Biochemical Mechanisms of Vasospasms

描述(摘自申请者摘要)：长期目标 这个应用是为了发现引起脑血管痉挛的分子(S) 蛛网膜下腔出血(SAH)后。血管痉挛是一种常见的 蛛网膜下腔出血患者迟发性缺血性卒中。有人建议，氧化 SAH后的胆红素会产生抑制蛋白磷酸酶的化合物， 这种抑制会导致血管收缩和血管增殖 见于血管痉挛患者。我们已经确定了候选分子 是似乎产生血管收缩的胆红素的过氧化片段 体外培养颈动脉血管和血管平滑肌细胞的增殖。 此外，这些分子在体外对血管产生代谢作用， 与脑脊液产生的血管痉挛相同。最后， 过氧化胆红素分子存在于脑脊液中 血管痉挛。此应用程序中的前两个目标将决定 脑血管痉挛患者的脑脊液中存在胆红素的过氧化形式； 哪些片段与临床血管痉挛有关；哪些片段与临床血管痉挛有关 过氧化形式的胆红素引起血管收缩和血管 体外增殖。这些研究将采用生化提纯。 胆红素氧化形式的分离程序和鉴定方法 它们的结构。化合物的鉴定可能会使我们有可能 开发一种针对血管痉挛的测试。体外颈动脉环试验用于 评估氧气消耗、等长力、高能磷酸盐和 有无血管痉挛患者脑脊液的磷酸酶活性。文化 将被用来确定脑脊液是否来自于 血管痉挛和过氧化胆红素化合物刺激血管增殖 将细胞在体外与对照溶液进行比较，以及这是否增加了 细胞增殖与蛋白磷酸酶抑制有关。第三 AIM将测试血管收缩是否可能是由于抑制 由过氧化胆红素片段引起的平滑肌磷酸酶，如果是这样的话 这发生在哪个亚细胞室，以及哪些磷酸酶是 被禁止了。最后一个目标将确定脑脊液是否来自血管痉挛患者 和纯化的氧化胆红素分子(当注射到蛛网膜下腔时 在这个活体模型中，(啮齿类动物的空间)引起血管痉挛和脑损伤。 在未来的研究中，该模型将被用于筛选可能的治疗方法。这个 该项目的最终长期目标是确定 血管痉挛，以发展有效的诊断，治疗和 这种脑血管疾病的预防方法。

DESCRIPTION (Adapted from applicant's abstract): The long-term objective of this application is to discover the molecule(s) that cause cerebral vasospasm following subarachnoid hemorrhage (SAH). Vasospasm is a frequent cause of delayed ischemic stroke in SAH patients. It is proposed that oxidation of bilirubin following SAH produces compounds that inhibit protein phosphatases, and that this inhibition causes the vasoconstriction and vascular proliferation seen in patients with vasospasm. We have identified candidate molecules that are peroxidized fragments of bilirubin that appear to produce vasoconstriction of carotid vessels and proliferation of vascular smooth muscle cells in vitro. In addition, these molecules produce metabolic effects on vessels in vitro that are identical to those produced by CSF from patients with vasospasm. Lastly, the peroxidized bilirubin molecules are present in the CSF of patients with vasospasm. The first two aims in this application will determine which peroxidized forms of bilirubin are found in the CSF of patients with vasospasm; which fragments correlate with the presence of clinical vasospasm; and which peroxidized forms of bilirubin cause vascular constriction and vascular proliferation in vitro. These studies will employ biochemical purification procedures to isolate the oxidized forms of bilirubin and methods to identify their structures. Identification of the compounds may make it possible to develop a test for vasospasm. An in vitro carotid artery ring assay is used to assess the oxygen consumption, isometric forces, high-energy phosphates, and phosphatase activity of CSF from patients with and without vasospasm. Cultures of smooth muscle cells will be used to determine whether CSF from patients with vasospasm and the peroxidized bilirubin compounds stimulate proliferation of the cells in vitro compared to control solutions, and whether this increase in cell proliferation is related to inhibition of protein phosphatases. The third Aim will test whether the vascular constriction might be due to inhibition of smooth muscle phosphatases by the peroxidized bilirubin fragments, and if so which subcellular compartment this occurs in, and which phosphatases are inhibited. The last Aim will determine whether CSF from patients with vasospasm and purified oxidized bilirubin molecules (when injected into the subarachnoid space of rodents) causes vasospasm and cerebral injury in this in vivo model. This model will be used to screen for possible therapies in future studies. The ultimate long-term goal for this project is to define the molecular causes of vasospasm in order to develop effective diagnostic, therapeutic and preventative approaches for this cerebral vascular disease.