The Protective Effects And Mechanisms Of Dapagliflozin On Kidney Injury In New-onset Type 2 Diabetes Mellitus

Objective: Many large clinical studies have confirmed that SGLT2 i has secondary preventive effect on DKD,but it is not clear whether SGLT2 i has primary preventive effect on DKD.Functional magnetic resonance imaging(f MRI),as a non-invasive and high-resolution imaging method,can provide quantitative parameters for evaluating the changes of microstructure and function of organs and tissues.In this study,multimodal f MRI technique was used to evaluate the primary preventive effect of dapagliflozin on diabetic nephropathy.T2 DM mouse model was established and treated with SGLT2 i to explore the mechanism of renal protective effect of SGLT2 i on T2 DM.Method: 1.A total of 40 newly diagnosed T2 DM patients were recruited in the outpatient clinic of Metabolic Diseases Hospital of Tianjin Medical University from June 2017 to December 2018.Finally,30 eligible patients were enrolled and randomly divided into two groups,including 10 patients in the dapagliflozin group and 20 patients in the control group(acarbose treatment).They were treated with drugs for 24 weeks and visited regularly at baseline,4th week,12 th week and 24 th week.Basic information,glucose metabolism index,biochemical index and abdominal multimodal fMRI examination before and after treatment were completed.2.Taking 25 healthy male C57BL/6 mice as the research object,a high-fat diet and STZ-induced T2 DM model were established,and SGLT2 i were given for intervention treatment.The mice were randomly divided into 5 groups: NC group,DM group,Dapa group(1mg/kg/d),Empa group(1mg/kg/d)and Cana group(10mg/kg/d).The mice in the intervention group were given intragastric administration for 8 weeks.The changes of blood glucose and body weight of mice were detected every week,and the pathological changes of glomeruli and renal tubules in each group were observed by light microscope.The morphological changes of mitochondria in renal tubular epithelial cells were observed by TEM.RT-PCR and Western Blot techniques were used to detect renal mitochondrial fusion,fission-related protein m RNA and protein expression in each group.Result: 1.For patients with new-onset T2 DM,Hb A1 c decreased by 1.37% compared with the baseline after 24 weeks of treatment with dapagliflozin,which was similar to that of acarbose 100 mg tid.Compared with baseline,FBG decreased 1.92mmol/L in dapa group,and the decreasing degree of FBG and the rate of reaching the standard(FBG<7mmol/L)at 4th and 12 th weeks were better than those in control group(P<0 05).In terms of body weight,the body weight of dapa group decreased by 4.65 kg compared with the baseline(P<0 05).In terms of blood pressure reduction,SBP decreased by 2.22 mm Hg and DBP decreased by 3.78 mm Hg(P>0 05).Blood lipid index: compared with the baseline,HDL in the dapa group increased 0.19mmol/L(P<0 05).Renal function index: e GFR had no significant change compared with baseline,uric acid decreased 42.6umol/L compared with baseline(P>0 05).2.Abdominal f MRI examination was performed in patients with newly diagnosed T2 DM before and after treatment.BOLD MRI showed that the R2* value of renal cortex and medulla decreased after treatment in dapa group,suggesting that the partial pressure of oxygen in renal cortex and medulla was increased,and dapagliflozin could improve the state of renal hypoxia in patients with T2 DM.DTI MRI showed that the FA value of renal cortex and medulla increased after treatment,suggesting that the anisotropy of water molecular movement in renal cortex and medulla increased,and dapagliflozin could improve the pathological changes of renal microstructure in patients with T2 DM.Dixon MRI showed that the FF value of kidney decreased after treatment,suggesting that the fat content of kidney was lower than that before treatment,and dapagliflozin could reduce the lipid accumulation of kidney.3.Abdominal f MRI examination was performed in patients with newly diagnosed T2 DM before and after treatment.Dixon MRI showed that the FF values of liver and pancreas decreased after dapa treatment,suggesting that the fat content of liver and pancreas decreased after dapa treatment,suggesting that dapagliflozin can reduce visceral fat.4.The T2 DM model was established by feeding C57BL/6 mice with high-fat diet for 8 weeks and injecting STZ(60mg/kg)for 5 days.Under light microscope,PAS staining of mouse kidney showed that in DM group,the glomerulus became larger,the renal capsule narrowed or even disappeared,the structure of glomerular capillary loop was blurred,and the PAS positive area in Mesangial area became more severe.SGLT2 i can improve the above pathological changes,and the improvement effect of DM+Cana group is better than that of DM+Dapa group and DM+Empa group.TEM observation showed that in DM group,the arrangement of renal tubular epithelial cells was disordered,the renal tubular basement membrane thickened irregularly,the intracellular mitochondria became smaller and shorter,the structure of mitochondrial crest was not clear,and the outer membrane of mitochondria was degraded locally.Compared with DM group,the arrangement of renal tubular epithelial cells in SGLT2 i intervention group was improved,the thickness of renal tubular basement membrane was uniform,most of the intracellular mitochondria were long rod-shaped,and most of the mitochondrial crest structure was clear.5.The expression of mitochondrial fission-related protein Drp-1 m RNA and protein in DM group was significantly higher than that in NC group(P<0.05),while the expression of mitochondrial fusion-related protein Mfn-2 m RNA and protein in SGLT2 i group was significantly lower than that in NC group(P<0.05).SGLT2 i intervention group could down-regulate the expression of Drp-1 and up-regulate the expression of Mfn-2 at gene and protein level.Conclusion: Dapagliflozin has the effect of improving renal oxygenation and reversing early renal lesions in patients with new-onset T2 DM,and this protective effect is independent of hypoglycemic effect and "tube-ball feedback" mechanism.The protective effect of SGLT2 inhibitor on diabetic kidney can be achieved by improving the function of renal mitochondrial fusion and fission.