Role of TRP channels in secondary injury after brain trauma

TRP通道在脑外伤继发性损伤中的作用

• 批准号: 8501703
• 负责人: Vladimir Gerzanich
• 金额: $ 32.4万
• 依托单位: UNIVERSITY OF MARYLAND BALTIMORE
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-09-01 至 2015-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8501703
• 关键词: Acute; Blood; Blood capillaries; Brain; Brain Death; Capillary Endothelial Cell; Cations; Cause of Death; Cell Line; Cells; Cellular biology; Cessation of life; Clinical; Clinical Trials; Contusions; Data; Edema; Electrophoretic Mobility Shift Assay; Electrophysiology (science); Endothelial Cells; Endothelium; Extravasation; FDA approved; Failure; Family; Functional disorder; Genes; Goals; Hemorrhage; Human; In Vitro; Inflammation; Inflammatory; Inflammatory Response; Injury; Ischemia; Knock-out; Knockout Mice; Lead; Lesion; Mechanics; Modeling; Molecular; Molecular Biology; Mus; Nervous System Physiology; Neurological outcome; Neurophysiology - biologic function; Nuclear; Outcome; Pharmaceutical Preparations; Physiological; Plasmids; Play; Property; Regulation; Reporter Genes; Riluzole; Role; Series; Stimulus; Stretching; Testing; Tissues; Transcriptional Regulation; Trauma; Traumatic Brain Injury; Up-Regulation; Wild Type Mouse; activating transcription factor; capillary; chromatin immunoprecipitation; controlled cortical impact; cytotoxic; disability; in vivo; injured; member; novel; p65; patch clamp; pre-clinical; preclinical study; receptor; research study; sulfonylurea receptor; therapeutic target; transcription factor

DESCRIPTION (provided by applicant): Traumatic brain injury (TBI) is inevitably complicated by microvascular dysfunction and/or failure, resulting in formation of edema and, even worse, "progressive secondary hemorrhage" (PSH). These abnormalities lead to ischemia, extravasation of blood and inflammatory cells, autodestruction of CNS tissues, and worsening of neurological outcome. We recently discovered that the NCCa-ATP channel is critically involved in PSH, with strong evidence showing critical involvement of sulfonylurea receptor 1 (SUR1), the regulatory subunit of channel. New preliminary data strongly suggest that an unusual member of the transient receptor potential (TRP) family, TRPM4, acts as the pore-forming subunit of the channel, and that the channel is transcriptionally upregulated in microvascular endothelial cells via mechano-activation of NF-?B, which occurs within minutes of force delivery to tissues in vivo and to cultured endothelial cells in vitro. Our overarching hypothesis is that trauma-associated mechanical forces activate the mechano-sensitive transcription factor, NF-?B, resulting in transcriptional upregulation of TRPM4 channels in microvascular endothelium, and that these channels play a critical role in the pathophysiology of progressive secondary hemorrhage (PSH) following TBI. Numerous ramifications of this hypothesis will be tested using molecular and cell biology, and patch clamp electrophysiology. In Specific Aim (SA) 1, we will use TRPM4 gene knock-out mice to determine the extent to which TRPM4 channels contribute to the overall pathophysiology of PSH. In the same Aim, we will also evaluate 2 treatments with strong translational potential: brief (24-hr) TRPM4 gene suppression using IV antisense oligodeoxynucleotide (AS-ODN), and riluzole, a pleiotropic FDA-approved drug that is a potent blocker of TRPM4 channels. In SA2, we will examine physiological modulation of the channel in freshly isolated CNS capillaries from TBI-mice, and in cultured endothelial cells exposed to a "trauma-like" mechanical stimulus (stretch), and we will determine the specific role of the channel in dysfunction and death of capillary endothelial cells. In SA3, we will examine the role of the mechano-activated transcription factor, NF-?B, in transcriptional regulation of the channel, and we will assess whether targeting this transcription factor in TBI is beneficial due to interference with expression of TRPM4 channels. TBI continues to be a leading cause of death and disability, with no treatments yet available to reduce secondary injury. We expect to find that TRPM4 plays a critical role as end-executioner in capillary destruction leading to PSH following TBI, and that block of TRPM4 by gene suppression, at the transcriptional level, or pharmacologically will yield significant reductions in the multiple cascades of secondary injury attributable to PSH and hemorrhage. These findings would establish a novel and powerful therapeutic target in TBI, leading to further pre-clinical as well as clinical trials with AS-ODN and/or riluzole, which we predict will have a major impact on acute treatment of TBI in humans.

描述(申请人提供)：创伤性脑损伤(TBI)不可避免地合并微血管功能障碍和/或衰竭，导致浮肿形成，更糟糕的是，“进行性继发性出血”(PSH)。这些异常导致缺血、血液和炎性细胞外溢、中枢神经系统组织的自体破坏，以及神经预后的恶化。我们最近发现NCCA-ATP通道与PSH密切相关，有强有力的证据表明该通道的调节亚单位--磺脲类受体1(SUR1)参与了该通道的关键作用。新的初步数据有力地表明，瞬时受体潜力(TRP)家族中的一个不寻常的成员TRPM4是该通道的成孔亚单位，该通道通过机械激活核因子-βB在微血管内皮细胞中转录上调，这发生在强制传递到体内组织和体外培养的内皮细胞的几分钟内。我们的主要假设是，创伤相关的机械力激活了机械敏感的转录因子--核因子-β，导致微血管内皮细胞中TRPM4通道转录上调，这些通道在脑外伤后进行性继发性出血(PSH)的病理生理学中起着关键作用。这一假说的许多分支将通过分子和细胞生物学以及膜片钳电生理学进行测试。在特定目的(SA)1中，我们将使用TRPM4基因敲除小鼠来确定TRPM4通道在PSH整体病理生理中的作用程度。在同一目标中，我们还将评估两种具有强大翻译潜力的治疗方法：使用IV反义寡核苷酸(AS-ODN)短暂(24小时)抑制TRPM4基因，以及利鲁唑，一种FDA批准的多效性药物，是TRPM4通道的有效阻断剂。在SA2中，我们将检测从脑创伤小鼠新鲜分离的CNS毛细血管中该通道的生理调节，以及暴露在“创伤样”机械刺激(拉伸)下的培养内皮细胞，我们将确定该通道在毛细血管内皮细胞功能障碍和死亡中的具体作用。在SA3中，我们将研究机械激活的转录因子NF-βB在通道转录调控中的作用，并评估在TBI中靶向该转录因子是否由于干扰TRPM4通道的表达而有益。颅脑损伤仍然是导致死亡和残疾的主要原因，目前还没有治疗方法来减少继发性伤害。我们期望发现TRPM4在脑外伤后导致PSH的毛细血管破坏中发挥关键作用，通过在转录水平或药物水平上通过基因抑制来阻断TRPM4将显著减少可归因于PSH和出血的多种继发性损伤。这些发现将建立一个新的和强大的脑损伤治疗靶点，导致进一步的AS-ODN和/或利鲁唑的临床前和临床试验，我们预测这将对人类脑损伤的急性治疗产生重大影响。