雷米普利对舒张性心力衰竭大鼠心房肌L型钙通道影响.docVIP

雷米普利对舒张性心力衰竭大鼠心房肌L型钙通道影响 　　[摘要] 目的 观察雷米普利对舒张性心力衰竭（DHF）大鼠模型心房肌细胞L型钙通道的影响。 方法 腹主动脉缩窄术建立DHF大鼠模型，随机分为DHF组及雷米普利大、中、小剂量组，另设对照组。全细胞膜片钳检测心房肌细胞L型钙通道电流（ICa-L）；RT-PCR检测L型钙通道上Cav1.2 mRNA的表达；Western blot检测钙调控相关蛋白LTCC的表达。 结果 与对照组比较，DHF组大鼠心房肌细胞ICa-L电流密度峰值、Cav1.2 mRNA表达及钙调蛋白LTCC的表达明显降低（P　　[关键词] 舒张性心力衰竭；L型钙通道；雷米普利；Cav1.2 　　[中图分类号] R972[文献标识码] A[文章编号] 1674-4721（2014）06（b）-0015-04 　　 　　Influences of ramipril on L-type calcium channel in rats with diastolic heart failure 　　GUO Zuo-bing1,2 SUN Shu-juan1,3▲ 　　1.School of Pharmaceutical Sciences,Shandong University,Jinan 250012,China;2.Department of Pharmacy,Central Hospital of Qilu Petrochemical Hospital Group,Zibo 255400,China;3.Department of Pharmacy,Shandong Qianfoshan Hospital,Jinan 250014,China 　　[Abstract] Objective To observe the influence of ramipril on L-type calcium channel in rats with diastolic heart failure (DHF). Methods The rat model of DHF was established by using abdominal aortic coarctation,and then the rats were randomly divided into DHF group,high-dose,mid-dose and low-dose ramipril groups and control group.L-type calcium channel current (ICa-L) was investigated by using a whole cell patchclamp technique.The expression levels of Cav1.2 mRNA were measured by RT-PCR analysis.The expression levels of LTCC were measured by Wsetern blot analysis. Results Compared with control group,the peak density of atrial ICa-L,the expression levels of Cav1.2 mRNA and the expression levels of LTCC was lower in DHF group (P 　　1.2.2 全细胞膜片钳记录ICa-L 　　 胶原酶解法分离大鼠心房肌细胞[5-6]。取得适合细胞，放入显微镜的细胞池中，贴壁。玻璃电极灌入电极液，电极与细胞表面形成高电阻，完成封接破膜补偿快慢电容。MODE置向VC后进行ICa-L记录。 　　1.2.3 RT-PCR检测Cav1.2 mRNA的表达 　　1.2.3.1 提取RNA剪取大鼠心房组织，称取75 mg，放入Trizol匀浆，离心，取上清液，氯仿充分混匀，离心20 min，取上清液移，异丙醇沉淀，再离心弃上清，后将RNA用1 ml 75%乙醇洗涤，离心，除乙醇后加0.01%焦碳酸二乙酯，温浴15 min，后在A260/A280下定量RNA。 　　1.2.3.2 RT-PCR反应混合好RT反应及PCR反应体系，加水至10 μl混匀，置于PCR仪上，94℃变性5 min，后扩增30个循环，72℃延伸5 min。凝胶电泳，染色。 　　1.2.4 Western blot检测LTCC的表达 　　提取心房肌细胞膜蛋白，取大鼠左心房组织120 mg，剪碎。后加入裂解液和蛋白酶抑制剂。于匀浆器中打碎，后4℃，离心5 min。取上清液置于EP管，4℃离心。测定膜蛋白浓度。SDS电