PKC/NADPH氧化应激途径对大鼠心脏微血管内皮细胞eNOS脱偶联的影响  

Effects of PKC/NADPH oxidase on AGEs-induced eNOS uncoupling in rat cardiac microvascular endothelial cells

作　　者：成永霞[1] 刘贵波[2] 颜彬[1] 冯玉宽[2] 郭素芬[1] 王宏伟[1] 杨向红[3] 

机构地区：[1]牡丹江医学院病理教研室,黑龙江牡丹江157011 [2]牡丹江医学院解剖教研室,黑龙江牡丹江157011 [3]中国医科大学附属盛京医院病理科,辽宁沈阳100011

出　　处：《基础医学与临床》2013年第1期82-87,共6页Basic and Clinical Medicine

基　　金：教育部科学技术研究重点项目(211050);黑龙江省青年科学基金(QC2010040)

摘　　要：目的观察体外原代培养大鼠心脏微血管内皮细胞内PKC/NADPH氧化应激途径在eNOS脱偶联中的作用。方法用牛血清白蛋白糖基化终末产物(BSA-AGEs)(100 mg/L)与LY33531(PKC抑制剂)和DPI(NADPH抑制剂)分别作用大鼠心脏微血管内皮细胞(24 h),HPLC法检测BH4,试剂盒检测NO和O2-生成,免疫组化检测eNOS蛋白表达,Western blot法检测P47phox蛋白表达。结果随着LY33531和DPI浓度(5、10和20μmol/L)的增加,eNOS脱偶联状态减轻:NO生成逐渐增加(90.7±0.3～122.6±0.3,160.6±0.6,P<0.05),而O2-生成逐渐减少(P<0.05),eNOS表达逐渐减少(126.1±3.5～112.6±1.7,114.4±1.8,P<0.05),而BH4含量逐渐增加(P<0.05)。同时,ROS表达和P47phox表达减少(P<0.05)。结论 NADPH氧化应激途径可能参与AGEs诱导的心脏微血管内皮细胞eNOS脱偶联的发生。而AGEs诱导的心脏微血管内皮细胞内NADPH氧化应激的发生是依赖PKC激活的。Objective To observe the effects of PKC/NADPH oxidase on AGEs-induced eNOS uncoupling in rat cardiac microvascular endothelial cells.Methods To culture the cardiac microvascular endothelial cells in vitro.AGEs（100 mg/L）,DPI and LY33531（2.5,5 and 10 μmol/L）was respectively added and resumed incubation for 24 hrs.BH4,NO,O2-generation,eNOS,ROS and P47phox protein expression were examined.Results As LY33531 and DPI concentration was increased,production of NO was noted increased gradually（90.7±0.3~122.6±0.3,160.6±0.6,P0.05）,while production of O2-was decreased gradually（P0.05）,BH4 expression was increased（P0.05）,eNOS expression was gradually decreased（126.1±3.5~112.6±1.7,114.4±1.7,P0.05）.Meanwhile,P47phox protein expression and ROS decreased gradually（P0.05）.ConclusionsAGEs may trigger the activation of PKC/NADPH oxidative stress pathway inducing intracellular eNOS uncoupling in cardiac microvascular endothelial cells.

关 键 词：糖基化终末产物 心脏微血管内皮细胞 一氧化氮 内皮型一氧化氮合酶 PKC NADPH LY33531 DPI 

分 类 号：R363[医药卫生—病理学]