瞬时受体电位C1通道在糖尿病大鼠冠状动脉平滑肌细胞上的表达及对细胞内钙离子浓度的影响  

The expression of classical transient receptor potential canonical channel-1 in coronary smooth muscle cells in diabetic rats and its effects on cytosolic calcium concentrations

作　　者：孙曼青[1] 汤徐 钱玲玲[1] 党时鹏[1] 吴莹[1] 杨承健[2] 肖春晖[3] 刘晓宇[1] 夏大云[1] 柴强[4] 王如兴[1] 

机构地区：[1]南京医科大学附属无锡市人民医院心内科,214023 [2]南京医科大学附属无锡市第二人民医院心内科 [3]杭州师范大学附属医院干部科 [4]山东省医学科学院基础医学研究所心血管病研究室

出　　处：《中华糖尿病杂志》2017年第5期298-303,共6页CHINESE JOURNAL OF DIABETES MELLITUS

基　　金：国家自然科学基金（81370303、81400297、81500249、81500323）;江苏省自然科学基金（BK2015110）;江苏省人事厅“六大人才高峰”第七批高层次项目（006）;江苏省医学重点人才资助项目（RC201134）

摘　　要：目的 探讨瞬时受体电位C1通道（TRPC1）在糖尿病大鼠冠状动脉平滑肌细胞中的表达变化及对糖尿病冠状动脉功能影响的可能机制.方法 选用8~12周龄、体重（200±20）g的健康雄性SD大鼠160只,采用随机数字表法随机分为正常对照组（80只）和糖尿病组（80只）.采用腹腔链脲佐菌素注射建立1型糖尿病大鼠动物模型;采用酶消化法急性分离正常和糖尿病大鼠冠状动脉平滑肌细胞;采用Western blotting和实时荧光定量PCR分别测定正常组和糖尿病组冠状动脉TRPC1通道蛋白和基因的表达;采用细胞内钙离子荧光成像技术测定正常和糖尿病大鼠冠状动脉平滑肌细胞内钙离子浓度及加入TRPC1通道阻滞剂SKF96365后细胞内钙离子浓度的变化;采用血管张力测定技术测定TRPC1通道阻滞剂SKF96365对糖尿病冠状动脉功能的影响.2组间比较采用t检验.结果 糖尿病组冠状动脉平滑肌细胞上TRPC1通道蛋白表达量是正常组的（1.456±0.081）倍（t=-2.210,P〈0.05）;糖尿病组TRPC1通道基因表达量为正常组的（2.198±0.251）倍（t=-3.864,P〈0.05）;糖尿病组较正常组钙离子浓度变化增加（1.217±0.044比0.869±0.029,t=-6.644,P〈0.05）,正常组加入SKF96365后较加入前钙离子浓度减低（0.067±0.039比0.842±0.020,t=15.726,P〈0.05）,糖尿病组加入SKF96365后较加入前钙离子浓度减低（0.195±0.028比1.217±0.043,t=-19.807,P〈0.05）;正常组和糖尿病组冠状动脉在ET-1收缩血管后,加入10μmol/L TRPC1通道抑制剂SKF96365,糖尿病组较正常组冠状动脉舒张的百分比明显升高[（91.6±2.6）%比（69.2±6.0）%,t=-3.529,P〈0.05].结论 糖尿病时冠状动脉平滑肌细胞上TRPC1通道表达增加,细胞内钙离子浓度增加,导致血管收缩和功能紊乱,这可能是糖尿病患者易发生冠状动脉病变的机制之一.Objective To investigate the expression of transient receptor potential canonical channel-1 （TRPC1） in coronary smooth muscle cells （CSMCs） in diabetic rats and the possible mechanisms of TRPC1 channels on diabetic coronary function. Methods A total of 160 healthy male Sprague-Dawley （SD） rats with 8-12 weeks of age and weight （200±20） g were randomly divided into normal control group （80 rats） and diabetic group （80 rats）. Type 1 diabetes was established by intraperitoneally injection of streptozotocin. Normal and diabetic CSMCs from Sprague-Dawley rats were isolated by enzyme digestion. The protein and gene expressions of TRPC1 channels in normal and diabetic CSMCs were determined by Western blotting and real-time fluorescent quantitative PCR technique, respectively. Cytosolic calcium concentrations before and after incubating with TRPC1 channels blocker SKF96365 in normal and diabetic CSMCs were examined by recording the changes of fluorescence intensity ratios. Vascular reactivity after appliance of SKF96365 on coronary arteries in normal and diabetic rats was detected by using vascular tension measurement. The experimental data were analyzed by t test between two groups. Results The protein expressions of TRPC1 channels in diabetic CSMCs were （1.456 ± 0.081） times of the normal groups （t=-2.210, P〈0.05）. The gene expressions of TRPC1 channels in diabetic CSMCs were （2.198±0.251） times of the normal group（t=-3.864, P〈0.05）. ΔRatios of normal and diabetic CSMCs were 0.869 ± 0.029 and （1.217 ± 0.043, t=-6.644, P〈0.05）, respectively. ΔRatios were 0.842 ± 0.020 and 0.067 ± 0.039 before and after the appliance of TRPC1 channels blocker SKF96365 in normal CSMCs （t=15.726,P〈0.05）. ΔRatios were 1.217±0.043 and 0.195±0.028 before and after appliance of SKF96365 in diabetic CSMCs （t=-19.807, P〈0.05）. When the vascular tension increased to the maximum after application of ET-1, then coronary arteries were exposed to 10μmol/L SKF96365, and the dil

关 键 词：瞬时受体电位C1通道 糖尿病 冠状动脉 平滑肌细胞 细胞内钙离子 

分 类 号：R36[医药卫生—病理学]