蛋白激酶信号通路及心肌缝隙连接蛋白CX43改变在阿片药物预处理中的作用

Studies suggest that the cardio protection effect is achieved through protein kinase (PKC). Cardiac gap junction protein Cx43 is a target protein for a variety of protein kinase. Cx43 expression is related to the arrhythmia. Whether the effect of expression of Cx43 is involved to different subtypes of protein kinase phosphorylation levels is currently a urgent project needs to be proofed. This project is carried out on the basis of preliminary studies. According to the study result of the mechanism of action, the intersection is existed between the drug pretreatment and ischemic preconditioning, the rat heart and neonatal rat myocardial ischemia-reperfusion injury model were adopted. The classic morphine and a new opioid pretreatment drug- sufentanil were applied as interventions. In different status of subtypes of phosphorylation, protein kinase(PKC) and mitogen-activated protein kinase(MAPK) MAPK were systematically observed. In order to reveal the opioid drugs with subtypes of different kinase and regularity of cardiac gap junction protein Cx43 in variation，Connexin Cx43 was used as a target, using western-blotting immunoblotting technique and laser scanning confocal microscopy immunofluorescence detect rat cardiac Cx43 protein expression and Ser368 phosphorylation levels. Revealing the protein kinase activation during myocardial opioid pretreatment and Cx43 expression and Ser368 phosphorylation between the signal path, from the cellular and molecular levels to reflect the experimental effects, providing a theoretical foundation for the reasonable and effective clinical application of opioids play a myocardial protective effect and to provide new ways of thinking for perioperative myocardial protection..

研究认为阿片参与的心肌保护作用是通过蛋白激酶PKC途径实现的，而心肌缝隙连接蛋白Cx43是各种蛋白激酶的靶蛋白，Cx43的表达与心律失常发生有关。阿片药物对Cx43表达的影响是否涉及了蛋白激酶不同亚型磷酸化水平，是目前亟待解决的一项课题。本项目是在阿片药物对缺血再灌注心肌具有保护作用的研究基础上，采用在体大鼠心肌及乳鼠心肌细胞缺血再灌注损伤模型，运用经典的吗啡和新型阿片药物舒芬太尼作为干预，系统观察了蛋白激酶PKC、MAPK不同亚型磷酸化状态，以心肌缝隙连接蛋白Cx43作为靶点利用免疫印迹western-blotting 技术，激光共聚焦显微镜免疫荧光法检测大鼠心肌Cx43的蛋白表达及Ser368磷酸化水平，揭示阿片药物与蛋白激酶C不同亚型的关系及心肌缝隙连接蛋白Cx43的变化规律，更深入的比较其对心律失常的影响，为临床合理有效的应用阿片药物发挥心肌保护作用提供理论依据。

研究认为阿片参与的心肌保护作用是通过蛋白激酶PKC途径实现的，而心肌缝隙连接蛋白Cx43是各种蛋白激酶的靶蛋白，Cx43的表达与心律失常发生有关。阿片药物对Cx43表达的影响是否涉及了蛋白激酶不同亚型磷酸化水平，是目前亟待解决的一项课题。本项目是在阿片药物对缺血再灌注心肌具有保护作用的研究基础上，采用在体大鼠心肌及乳鼠心肌细胞缺血再灌注损伤模型，运用经典的吗啡和新型阿片药物舒芬太尼作为干预，系统观察了蛋白激酶PKC、MAPK不同亚型磷酸化状态，以心肌缝隙连接蛋白Cx43作为靶点利用免疫印迹western-blotting 技术，激光共聚焦显微镜免疫荧光法检测大鼠心肌Cx43的蛋白表达及Ser368磷酸化水平，揭示阿片药物与蛋白激酶C不同亚型的关系及心肌缝隙连接蛋白Cx43的变化规律，更深入的比较其对心律失常的影响，为临床合理有效的应用阿片药物发挥心肌保护作用提供理论依据。通过研究发现，等效剂量吗啡和舒芬太尼预处理后，心肌Cx43蛋白表达较未做预处理的心肌细胞明显提升；但是给予蛋白激酶抑制剂后，Cx43蛋白较单纯使用吗啡、舒芬太尼预处理有所下降。这就提示吗啡、舒芬太尼具有很强的Cx43蛋白保护作用；这种作用与PKC、MAPK信号通路有关。通过本次研究是我们对阿片类药物的保护作用有了较为深刻的认识，对临床积极开展以阿片药物为核心的围术期心肌保护相关策略，奠定了坚实的理论基础。