1,25-二羟维生素D3对高糖诱导肾小管上皮细胞解耦联蛋白2表达及氧化应激的影响

摘要

体外培养的HK-2细胞分为正常对照组(NG组,5.5 mmol/L D-葡萄糖)、高糖处理组(HG组,30 mmol/L D-葡萄糖)、高渗对照组(MG组,5.5 mmol/L D-葡萄糖+24.5 mmol/L D-甘露醇)、高糖培养液中又加入不同浓度的1,25-二羟维生素D3[1,25-(OH)2D3]组(V1～V3组)、N-乙酰半胱氨酸药效对照组(NAC组,30 mmoL/L D-葡萄糖+1.0 mmol/L N-乙酰半胱氨酸)和无水乙醇溶剂对照组(SG组,30 mmol/LD-葡萄糖+6.86×10-2 mol/L乙醇).检测细胞活性氧簇荧光强度、线粒体膜电位、解耦联蛋白2(UCP2)mRNA、蛋白表达、总超氧化物歧化酶(SOD)活力和丙二醛水平.结果HG组细胞的线粒体膜电位明显低于NG组(P＜0.01),1,25-(OH)2D3处理后细胞的线粒体膜电位与HG组比较显著下降(均P＜0.01)；HG组总SOD活力显著低于NG组(P＜0.01),丙二醛含量明显高于NG组(P＜0.01),而1,25-(OH)2D3处理后细胞总SOD活力较HG组显著增高(P＜0.05),丙二醛水平显著低于HG组(P＜0.01)；HG组UCP2 mRNA与蛋白表达显著高于NG组(P＜0.05),而1,25-(OH)2D3处理后细胞UCP2的表达与HG组比较显著降低(P＜0.01).结果表明高糖可诱导体外培养的HK-2细胞氧化应激损伤；1,25-(OH)2D3可能通过降低线粒体膜电位、活性氧簇生成及调节胞内UCP2表达抑制高糖诱导的氧化应激反应.%[Summary] The HK-2 cells with different culture media were divided into normal glucose group (NG group,5.5 mmol/L D-glucose) ; high glucose group (HG group,30 mmol/L D-glucose) ; mannitol group (MG group,5.5mmol/L D-glucose+24.5 mmol/L mannitol) ; 1,25-dihydroxyvitamin D3 [1,25-(OH)2D3] groups (V1-V3 group)which were exposed to medium containing 30 mmol/L D-glucose and different concentrations of 1,25-(OH)2D3 ;Nethyl-cysteim control group (NAC group,30 mmoL/L D-glucose + 1.0 mmol/L N-Nethyl-cysteim) ; and ethanol control group(SG group,30 mmol/L D-glucose+6.86 × 10-2 mol/L ethanol).The level of intracellular reactive oxygen species,mitochondrial membrane potential,activity of total-superoxide dismutase (T-SOD),level of malondialdehyde,expression of UCP2 mRNA and protein in HK-2 cells were detected.Compared with NG group,the mitochondrial membrane potential significantly decreased in HG group (P＜0.01),and the mitochondrial membrane potential in V group was lower than that in HG group(P＜0.01).The activity ofT-SOD in HG group was significantly lower than that in NG group(P＜0.01),while its level of malondialdehyde was significantly higher than that in NG group(P＜0.01).Compared with HG group,the activity of T-SOD in V groups was significantly increased (P＜0.05)and the level of malondialdehyde in these groups significantly decreased (P＜0.01).The mRNA expression of UCP2 in HG group was increased significantly in comparison with NG group (P ＜ 0.05) and the expression in V groups was significantly decreased in comparison with HG group (P＜0.01).The results suggest that 1,25-(OH)2D3 could reduce the mitochondrial membrane potential,the production of reactive oxygen species,and regulate the expression of UCP2 in order to suppress the oxidative stress induced by high glucose.