降钙素基因相关肽及IP3信号通路介导1型糖尿病大鼠离体心脏缺血预适应保护作用

Role of calcitonin gene-related peptide and IP3 pathway in ischemic preconditioning in isolated perfused hearts of rats with type 1 diabetes mellitus

目的研究降钙素基因相关肽(CGRP)及IP3信号通路在1型糖尿病大鼠离体心脏缺血预适应(IPC)中的作用,探讨1型糖尿病大鼠心脏IPC保护作用减弱的机制。方法将80只造模成功的SD大鼠随机分为1型糖尿病4周(D-4w)组(n=40)和8周(D-8w)组(n=40),并进一步将两组各自随机分为模型对照(D-Cont)组、1型糖尿病缺血再灌注(IR)组、IPC组、CGRP(IPC+CGRP)组和IP3抑制剂wortmanin(IPC+WMN)组5个亚组;另取16只大鼠作为正常对照(N-Cont)组。采用Langendorff方法建立离体心脏体外灌流模型,灌流期间外源性给予CGRP或wortmanin(WMN)。采用多道生物信号分析系统监测各组大鼠离体心脏左室功能,记录冠脉流量,ELISA法检测大鼠血清CGRP含量;生化法测定冠脉流出液中乳酸脱氢酶(LDH)及肌酸激酶(CK)的活性;NBT染色法测量心肌梗死面积;TUNEL法检测心肌细胞凋亡情况。结果与N-Cont组大鼠相比,1型糖尿病大鼠血清CGRP含量随着时间进行性降低,且心脏基础左心功能降低,冠脉流出液中LDH和CK活性增加,心肌梗死面积和心肌细胞凋亡指数增加(P<0.05);IR组与D-Cont组相比,左心功能更为降低,心肌损伤明显;IPC可改善D-4w IR心肌损伤情况,而对D-8w IR损伤无保护作用;IPC+CGRP组与IPC组相比,其左心功能明显改善;IPC+WMN组可阻断IPC对D-4w组大鼠心肌保护作用。结论CGRP及IP3信号通路参与1型糖尿病大鼠离体心脏的IPC保护作用。

Objective To investigate the role of calcitonin gene-related peptide （CGRP） and IP3 signal pathway in ischemic preconditioning （IPC） in the isolated perfused hearts of rats with type 1 diabetes mellitus. Methods Type 1 diabetes mellitus rat models were established in 80 SD rats, and were randomly divided into 4 week （D-4w） group and 8 week （D-8w） group. These two groups were randomly subdivided into model control （D-Cont） group, type 1 diabetes mellitus ischemia-reperfusion （IR） group, IPC group, CGRP （ IPC ＋ CGRP） group and IP3 inhibitor wortmanin （ IPC ＋ WMN） group. Another 16 rats were served as normal control （N-Cont） group. In vitro perfusion models of isolated hearts were established by Langendorff methods, and CGRP or wortmanin （WMN） were administered during perfusion. The left ventricle function of isolated heart in each group was monitored by muhiehannel biosignal analysis system, and coronary artery flow was recorded. The serum CGRP levels were detected by ELISA. The activity of lactate dehydrogenase （LDH） and ereatine kinase （CK） in effluent of coronary artery was detected by biochemical method. The size of myocardial infarction was determined by NBT staining, and apoptosis of cadiocytes was detected by TUNEL method. Results Compared with N- Cont group, the CGRP level in serum of rats with type 1 diabetes mellitus decreased with time, the basic left ventricle function decreased, while the activity of LDH and CK in effluent of coronary artery, size of myocardial infarction and cardiomyocyte apoptosis index increased （P 〈 0. 05）. Compared with N-Cont group, the left ventricle function was significantly lower in IR group, and more severe myocardial damage was observed. IPC improved myocardial damage of D-4w IR group, while had no protection on D-Sw IR group. Compared with IPC group, the left ventricle function was significantly improved in IPC ＋ cGRP group. IPC ＋ WMN blocked the myocardial protection of D-4w group from IPC. Conclusion CGRP and IP3 signal pathway are involved in the protection provided by IPC in isolated hearts of rats with type 1 diabetes mellitus.