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药物治疗线粒体功能障碍是关键组成部分的疾病。实例包括急性器官衰竭（例如创伤性脑障碍（VETERAN）、中风（VETERAN）和肾脏）、慢性疾病（例如PTSD（VETERAN）、糖尿病、神经变性）和线粒体疾病。由于没有真正有效的疗法来促进细胞和器官修复/再生，以及急性/慢性疾病中器官功能的恢复，因此迫切需要药物疗法。缺血/再灌注（I/R）期间氧化应激后发生线粒体功能障碍伴ATP丧失，导致细胞损伤和死亡以及器官衰竭。细胞通过线粒体生物发生（MB）取代旧的和功能失调的线粒体。因此，我们开始了MB的药物发现计划，最近确定了5-HT 1F受体作为MB的新靶点。5-HT 1F受体是一种研究较少的5-HT受体，存在于CNS和其他组织（包括肾脏）中。LY 344864是一种有效和选择性的5-HT 1F激动剂，在1-100 nM浓度下24 h内诱导肾近端小管细胞（RPTC）中的MB。使用siRNA敲除5-HT 1F受体证实LY 334864通过5-HT 1F受体产生MB。在未经处理的小鼠中进行的实验证明，LY 344864在24小时内诱导了肾脏中的MB。概念验证实验确定，LY 344864加速了小鼠I/R诱导的急性肾损伤（阿基）后的肾功能恢复。这些令人兴奋的结果支持以下假设：AKI损伤后5-HT 1F激动剂诱导的MB恢复线粒体功能并刺激细胞和肾脏修复/再生。具体目的1：阐明5-HT 1F受体激动剂LY 344864和拉司米坦诱导RPTC中MB的信号通路。Lasmiditan是一种正在进行偏头痛治疗临床试验的新型5-HT 1F受体激动剂，在我们的MB筛查试验中检测为阳性。由于lasmiditan很可能成为FDA批准的药物，并且可能被重新利用，因此我们将研究LY 344864和lasmiditan诱导MB的信号通路。具体目标二：在最近开发的5-HT 1F受体敲除小鼠和年龄不断增加的野生型小鼠中确定肾脏线粒体功能和稳态以及MB。具体目标3：在肾I/R诱导的阿基小鼠模型中，阐明5-HT 1F受体在AKI及其恢复中的作用，以及LY 344864和lasmiditan在促进MB和加速肾功能恢复方面的疗效。这些研究将使用细胞和体内模型检查1）MB的新靶点5-HT 1F受体，2）MB的新途径，3）5-HT 1F受体在线粒体功能和稳态中的作用，以及4）5-HT 1F受体激动剂在治疗急性器官损伤（特别是阿基）中的疗效。将使用分子生物学、生物化学和药理学方法以及基因敲除小鼠的组合来完成上述目标。最终，这些研究可能会导致新的治疗方法，以增加细胞和器官的生存和功能，在许多病理情况。

 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.