Effect Of DPP-â…£ Inhibitor Sitagliptin On P38MAPK Signal Pathway In High Glucose Exposed Renal Tubular Epithelial Cells

Objective:1.To observe the form of cultured rat renal tubular epithelial cell, and grasp the ability of cell-culture.2.To research the effects of DPP-IV inhibitor sitagliptin on p38 MAPK signal pathway in high glucose exposed renal tubular epithelial cells, explore the mechanism of sitagliptin in diabetic nephropathy.Methods:Recover the frozen renal tubular epithelial cells in DMEM low-glucose medium containing 10% fetal calf serum with the condition of 37 ℃ and 5% CO2 incubator,it show cobblestone-appearance,when the vast majority of the cells adherent, change the calf-free DMEM medium in the frozen renal tubular epithelial cells for 24 hours.The synchronized renal tubular epithelial cells were grouping:normal glucose group,high glucose group,hyperosmosis group, SB202190 group(the p38 MAPK inhibitor) with high glucose and DPP-IV inhibitor sitagliptin with high glucose.The activation condition of p38 MAPK were detected by western blot. The rate of renal tubular epithelial cells apoptosis was evaluated by flow cytometry and PCR, include bax,bcl-2,caspase3.Results:1.High glucose could activate the p38 MAPK signaling pathway of renal tubular epithelial cell,increased the expression of p-p38 protein,normal glucose group and hyperosmosis group could detected a small amount of activation of thepathways.SB202190 can reduce the expression of p-p38 protein which triggered by glucose with 25mmol/L.DPP-IV inhibitor sitagliptin also can reduce the activation of p38 MAPK signaling pathway.2.PCR was used to inspect mRNA of apoptosis factor bax and bcl-2.The presentation of bax mRNA in high glucose group was higher than normal group and hyperosmosis group;SB202190 could obviously decrease the relative amount of bax-mRNA triggered by high glucose.DPP-IV inhibitor sitagliptin also decrease the expression due to 25mmol/L glucose.The presentation of bcl-2 mRNA was on the contrary.To compare the ratio of bax mRNA and bcl-2 mRNA between high glucose group and SB202190 group, the difference was statistically significant( P <0.05). It was also statistically significant between high glucose group and sitagliptin group.3.Flow cytometry detected the apoptosis of renal tubular epithelial cell triggered by glucose with 25mmol/L appeared after 24 h, after 48 h the apoptosis was added obviously,the most obvious to 48h(17.5 ± 0.35)%, higher than normal group(4.8 ± 0.24)% and hyperosmosis group(5.2±0.32)%. The apoptosis of SB202190 group(5.2 ± 0.32)%,decreased clearly compared with high glucose group.48 h apoptosis of SB202190 group(11.3 ± 0.19)% decreased 35% compared with the high glucose group.High glucose with DPP-IV inhibitor sitagliptin can reduce apoptosis,The apotosis rate of the group with10μmol/L sitagliptin(14.2 ± 0.31)% decreased 19% compared with the high glucose group,the difference is statistically significant( P <0.05). Just like the expression of caspase3,the expression rate with 10μmol/L sitagliptin decreased 25% compared with the high glucose group,the difference is statistically significant( P <0.05). The ratio of bax and bcl-2 with 10μmol/L sitagliptin was(0.78 ± 0.21),the difference is statistically significant( P <0.05) compared with Group HG.Conclusion:DPP-IV inhibitor sitagliptin can reduce the activation of p38 MAPK trigger by high glucose in renal tubular epithelial cell,always reduce the expression quantity of bax and caspase3,protect the cells in the end.