5-HT Stimulation of Mitochondrial Biogenesis and Acute Kidney Injury

 描述(由申请人提供)： 该项目的长期目标是确定对线粒体功能障碍是关键组成部分的疾病的药物治疗。例子包括急性器官衰竭(例如创伤性脑功能障碍(退伍军人)、中风(退伍军人)和肾脏)、慢性疾病(例如创伤后应激障碍(退伍军人)、糖尿病、神经退行性疾病)和线粒体疾病。由于没有真正有效的疗法来促进急/慢性疾病的细胞和器官修复/再生，以及器官功能的恢复，因此迫切需要药物疗法。缺血/再灌注(I/R)时氧化应激(I/R)后线粒体功能障碍伴随着ATP的丢失，导致细胞损伤和死亡，导致器官衰竭。细胞通过线粒体生物发生(MB)取代陈旧和功能失调的线粒体。因此，我们启动了MB的药物发现计划，最近确定5-HT1F受体是MB的新靶点。5-HT1F受体是一种研究较少的5-羟色胺受体，在中枢神经系统和其他组织中发现，包括肾脏。LY344864是一种高效、选择性的5-HT1F激动剂，在1-100 nM作用24小时可诱导肾小管上皮细胞(RPTC)产生MB。用siRNA敲除5-HT1F受体证实LY334864通过5-HT1F受体产生MB。在幼稚小鼠身上的实验证明，LY344864在24小时内在肾脏中诱导了MB。一项概念验证实验确定，LY344864加速了小鼠I/R诱导的急性肾损伤(AKI)后的肾功能恢复。这些令人兴奋的结果支持了5-HT1F激动剂诱导的MB在AKI后损伤后恢复线粒体功能并刺激细胞和肾脏修复/再生的假说。具体目的1：阐明5-HT1F受体激动剂LY344864和Lasmiditan诱导RPTC MB的信号通路。Lasmiditan是一种新的5-HT1F受体激动剂，正在进行临床试验，用于治疗偏头痛，在我们的MB筛查试验中检测呈阳性。由于Lasmiditan很有可能成为FDA批准的药物，并且可能被重新利用，我们将研究LY344864和Lasmiditan诱导MB的信号通路。具体目的2：测定新近建立的5-HT1F受体基因敲除小鼠和增龄野生型小鼠肾脏线粒体功能和动态平衡及MB。目的3：探讨5-HT1F受体在肾I/R所致AKI中的作用及其恢复，以及LY344864和Lasmiditan在促进MB和促进肾功能恢复中的作用。这些研究将检测1)MB的新靶点5-HT1F受体，2)MB的新途径，3)5-HT1F受体在线粒体功能和动态平衡中的作用，以及4)5-HT1F受体激动剂在治疗急性器官损伤，特别是AKI中的疗效，使用细胞和体内模型。将结合分子生物学、生化和药理学方法和基因敲除小鼠来完成上面确定的目标。最终，这些研究可能导致新的治疗方法，以提高细胞和器官的存活率和功能，在许多病理情况下。

 DESCRIPTION (provided by applicant): The long-term goal of this project is to identify pharmacological treatments for diseases in which mitochondrial dysfunction is a critical component. Examples include acute organ failures (e.g. traumatic brain disorder (VETERAN), stroke (VETERAN), and kidney), chronic diseases (e.g. PTSD (VETERAN), diabetes, neurodegenerative) and mitochondrial diseases. Because there are no truly effective therapies that promote cell and organ repair/regeneration, and recovery of organ function in acute/chronic diseases, drug therapies are desperately needed. Mitochondrial dysfunction with the loss of ATP occurs after oxidative stress during ischemia/reperfusion (I/R) and leads to cell injury and death, and organ failure. Cells replace old and dysfunctional mitochondria through mitochondrial biogenesis (MB). Consequently, we started a drug discovery program in MB and recently identified the 5-HT1F receptor as a novel target for MB. The 5-HT1F receptor is a lesser studied 5-HT receptor found in the CNS and other tissues, including the kidney. LY344864, a potent and selective 5-HT1F agonist, induced MB in renal proximal tubular cells (RPTC) at 1-100 nM in 24 hr. Knockdown of the 5-HT1F receptor using siRNA confirmed LY334864 produced MB through the 5-HT1F receptor. Experiments in naïve mice documented that LY344864 induced MB in the kidney within 24 hr. A proof-of-concept experiment determined that LY344864 accelerated recovery of renal function after I/R-induced acute kidney injury (AKI) in mice. These exciting results support the hypothesis that 5-HT1F agonist-induced MB post-AKI injury restores mitochondrial function and stimulates cell and renal repair/regeneration. Specific Aim 1: Elucidate the signaling pathway by which 5-HT1F receptor agonists LY344864 and lasmiditan induce MB in RPTC. Lasmiditan, a newer 5-HT1F receptor agonist undergoing clinical trials for the treatment of migraines, tested positive in our MB screening assay. Because lasmiditan is likely to become a FDA-approved drug and could be repurposed, we will study the signaling pathway by which LY344864 and lasmiditan induce MB. Specific Aim 2: Determine renal mitochondrial function and homeostasis, and MB in recently developed 5-HT1F receptor knockout mice and wild-type mice of increasing age. Specific Aim 3: Elucidate the role of the 5-HT1F receptor in AKI and the recovery thereof and the efficacy of LY344864 and lasmiditan in promoting MB and accelerating the recovery of renal function in a mouse model of renal I/R-induced AKI. These studies will examine 1) a new target, 5-HT1F receptor, for MB, 2) a novel pathway of MB, 3) the role of 5- HT1F receptor in mitochondrial function and homeostasis, and 4) the efficacy of 5-HT1F receptor agonists in the treatment of acute organ injury, specifically AKI, using cellular and in vivo models. A combination of molecular biological, biochemical, and pharmacological approaches and a knockout mouse will be used to complete the aims identified above. Ultimately, these studies may lead to new therapeutic approaches to increase cell and organ survival and function in numerous pathologic situations.