Effect of sitagliptin on apoptosis and p38 mitogen activated protein kinase pathway in renal tubular epithelial cells induced by high glucose

Abstract

Objective To observe the effect of dipeptidyl peptidase-Ⅳ (DPP-4) inhibitor sitagliptin on the apoptosis and p38 mitogen activated protein kinase (p38MAPK) signaling pathway in renal tubular epithelial cells induced by high glucose. Methods In vitro cultured rat renal tubular epithelial cell line (NRK-52E) was divided into normal glucose control group (NG), hypertonic control group (MG), high glucose stimulation group (HG), high glucose plus p38MAPK blocker (SB202190) group (HG+SB), and high glucose plus sitagliptin group (HG+ST). Trypan blue experiment was conducted to find out the appropriate concentration of sitagliptin. The activation of p38MAPK signaling pathway was detected by western blot. The relative amount of pro-apoptosis factor anti-apoptotic factor B-cell lymphoma-2 (bcl-2) mRNA and Bcl-2 associated X protein (bax) mRNA was detected by real-time fluorescence quantitative PCR and the ratio of the two was determined. The cell apoptosis rate, was detected by flow cytometry. Transforming growth factor beta (TGF-β) and fibronectin (FN) released into media were quantitatively analyzed by enzyme-linked immunosorbent assay (ELISA). One-way analysis of variance was used for multigroup comparison. Results Western blot revealed that high glucose could increase the expression of p-p38 protein in renal tubular epithelial cells. SB202190 and sitagliptin could significantly reduce the high expression of p-p38 in renal tubular epithelial cells induced by high glucose (0.82±0.12 vs 1.39±0.22, 1.03±0.14 vs 1.39±0.22, t=4.01, 3.71, P<0.05) . Quantitative PCR showed that bax/bcl-2 mRNA in group HG+SB and group HG+ST was significantly lower than that in HG group (1.32±0.54 vs 4.42±0.18, 2.14±0.29 vs 4.42±0.18, t=9.43,11.57, P<0.05). Flow cytometry indicated that compared with NG group, apoptosis rate of HG group increased significantly (17.50±0.35 vs 4.80±0.54, t=34.18, P<0.05) . Compared with the HG group, apoptosis rate in the HG+SB group and the HG+ST group significantly decreased (11.30±1.19 vs 17.50±0.35,14.20±0.81 vs 17.50±0.35, t=8.66, 6.48, P<0.05 respectively). The ELISA results showed that compared with NG group, the expression of TGF-β and FN in HG group increased significantly (536.0±37.9 vs 180.0±17.6, 19.4±2.4 vs 5.3±0.8, t=14.76, 9.64, P<0.05 respectively) . The expression of TGF-β and FN in HG+SB and HG+ST significantly decreased compared with that of HG group (356.0±33.8 vs 536.0±37.9, 390.0+42.3 vs 536.0±37.9, t=6.13, 4.45; 12.5±2.1 vs 19.4±2.4, 14.6±2.7 vs 19.4±2.4, t=3.72, 2.85, P<0.05 respectively) . Conclusion Sitagliptin can reduce the apoptosis and fibrosis of renal tubular epithelial cells through inhibiting the activation of p38MAPK signaling pathway induced by high glucose, thereby play a protective role in kidney. Key words: Diabetic nephropathies; Sitagliptin; p38 mitogen activated protein kinase