Mechanisms of Nitrite Mediated Hepatic Preconditioning: Role of the Mitochondria

肝脏缺血/再灌注（I/R）导致的肝功能障碍或衰竭是发病的主要原因 以及接受肝移植手术的患者的死亡率。目前，没有治疗 战略存在，捐助者供需之间日益扩大的差距迫使 尸体或脂肪变性图像的重建，这些图像对I/R非常敏感。因为干预 细胞和器官衰竭的恢复可能需要一个以上的水平， 预处理是近年来研究的一种很有前途的抗I/R损伤的保护策略。因此，我们认为， 缺血预处理或模拟这些作用的药物干预可能具有最大的 改善肝移植和肝脏手术临床结果的潜力。最近，亚硝酸盐通过 一氧化氮（NO）的产生已经显示在肝I/R的情况下具有细胞保护作用， 这表明亚硝酸盐可能作为NO的生物储存储备，在组织中发挥关键作用 防止缺血性损伤。此外，我们实验室的初步数据表明， 在I/R前24小时注射亚硝酸盐可以保护肝脏免受损伤。此外，亚硝酸盐预处理被发现， 减弱I/R诱导的线粒体呼吸抑制。因此，本提案的目的 研究线粒体在亚硝酸盐介导的肝脏预适应中的作用， 假设亚硝酸盐预处理保留了线粒体的结构和功能 在肝I/R之后，从而减轻肝细胞损伤。拟议研究的基本原理是 确定亚硝酸盐预适应的机制可能为扩大临床应用提供基础。 适用于面临肝移植或肝脏手术的患者。所以，我们计划测试我们的中心假设 并通过两个具体目标实现拟议研究的目标。具体目标1将进一步 通过评估线粒体的保护作用来扩展我们的初步数据， 线粒体基质体积，线粒体膜电位，线粒体钙摄取， 不同复合物的线粒体呼吸，ATP产生，氧化还原电位，ROS产生， 最后，细胞色素C从线粒体释放到细胞质中。具体目标2将采取步骤 通过检测线粒体的作用， K+通道。我们认为，亚硝酸盐，通过其转化为NO，保护线粒体免受 随后的I/R损伤通过介导线粒体K+通道mKATP和mKCa的开放， PKC和PKA的分子信号传导。

Liver dysfunction or failure, as a result of hepatic ischemia/reperfusion (I/R), is a leading cause of morbidity and mortality for patients who have undergone liver transplantation surgery. Currently, no therapeutic strategy exists and the ever growing gap between supply and demand for donors has forced the considertation of cadaveric or steatotic graphs, which are very susceptible to I/R. Because intervention on more than one level is likely needed to allow for the recovery of cellular and organ failure, the most promising protective strategy against I/R injury explored in the last few years is preconditioning. Therefore, ischemic preconditioning or pharmacological interventions that mimic these effects may have the greatest potential to improve clinical outcome in liver transplantation and liver surgery. Recently, nitrite through the generation of nitric oxide (NO) has been shown to have cytoprotective effects in the setting of hepaticI/R, suggesting that nitrite may serve as a biological storage reserve of NO subsurving a critical function in tissue protection from ischemic injury. Furthermore, preliminary data from our lab indicates that the administration of nitrite 24 hours prior to I/R protects the liver against injury. Moreover, nitrite preconditioning was found to attenuate the l/R-induced suppression of mitochondrial respiration. Therefore, the objective of this proposal is to delve into the role of the mitochondria in nitrite mediated hepatic preconditioning with the central hypothesis being that nitrite preconditioning preserves the structure and function of the mitochondria following hepatic I/R, thereby attenuating hepatocellular injury. The rationale for the proposed research is that identifying the mechanisms of nitrite preconditioning may provide a basis for extending the clinical application to patients facing liver transplantation or liver surgery. So, we plan to test our central hypothesis and accomplish the objective of the proposed study by pursuing two specific aims. Specific aim 1 will further expand on our preliminary data by exploring the-protectionof the mitochondria through the evaluation of mitochondrial matrix volume, mitochondrial membrane potential, mitochondiral uptake of calcium, mitochondrial respiration of the different complexes, ATP production, redox potential, ROS production, and finally the release of cytochrome C from the mitochondria into the cytosol. Specific aim 2 will take a step back and explore the signaling mechanisms of nitrite preconditioning by examining the role of mitochondrial K+ channels. We believe that nitrite, through its conversion to NO, protects the mitochondria from a subsequent I/R injury by mediating the opening of the mitochondrial K+ channels, mKATP and mKCa, via the molecular signaling of PKC and PKA, respectively.