG-or H₂O₂-induced senescent endothelial cells（Similar to the aging model group）.Conclusions:Dapagliflozin attenuates endothelial senescence and improves endothelium-dependent vasorelaxation by activating SIRT1/e NOS signaling pathway in type 2 diabetes mellitus.Figures 33,Tables 8,References 56...