油茶皂苷上调AE3表达及其通过AE3调控缺血/再灌注心肌细胞内Cl-外流与线粒体保护机制研究

阴离子交换蛋白3（anion exchanger 3, AE3）属于SLC4蛋白家族，具有可逆性电中性Cl-/HCO3-交换活性，在调节细胞内Cl-的浓度等方面起着重要作用。最近，我们在前一基金资助下证实，SQS预处理能上调AE3蛋白的表达，并通过AE3蛋白促使缺血/再灌注（I/R）心肌细胞内过载的Cl-外排，对抗I/R所诱发的氧化应激、钙超载和细胞凋亡。在此工作基础上，本研究拟在细胞和整体水平，借助信号通路抑制剂，利用一系列分子生物学实验手段，从NO/PKCε/Ras/ERK1/2信号通路的变化以及AE3蛋白Ser67位点磷酸化程度改变的角度，进一步阐明SQS对AE3蛋白表达及其促Cl-/HCO3-反向交换的调控机制；同时，从细胞内Cl-动态变化及mPTP功能改变入手深入全面地揭示SQS通过AE3蛋白发挥心肌保护效应的新靶点和新机制。该研究的完成将为进一步研发SQS提供新的理论和实验依据

Anion exchanger 3 (AE3), a member of the Solute Carrier 4 (SLC4) protein family, mediates the reversible electroneutral exchange of Cl- for HCO3- across the plasma membrane and exerts a central role in the regulation of intracellular Cl- concentration, intracellular pH, and cardiovascular homeostasis. Our previous work shown that Sasanquasaponin (SQS) can inhibit ischemia/reperfusion-induced elevation of intracellular Cl- concentration ([Cl-]i) followed by the attenuation of Ca2+ overload, ROS production, and cell apoptosis by up-regulating AE3 protein expression. In the present study, by using a variety of signal pathway inhibitor and molecular biology methods, we will futher analyse the regulatory mechanisms by which SQS up-regulates AE3 expression and increases Cl-/HCO3- exchange activity of AE3 protein from NO/PKCε/Ras/ERK1/2 signaling pathway and phosphorylation level of serine 67 of AE3 protein at the cellular and whole-animal levels. Moreover, this study will further explore novel targets and mechanisms of AE3-mediated SQS cardioprotection from aspect of mPTP opening and chloridion transport. This study will provide new experimental and theoretical basis for further developing SQS.