Effect of forkhead transcription factor O1 on oxidative stress in rat mesangial cells cultured under high glucose conditions

Abstract

Objective To study the effect and mechanism of forkhead transcription factor O1(FoxO1) on oxidative stress in rat mesangial cells(MCs) cultured under high glucose conditions. Methods The MCs cultured in normal glucose (5.6 mmol/L) medium were transfected by the plasmid vector of the FoxO1 short hairpin RNAs (FoxO1 shRNA), and then cultured in high glucose (30.0 mmol/L) medium and resveratrol (Resv 20.0 μmol/L). The MCs cultured in normal glucose medium served as normal control group (group NG), and the MCs cultured in high glucose medium were divided into five groups: simple high glucose group (group HG), HG+ Resv group, HG+ FoxO1 shRNA group, HG+ Resv+ FoxO1 shRNA group, HG+ Negative control group (shNC). MCs in each group were cultured for 72 h. The level of reactive oxygen species (ROS) was assessed by fluorescence microplate reader. The mRNA expression of silent information regulator 1(Sirt1), FoxO1, manganese superoxide dismutase (MnSOD) were detected by reverse transcription polymerase chain reaction(RT-RCR). The protein expression of FoxO1 and phosphorylation FoxO1(p-FoxO1) were determined with Western blotting methods. One-way analysis of variance was applied in the data comparisons among the groups. Results Compared with group NG, the mRNA expression of Sirt1 and MnSOD were significantly reduced but the p-FoxO1 and ROS were enhanced in group HG (t=12.38, 14.13, 13.27, 8.36, all P<0.05). Compared with group HG, the mRNA expression levels of FoxO1 and MnSOD were obviously inhibited, but the level of ROS was enhanced in group HG+ FoxO1shRNA(t=20.61, 13.61, 10.13, all P<0.05); and the mRNA expression of Sirt1 and MnSOD increased, whereas the level of p-FoxO1 and ROS decreased significantly in group HG+ Resv (t=6.02, 5.39, 10.69, 5.37, all P<0.05). Compared with group HG+ Resv, the mRNA expression of FoxO1 and MnSOD was inhibited but the ROS was enhanced significantly in group HG+ Resv+ FoxO1shRNA (t=22.53, 15.31, 14.63, all P<0.05). Conclusion FoxO1 is the important transcription factors to protect MCs against the damage of oxidative stress by activating the expression of downstream antioxidant target gene of MnSOD. Key words: Forkhead transcription factors; Oxidative stress; Mesangial cells; High glucose; Rats