Cyclophilin D as a Therapeutic Target following Traumatic Brain Injury

描述(由申请人提供)：本研究的目标是将创伤性脑损伤(TBI)后继发性细胞死亡和由此导致的发病率降至最低。改善脑外伤后预后的临床干预措施极其有限。线粒体功能障碍是创伤性脑损伤神经病理后遗症的关键环节。脑损伤后线粒体钙循环/超载增加，最终导致线粒体通透性转换孔(MPTP)开放。这个位于线粒体膜上的孔洞在钙离子升高和氧化应激时打开，并被线粒体蛋白亲环素D(CypD)门控。如果时间延长，MPTP的开放是灾难性的，因为线粒体膜电位的丧失，以及线粒体释放钙和死亡相关蛋白。这项研究的目的是通过限制MPTP的开放，最大限度地减少脑外伤后继发性细胞死亡和由此导致的发病率。免疫抑制剂环孢素A(CsA)通过与CypD结合来抑制MPTP的开放。我们和其他人之前证明，环孢素A在实验性脑损伤后应用时，可减少组织损伤的程度。不幸的是，CsA在高浓度时是有毒的，这是因为它抑制了钙调神经磷酸酶。最近，我们已经证明了原代神经元中CypD的水平大约是星形胶质细胞中的两倍，并且突触线粒体(神经元起源)的CypD水平也显著高于非突触线粒体(主要是非神经元起源)。由于其高CypD含量，我们假设神经元线粒体更容易受到MPTP开放的影响，也需要更高的CsA水平来抑制MPTP开放。为了验证这一假设，我们建议使用遗传和新的药理学方法来抑制神经元损伤后的CypD。具体地说，我们将使用CypD基因敲除小鼠和CsA衍生物NIM811，NIM811不结合或抑制钙调神经磷酸酶。其具体目的是：1：评估神经元线粒体高CypD含量增加MPTP开放易感性的假说，这些损伤导致细胞内钙离子升高和氧化应激。2：评估CypD抑制剂NIM811保护神经元免受兴奋性毒性损伤所需的水平与其CypD含量成正比的假设。3：验证线粒体CypD水平调节小鼠脑外伤后神经病理和功能转归的假说。4：验证CypD抑制剂NIM811减少脑损伤后组织损伤和改善功能预后的假说。根据这些研究的结果，我们预计NIM811将显示出强大的潜力，作为新的治疗脑外伤的方法。公共卫生相关性：创伤性脑损伤(TBI)在美国是一个毁灭性的医疗问题，目前还没有被批准用于创伤后临床干预的药物治疗。迫切需要改进的创伤性脑损伤治疗方案。这项建议研究了CSA的衍生物NIM811在限制脑损伤引起的脑损伤和功能障碍方面的潜力。

DESCRIPTION (provided by applicant): The goal of this research is to minimize the secondary cell death and resultant morbidity following traumatic brain injury (TBI). Clinical interventions to improve outcome following TBI are extremely limited. Mitochondrial dysfunction is a pivotal link in the neuropathological sequalae of traumatic brain injury (TBI). TBI-induced increases in mitochondrial Ca2+ cycling/overload ultimately lead to opening of the mitochondrial permeability transition pore (mPTP). This pore, located on the inner mitochondrial membrane, opens in response to elevated Ca2+ and oxidative stress, and is gated by the mitochondrial protein cyclophilin D (CypD). When prolonged, mPTP opening is catastrophic as a result of the loss of mitochondrial membrane potential, and the release of calcium and death-related proteins from mitochondria. The goal of this research is to minimize the secondary cell death and resultant morbidity following TBI by limiting mPTP opening. The immunosuppressant Cyclosporin A (CsA) inhibits mPTP opening by binding to CypD. We and others previously demonstrated that CsA reduces the extent of tissue damage when administered following experimental TBI. Unfortunately, CsA is toxic at high concentrations, resulting from its inhibition of calcineurin. Recently we have demonstrated that CypD levels in primary neurons are approximately double the levels found in astrocytes and that CypD levels are also significantly higher in synaptic mitochondria (neuronal origin) compared to non-synaptic mitochondria (predominately non-neuronal origin). As a result of their high CypD content, we hypothesize that neuronal mitochondria are more vulnerable to mPTP opening and also require greater CsA levels to inhibit mPTP opening. To test this hypothesis we propose to use genetic and newer pharmacologic approaches to inhibit CypD following neuronal injury. Specifically, we will use CypD knockout mice and a CsA derivative, NIM811, which does not bind to or inhibit calcineurin. The specific aims are: 1: To evaluate the hypothesis that the high CypD content of neuronal mitochondria enhances vulnerability to mPTP opening following insults that result in elevated intracellular Ca2+ and oxidative stress. 2: To evaluate the hypothesis that the levels of the CypD inhibitor NIM811 required to protect neurons from excitotoxic insult is proportional to their CypD content. 3: To examine the hypothesis that mitochondrial CypD levels modulate the neuropathologic and functional outcome following TBI in mice. 4: To examine the hypothesis that CypD inhibitor NIM811 reduces tissue damage and improves functional outcome following TBI. Based on the results of these studies, we anticipate that NIM811 will exhibit strong potential as novel therapy for TBI. PUBLIC HEALTH RELEVANCE: Traumatic brain injury (TBI) is a devastating healthcare problem in the United States, with no pharmacological treatments currently approved for clinical intervention following injury. Improved TBI treatment options are urgently needed. This proposal examines the potential of NIM811, a derivative of CsA, to limit the brain damage and dysfunction resulting from TBI.