高糖通过血清和糖皮质激素诱导的蛋白激酶1通路促进人系膜细胞合成纤连蛋白

目的研究血清和糖皮质激素诱导的蛋白激酶1(SGK1)在高糖诱导人肾小球系膜细胞(HMC)产生纤连蛋白(FN)中的作用,探讨SGK1在糖尿病肾病(DN)肾小球硬化中的作用机制。方法将带有SGK1显性激活型突变体质粒(PIRES2EGFPS422DSGK1mutant,SD)瞬时转染HMC;同时,设空质粒(PIRES2EGFP,FP)转染组和未转染组(NT)为对照。分别用正常糖(NG,5.5mmol/LD葡萄糖)和高糖(HG,25mmol/LD葡萄糖)刺激8h后,采用RTPCR方法和Western印迹方法来观察SGK1和FN的mRNA及蛋白的表达。结果高糖环境下,转染SD的HMC与转染FP、NT组比较,其SGK1mRNA表达显著性增高(0.704vs0.497,0.491,P<0.01),SGK1蛋白过度活化(1178497vs193875,195597,P<0.01),其FNmRNA和蛋白的表达显著性的增加(0.749vs0.463,0.475,P<0.01;659550vs342354,340428,P<0.01)。结论在糖尿病肾病中,高糖可以通过SGK1介导的信号通路来诱导人肾小球系膜细胞增加合成FN,这种新发现的信号通路,表明SGK1可能参入糖尿病肾病肾小球纤维化的发生。

High glucose stimulates synthesis of fibronectin in human mesangial cells via serum and glucocorticoid induced protein kinase-1 signaling pathway

OBJECTIVE: To investigate the role of serum and glucocorticoid induced kinase-1 (SGK(1)) pathways in fibronectin (FN) synthesis in human mesangial cell (HMC) under high glucose condition and the mechanism by which SGK(1) contributes to glomerulosclerosis in diabetic nephropathy (DN). METHODS: HMCs were cultured and transfected with (P)IRES2-EGFP-(S422D) SGK(1) mutant (SD), plasmid containing SGK(1) dominant activation mutant, or blank plasmid. Non-transfected HMCs were used as control group. Then the HMCs were divided into 6 groups: transfected with SD + high glucose (SD-HG, 25 mmol/L D-glucose), transfected with FP + high glues (FP + HG), non-transfected + high glucose (NT-HG), transfeted with SD + normal glucose (SD-NG, 5.5 mmol/L D-glucose), transfected with FP + normal glues (FP + NG), and non-transfected + normal glucose (NT-NG). Eight hours after the glucose stimulation, RT-PCR was used to examine the SGK(1) mRNA expression and fibronectin (FN). Western blotting was used to detect the fibronectin (FN) protein expression. RESULTS: The SGK(1) mRNA expression of the SD + HG group was 0.709, significantly higher than those of the FP + HG and NT + HP groups (0.497 and 0.491, both P < 0.01). The SGK(1) protein expression of the SD + HG group was 1,178,497, significantly higher than those of the FP + HG and NT + HP groups (193,875 and 195,597 respectively, both P < 0.01). The FN mRNA expression of the SD + HG group was 0.749, significantly higher than those of the FP + HG and NT + HP groups (0.463 and 0.475 respectively, both P < 0.01). The FN protein expression of the SD + HG group was 659,550, significantly higher than those of the FP + HG and NT + HG groups (342,354 and 340,428 respectively, both P < 0.01). There were not significant differences in the expressions of FN mRNA and protein among different NG groups. CONCLUSION: SGK(1) may be involved in the signal transduction leading to the increase of fibronectin production in DN and therefore may play an active part in glomerulosclerosis in DN.