Effects Of1,25-dihydroxyvitamin D₃on High Glucose-induced Expression Of UCP2and Oxidative Stress In Human Renal Tubular Epithelial Cells

Objective To investigate the influence of1,25-dihydroxyvitamin D3(1,25-(OH)2VitD3) on the expression of uncoupling protein2(UCP2) and OxidativeStress in human renal tubular epithelial cells (HK-2cells) induced by high glucose.Methods The HK-2cells with different culture media were divided into severalgroups at random: Normal glucose group (NG,5.5mmol/L D-glucose), High glucosegroup (HG,30mmol/L D-glucose) and Mannitol group (MG,5.5mmol/LD-glucose+24.5mmol/L mannitol), three groups (V1、V2、V3) which were exposed tomedium containing30mmol/L D-glucose and different concentrations of1,25-(OH)2VitD3(V1group10-9mol/L、V2group10-8mol/L、V3group10-7mol/L),N-Acetyl-L-cysteine positive control group (NAC) and solvent control group (SG).The level of intracellular reactive oxygen species (ROS) was examined byfluorescence microscope. With flow cytometry, the changes of mitochondrialmembrane potential (ΔΨm) were tested. The activity of Total-superoxide dismutase(T-SOD) and the level of malondialdehyde (MDA) in cytoplasm were detected bycolorimetry. The expression of UCP2mRNA and protein were determined by RT-PCRand Western blot. Results (1) The fluorescence intensity of ROS in HG group wassignificantly higher than in NG group, while the fluorescence intensity of ROS in V1group and NAC group were both lower than in HG group.(2) Compared with NGgroup, the ΔΨm significantly decreased in HG group (P<0.01), and the ΔΨm in NACand V1group was lower than in HG group (both P<0.01).(3)The activity of T-SOD inHG group was significantly lower than in NG group (P<0.01), while its level of MDAwas significantly higher than in NG group (P<0.01). Compared with HG group, theactivity of T-SOD in V1、V2、V3groups could significantly increase (all P<0.05) and the level of MDA in these groups could significantly decreased in dose-dependentmanner at the same time (all P<0.01).(4)The mRNA and protein expression of UCP2existed in nomal cultured HK-2in NG group. The mRNA expression of UCP2in HGgroup at72hours increased significantly in comparison with NG group (P<0.05). Theexpression of UCP2mRNA and protein in V1、V2、V3groups was significantlydecreased in dose-dependent manner in comparison with HG group (all P<0.01)Conclusion (1)High glucose could stimulate oxidative stress injury in HK-2cells.(2)1,25-(OH)2VitD3could reduce the ΔΨm, the production of ROS, and regulate theexpression of UCP2in order to suppress the oxidative stress induced by high glucose.