CAJ | 학술논문

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（eNOS）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-NaHS：③10-15 M NaHS组：加入10-15 mol/L NaHS+10-8 mol/l ET-1；④10-14 M NaHS组：加入10-14 mol/L NaHS+10-8 mol/L ET-1；⑤10-13 M NaHS组：加入10-13 mol/L NaHS+10-8 mol/L ET-1；⑥10-12 M NaHS组：加入10-12 mol/L NaHS+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）、eNOS mRNA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 mRNA的表达量也明显增加（P＜0.05），但PI3K、AKt、eNOS mRNA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NaHS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/eNOS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-eNOS信号通路有关。
목적：관찰린지선기순-3격매（PI3K）-단백질사안산/소안산격매（AKt）-내피형일양화담합매（eNOS）신호전도통로재류화경（H2S）억제내피소-1（endothelin-1，ET-1）유도심기비대과정중적작용。방법체외배양원대심기세포，장기수궤분위6조，매조4공，①대조조：가입등체적무혈청적DMEM배양기；②비대（ET-1）조：가입종농도위10-8 mol/L적ET-1；잉여4조위실험조，각조분별가입불동종농도적H2S공체-NaHS：③10-15 M NaHS조：가입10-15 mol/L NaHS+10-8 mol/l ET-1；④10-14 M NaHS조：가입10-14 mol/L NaHS+10-8 mol/L ET-1；⑤10-13 M NaHS조：가입10-13 mol/L NaHS+10-8 mol/L ET-1；⑥10-12 M NaHS조：가입10-12 mol/L NaHS+10-8 mol/L ET-1。상술각조약물분별자격24 h후측정심기세포표면적、세포총단백함량、배양액NO함량，RT-PCR검측심기세포심방리납태（atrial natriuretic peptide，ANP）、뇌납태（B-type natriuretic peptide，BNP）、린지선기순-3격매（phosphatidylinositol-3-kinase，PI3K）、단백격매B（protein kinase B，PKB/AKt）、eNOS mRNA수평，Western Blot기술검측총AKt화린산화AKt단백표체함량。결과비대（ET-1）조적심기세포표면적（1933.80±143.06）화세포총단백함량（367.51±25.9）균고우대조조（787.27±107.66，218.55±21.28，P＜0.05），ANP급BNP mRNA적표체량야명현증가（P＜0.05），단PI3K、AKt、eNOS mRNA표체수평，린산화AKt정도화NO적석방량（4.60±0.73）저우대조조（8.63±0.30，P＜0.05），각실험조급여불동농도NaHS자격후능구농도의뢰성적억제저충비대효응（P＜0.05），동시상조료PI3K/AKt/eNOS통로각신호분자적표체량（P＜0.05）。결론H2S대ET-1유도적심기비대유일정적억제작용，저충작용가능여격활PI3K-AKt-eNOS신호통로유관。
Objective To observe the role of PI3K/AKt/eNOS signal pathway in course of H2S 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, ② hypertrophy group, with 10-8 mol/l ET-1, ③10-15 M NaHS group, with 10-15 mol/l NaHS+10-8 mol/l ET-1,④10-14 M NaHS group, with 10-14 mol/l NaHS+10-8 mol/l ET-1, ⑤10-13 M NaHS group, with 10-13 mol/l NaHS+10-8 mol/l ET-1, and ⑥10-12 M NaHS group, with 10-12 mol/l NaHS+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 mRNA 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 (eNOS) 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 mRNA expressions of ANP and BNP increased significantly (P<0.05). The mRNA expressions of PI3K, AKt and eNOS 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 NaHS groups, the hypertrophy effect was inhibited (P<0.05) showed a dose-dependent manner after NaHS treating, and the expressions of all signal molecules in PI3K/AKt/eNOS signal pathway were up-regulated (P<0.05). Conclusion H2S had some inhibitory effect on myocardial hypertrophy induced by ET-1, which is related to the activation of PI3K/AKt/eNOS signal pathway.