CONTROL OF MICROVASCULAR TONE IN TRAUMATIC BRAIN INJURY

创伤性脑损伤中微血管张力的控制

• 批准号: 6086110
• 负责人: JOSE A RAFOLS
• 金额: $ 25.71万
• 依托单位: WAYNE STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-03-10 至 2003-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6086110
• 关键词: blood circulation; brain circulation; brain injury; cerebral cortex; endothelin; flow cytometry; gene expression; genetic regulation; genetic transcription; genetic translation; immunocytochemistry; in situ hybridization; laboratory rat; messenger RNA; morphometry; neurobiology; neurogenetics; neuroregulation; nitric oxide; nitric oxide synthase; oligonucleotides; protein biosynthesis; ultrasound blood flow measurement; vascular endothelium; vascular smooth muscle

DESCRIPTION: (Verbatim from the Applicant's Abstract) The endothelial cells which form the vascular wall play a pivotal role in maintaining cerebral microvascular tone through the released of nitric oxide (NO) and endothelin (ET), which relax and constrict, respectively, the vascular smooth muscle. Although factors other than NO and ET participate in maintaining microvascular tone, reciprocal feedback mechanisms between NO and ET are important in the microvascular autoregulation in extracerebral arteries. However such a mechanism in brain microvessels in normal or abnormal states has never been investigated. From the three isoforms that participate in the synthesis of NO, the inducible nitric oxide synthase (iNOS) is expressed only after challenges to the CNS. NO derived from iNOS contributes to excessive amounts of NO released after trauma. Within the endothelin family, endothelin-1 (ET-1) is the most powerful vasoconstrictor produced in a variety of cellular types. Our long term objectives is to understand the interaction between iNOS and ET-1 in the control of microvascular tone during the acute phase post trauma that will ultimately lead to clinically effective therapy in the development of secondary injury. We propose that altered regulation of the genes that encode for iNOS and ET-1 in endothelial cells, at different time points, participate in the abnormal contractility of brain microvessels following TBI. Our specific aims are to: 1) identify vessels with abnormal diameter and the extent to which the endothelial cells that form these vessels synthesize iNOS and ET-1 proteins, 2) characterize quantitatively the expression of iNOS and ET-1 genes (mRNAs), 3) translationally inhibit the expression of iNOS and ET-1 genes in an attempt to restore impaired microcirculation. We will use morphometric analysis in combination with double immunocytochemistry at the ultrastructural level to detect temporal relationships between vascular diameter and protein synthesis of iNOS and ET-1, in situ hybridization for measurement of mRNA synthesis and in vivo intracerebroventricular application of antisense oligonucleotides to attenuate iNOS an ET-1 gene expression. On line laser Doppler flowmetry will be used to assess changes in cortical blood flow and determine how these changes are dependent on structural and molecular alterations as well as to infusions of antisense oligonucleotides. The results will provide valuable information on the expression of iNOS and ET-1 (proteins and mRNAs) and the causal relationship of this expression to alterations of brain perfusion following TBI. Precise time points of the dissociated expression of iNOS and ET-1 will be established which will serve as a baseline for future therapeutic interventions. This study will also provide direct evidence on the therapeutic value of suppressing (or attenuating) activity of genes in cerebral perfusion after TBI.

描述：（逐字研究来自申请人的摘要）内皮细胞 血管壁形成的哪个在维持脑中起着关键作用 一氧化氮（NO）和内皮素的释放的微血管张力 （ET），分别放松和收缩的是血管平滑肌。 尽管除非和ET以外的其他因素都参与维持微血管 音调，NO和ET之间的相互反馈机制在 脑外动脉的微血管自动调节。但是这样的 正常状态或异常状态的脑微血管机制从未有过 调查。来自参与NO合成的三种同工型 诱导型一氧化氮合酶（iNOS）仅在挑战之后表达 到CNS。没有源自INOS的毫无贡献的否 创伤后释放。在内皮素家族中，内皮素-1（ET-1）是 最强大的血管收缩剂以多种细胞类型产生。我们的漫长 术语目标是了解iNOS与ET-1之间的相互作用 在创伤后急性期间对微血管张力的控制 最终导致临床有效的疗法在次要的发展中 受伤。 我们建议改变编码iNOS和ET-1的基因调节 在内皮细胞中，在不同的时间点参与异常 TBI后大脑微血管的收缩力。我们的具体目的是：1） 鉴定直径异常的血管和内皮的程度 形成这些血管合成iNOS和ET-1蛋白的细胞，2） 定量表征iNOS和ET-1基因的表达（mRNA），3） 翻译抑制iNOS和ET-1基因的表达 恢复微循环受损。 我们将使用双重分析与双重分析 超微结构水平的免疫细胞化学检测时间 iNOS和ET-1的血管直径与蛋白质合成之间的关系， 原位杂交用于测量mRNA合成和体内 反室内室内施用反义寡核苷酸的衰减 iNOS A ET-1基因表达。在线激光多普勒流量仪将用于 评估皮质血流的变化并确定这些变化是如何变化的 取决于结构和分子改变以及输注 反义寡核苷酸。 结果将提供有关iNOS和 ET-1（蛋白质和mRNA）以及该表达与 TBI后大脑灌注的改变。精确的时间点 将建立iNOS和ET-1的解离表达，这将作为 未来治疗干预措施的基准。这项研究也将提供 直接证明抑制（或衰减）的治疗价值 TBI后脑灌注中基因的活性。