【摘要】 目的观察磷脂酰肌醇-3激酶(PI3K)-蛋白质丝氨酸/苏氨酸激酶(AKt)-内皮型一氧化氮合酶(e NOS)信号转导通路在硫化氢(H2S)抑制内皮素-1(endothelin-1,ET-1)诱导心肌肥大过程中的作用。方法体外培养原代心肌细胞,将其随机分为6组,每组4孔,1对照组:加入等体积无血清的DMEM培养基;2肥大(ET-1)组:加入终浓度为10-8 mol/L的ET-1;剩余4组为实验组,各组分别加入不同终浓度的H2S供体-Na HS:310-15 M Na HS组:加入10-15 mol/L Na HS+10-8 mol/l ET-1;410-14 M Na HS组:加入10-14 mol/L Na HS+10-8 mol/L ET-1;510-13 M Na HS组:加入10-13 mol/L Na HS+10-8 mol/L ET-1;610-12 M Na HS组:加入10-12 mol/L Na HS+10-8 mol/L ET-1。上述各组药物分别刺激24 h后测定心肌细胞表面积、细胞总蛋白含量、培养液NO含量,RT-PCR检测心肌细胞心房利钠肽(atrial natriuretic peptide,ANP)、脑钠肽(B-type natriuretic peptide,BNP)、磷脂酰肌醇-3激酶(phosphatidylinositol-3-kinase,PI3K)、蛋白激酶B(protein kinase B,PKB/AKt)、e NOS m RNA水平,Western Blot技术检测总AKt和磷酸化AKt蛋白表达含量。结果肥大(ET-1)组的心肌细胞表面积(1933.80±143.06)和细胞总蛋白含量(367.51±25.9)均高于对照组(787.27±107.66,218.55±21.28,P<0.05),ANP及BNP m RNA的表达量也明显增加(P<0.05),但PI3K、AKt、e NOS m RNA表达水平,磷酸化AKt程度和NO的释放量(4.60±0.73)低于对照组(8.63±0.30,P<0.05),各实验组给予不同浓度Na HS刺激后能够浓度依赖性的抑制这种肥大效应(P<0.05),同时上调了PI3K/AKt/e NOS通路各信号分子的表达量(P<0.05)。结论 H2S对ET-1诱导的心肌肥大有一定的抑制作用,这种作用可能与激活PI3K-AKt-e NOS信号通路有关。更多还原

【Abstract】 Objective To observe the role of PI3K/AKt/e NOS signal pathway in course of H2 S inhibiting myocardial hypertrophy induced by endothelin-1(ET-1). Methods The neonatal myocardial cells were cultured in vitro and randomly divided into 6 groups including 1control group treated with serum-free DMEM medium, 2hypertrophy group, with 10-8 mol/l ET-1, 310-15 M Na HS group, with 10-15 mol/l Na HS+10-8 mol/l ET-1, 410-14 M Na HS group, with 10-14 mol/l Na HS+10-8 mol/l ET-1, 510-13 M Na HS group, with 10-13 mol/l Na HS+10-8 mol/l ET-1, and 610-12 M Na HS group, with 10-12 mol/l Na HS+10-8 mol/l ET-1. After 24 h, cell surface area and total protein content of myocardial cells and nitric oxide(NO) content in nutrient solution were detected in all groups. The m RNA levels of atrial natriuretic peptide(ANP), B-type natriuretic peptide(BNP), phosphatidylinositol-3-kinase(PI3K), protein kinase B(PKB/AKt) and endothelial nitric oxide synthase(e NOS) were detected by using RT-PCR. The expressions of total AKt and phosphorylated AKt were detected by using Western blot test. Results In hypertrophy group, the surface area(1933.80±143.06) and total protein content(367.51±25.9) of myocardial cells were higher than those in control group(787.27±107.66, 218.55±21.28, P<0.05), and m RNA expressions of ANP and BNP increased significantly(P<0.05). The m RNA expressions of PI3 K, AKt and e NOS and phosphorylated AKt degree decreased in hypertrophy group, and NO release quantity were lower in hypertrophy group(4.60±0.73) than that in control group(8.63±0.30, P<0.05). In all M Na HS groups, the hypertrophy effect was inhibited(P<0.05) showed a dose-dependent manner after Na HS treating, and the expressions of all signal molecules in PI3K/AKt/e NOS signal pathway were up-regulated(P<0.05). ConclusionH2 S had some inhibitory effect on myocardial hypertrophy induced by ET-1, which is related to the activation ofPI3K/AKt/e NOS signal pathway.更多还原