氟伐他汀对醛固酮诱导的系膜细胞SGK1及CTGF表达的影响

目的:观察氟伐他汀对醛固酮(Ald)诱导的大鼠肾小球系膜细胞(HBZY-1)血清与糖皮质激素诱导蛋白激酶1(SGK1)和结缔组织生长因子(CTGF)表达的影响。方法:以大鼠系膜细胞为实验对象,分为:(1)对照组;(2)不同浓度及时间Ald组;(3)Ald(10-7mol/L)+SPI(安体舒通10-9mol/L)组;(4)Ald(10-7mol/L)+LY294002(PI3K抑制剂20μmol/L)组;(5)Ald(10-7mol/L)+SB203580(P38MAPK抑制剂20μmol/L);(6)Ald(10-7mol/L)+F(氟伐他汀10-7、10-6、10-5mol/L)组;(7)Ald(10-7mol/L)+F(氟伐他汀10-5mol/L)+MVA(甲羟戊酸10-4mol/L)组;(8)Ald(10-7mol/L)+F(氟伐他汀10-5mol/L)+FPP(法尼酯焦磷酸10-5mol/L)组;(9)Ald(10-7mol/L)+F(氟伐他汀10-5mol/L)+GGPP(焦磷酸牛儿基牛儿酯10-5mol/L)组。Western印迹方法检测SGK1、CTGF蛋白表达水平。酶联免疫吸附实验(ELISA)检测细胞培养液中细胞外基质成分纤维连接蛋白(FN)、单核细胞趋化因子(MCP-1)和细胞间黏附分子-1(ICAM-1)蛋白含量。结果:Ald呈剂量依赖性上调SGK1和CTGF蛋白表达(P0.05)。醛固酮刺激6h即可增加SGK1蛋白表达(P0.05),12h达高峰,CTGF蛋白水平呈时间依赖性增加。安体舒通和PI3K特异性抑制剂LY294002可阻断Ald对SGK1的激活作用(P0.01)。氟伐他汀可呈剂量依赖性下调Ald诱导的系膜细胞内SGK1和CTGF蛋白表达,以及其下游趋化因子MCP-1和ICAM-1表达(P0.05)。甲羟戊酸和GGPP可逆转氟伐他汀的作用(P0.05)。结论:醛固酮可通过抑制盐皮质激素受体(MR)和PI3K信号转导通路诱导大鼠系膜细胞内SGK1和CTGF蛋白的表达,氟伐他汀可通过SGK1信号转导通路缓解Ald的致纤维化作用。甲羟戊酸可部分逆转氟伐他汀的作用,可能与甲羟戊酸通路中的Rho蛋白活化有关。

Effect of fluvastatin on expression of SGK1 and CTGF induced by aldosterone in rat mesangial cells

AIM: To investigate the effect of fluvastatin on the expression of serum and glucocorticoid inducible kinase 1 (SGK1) and connective tissue growth factor (CTGF) induced by aldosterone (Ald) in rat mesangial cells (GMCs). METHODS: GMCs were divided into (1) control group; (2) aldosterone group with different concentrations and times; (3) Ald (10-7 mol/L)+spironolactone (10-9mol/L) group; (4) Ald (10-7mol/L)+LY294002 (20 μmol/L) group; (5) Ald (10-7mol/L) +SB203580 (20 mmol/L) group; (6) the group of Ald (10-7mol/L)+ fluvastatin at different concentrations (10-7, 10-6, 10-5 mol/L); (7) Ald (10-7mol/L) +fluvastatin (10-5mol/L)+mevalonate (10-4 mol/L) group; (8) Ald (10-7mol/L) +fluvastatin (10-5mol/L)+FPP (farnesyl pyrophosphate, 10-4 mol/L) group; (9) Ald (10-7mol/L) +fluvastatin (10-5 mol/L) +GGPP (geranylgerany pyrophosphate, 10-4 mol/L) group. The protein levels of SGK1 and CTGF were determined by Western blotting. The levels of fibronection (FN), monocyte chemoattractant protein-1 (MCP-1) and intercellular adhesion molecule-1 (ICAM-1) in the supernatants were determined by enzyme-linked immunosorbant assay (ELISA). RESULTS: Aldosterone stimulated the protein expression of SGK1 and CTGF in cultured mesangial cells in a dose-dependent manner (P<0.01). SGK1 expression was increased at as early as 6 h (P<0.05), peaked at 12 h after aldosterone treatment (P<0.01). This stimulatory effect of aldosterone on SGK1 and CTGF was blocked by mineralocorticoid receptor (MR) inhibitor and LY294002 (a specific inhibitor of PI3K). Incubation of cells with fluvastatin significantly inhibited the aldosterone-induced the activations of SGK1 and CTGF, and the expression of MCP-1 and ICAM was in a dose-dependent manner (P<0.05). Exogenous mevalonate prevented the effect of fluvastatin on SGK1 expression in GMCs. CONCLUSION: Fluvastatin reduces aldosterone-induced SGK1 expression via mineralocorticoid receptor and PI3K pathway in rat mesangial cells. Such effect of flurastatin is partly blocked by mevalonate.