The Effect Of Sitagliptin On Autophagy And Apoptosis In H9c2 Cardiomyocytes Induced By High Glucose And High Lipids

Objective:The rat H9c2 cardiomyocytes were cultured in vitro by high-sugar and high-lipid,and the diabetic environment of cardiomyocytes was simulated,and then sitagliptin was administered to observe the autophagy and apoptotic expression of high-sugar and high-fat model group and the normal control group and the drug-treated group.Methods:The experiment was divided into three parts:（1）Rat H9c2cardiomyocytes cultured in vitro were randomly divided into normal control group（Con group）,high glucose and high fat group（HP group）,high glucose and high fat+different concentrations of sitagliptin group(0.1,1,5,10,20,and 50μmol·L^-1)（HP+Sit group）,cell viability was measured by Cell Counting Kit-8（CCK-8）,and the optimal concentration of sitagliptin was selected.（2）Rat H9c2 cardiomyocytes cultured in vitro were randomly divided into normal control group（Con group）,high glucose and high fat group（HP group）,sitagliptin group（Sit group）,high glucose and high fat+sitagliptin group（HP+Sit group）,the expression of autophagy-related proteins LC3 and Beclin-1 was detected by Western blot;theexpressionofautophagosomesincellswasdetectedby single-dansylsulfonylamine（MDC）fluorescent staining,this method was used to detect the formation of autophagosomes in cells.This step was to observe the effect of sitagliptin on autophagy of H9c2 cells under high glucose and high fat conditions.（3）Rat H9c2cardiomyocytes cultured in vitro were randomly divided into normal control group（Con group）,high glucose and high fat group（HP group）,sitagliptin group（Sit group）and high glucose high fat+sitagliptin group（HP+Sit group）（same with the second part）.Western blot was used to detect the expression of apoptosis-related proteins Cleaved-PARP and Cleaved-Caspase3,flow cytometry were used to observe the effect of sitagliptin on cell viability and apoptotic rate of H9c2 cells under high glucose and high fat conditions.This step was to observe the effect of sitagliptin on apoptosis of H9c2 cells under high glucose and high fat conditions.Results:1.Cell viability was detected by CCK8:The results of CCK8 showed that the OD value of cell growth viability in high glucose and high fat group was significantly lower than that in normal control group（P<0.05）;after treatment with 0.1,1 and 2μmol·L^-1 of sitagliptin,the OD value was Significantly elevated（P<0.05）,and the highest OD value at a concentration of 0.1μmol·L^-1 of sitagliptin.Finally,the final concentration of sitagliptin was 0.1μmol·L^-1,and the action time was 24 hours,which was used as the treatment condition of the drug.2.Western blotting results showed that compared with the normal control group,the expression of autophagy-related proteins LC3 and Beclin-1 were significantly decreased in the high-sugar and high-fat group（P<0.05）.After treatment with sitagliptin,the expression levels of LC3 and Beclin-1 were significantly higher compared with that in the high-sugar and high-fat group（P<0.05）.The expression of each protein in the sitagliptin group was not significantly different from that in the normal control group.The results of single-dansylsulfonylamine（MDC）fluorescent staining showed that compared with the normal control group,the proportion of MDC-positive cells in the high-sugar and high-fat group decreased,and there was a significant increase after the administration of sitagliptin.There was no significant difference between the expression of sitagliptin alone and the normal control group.3.Western blotting results showed that compared with the normal control group,the expression of apoptosis-related proteins Cleaved-PARP and Cleaved-Caspase3 in the high-sugar and high-fat groups were significantly increased（P<0.05）.After treatment with sitagliptin,the expression of Cleaved-PARP and Cleaved-Caspase3 protein was significantly lower compared with that in the high-sugar and high-fat groups（P<0.05）.The expression of each protein in the sitagliptin alone group was not significantly different from that in the normal control group.The results of flow cytometry showed that the apoptosis rate of high glucose and high fat group was significantly increased（P<0.05）;after treatment with sitagliptin,the apoptosis rate was significantly lower（P<0.05）.The apoptotic rate of the sitagliptin alone group was not significantly different from that of the normal control group.Concusion:1.Under the condition of high glucose and high fat,the growth of H9c2 cardiomyocytes was inhibited.Sitagliptin can significantly reverse the growth inhibition of H9c2 cardiomyocytes induced by high glucose and high fat.2.There was a low level of autophagy expression in H9c2 cells in normal control group;after high glucose and high fat stimulation,the autophagy level was significantly decreased and the apoptosis was significantly enhanced.Administration of sitagliptin significantly enhanced the expression of autophagy in H9c2 cardiomyocytes and decreased the rate of apoptosis.This indicates that sitagliptin can promote the autophagy of H9c2cardiomyocytes,inhibit cell apoptosis,and alleviate the damage of H9c2 cardiomyocytes induced by high glucose and high fat,which may have a protective effect on the myocardium of diabetic patients.