Impact of heme and heme degradations products on cerebral vascular reactibility

迟发性脑血管痉挛和脑缺血是导致蛛网膜下腔出血患者死亡和严重神经功能损害的最常见原因。预防和医疗的选择仍然不足。血红素和血红素降解产物（HHDPs）参与血管痉挛的发展。在自己的实验中也有详细的分析，尤其是胆红素氧化终产物（box），来源于破裂血管周围降解的血红蛋白，通过抑制血管平滑肌细胞中的大电导BKCa钾通道诱导脑血管痉挛。将这些结果转化为新的治疗策略需要回答以下问题：(i)脑血管中HHDPs（特别是box）的分子靶点是什么，以及如何拮抗这种作用？（ii）迟发性脑缺血是否由毛细血管血流紊乱引起？（iii）体外观察能否转移到体内情况，特别是人类患者的情况？在该项目中，我们将结合钢丝肌图和BKCa通道亚基和剪接变异体的表达分析，确定和表征HHDPs在脑血管中的分子靶点，并研究不同动物差异敏感性的基础。在第二步，我们将分析毛细血管紊乱在体外继发性脑缺血中的作用，并量化HHDPs对小鼠脑血管反应性的影响。使用高分辨率磁共振成像（9.4 T MRI），我们将确定脑脊液和/或穿透性小动脉以及大血管对HHDP介导的缺血性损伤的贡献。应用体内已识别的小动脉和毛细血管的双光子成像，我们将细化我们对周细胞的调节作用的分析，周细胞是毛细血管血流的主要调节单位。在翻译的方法中，我们将开始知识转移到患有蛛网膜下腔出血的患者。我们将研究患者脑脊液中box和其他HHDPs与血管痉挛或继发性缺血的相关性，并详细阐述参数以进行全面的临床研究。建议的研究将有助于发展创新治疗策略的患者蛛网膜下腔出血。

Delayed cerebral vasospasm and brain ischemia are the most common causes of mortality and severe neurological impairment in patients with subarachnoid hemorrhage. Options for prevention and medical treatment are still insufficient. Heme and heme degradation products (HHDPs) are involved in the development of vasospasm. As also analyzed in detail in own experiments, especially bilirubin oxidation end products (BOXes), originating from degraded hemoglobin around ruptured blood vessels, induce cerebral vasospasm by inhibiting large conductance BKCa potassium channels in vascular smooth muscle cells. Translation of these results into new treatment strategies requires answers to the following questions: (i) What are the molecular targets of the HHDPs (and specifically BOXes) in cerebral blood vessels, and how can the action be antagonized? (ii) Is delayed cerebral ischemia due to disturbance of capillary blood flow? (iii) Can the in vitro observations be transferred to the in vivo situation, and especially the situation in human patients?In the proposed project, we will identify and characterize the molecular targets of HHDPs in brain vessels and investigate the basis for differential sensitivity in different animals combining wire myography with expression analysis of subunits and splice variants of the BKCa channel. In a second step, we will analyze the role of capillary disturbances for secondary brain ischemia in vitro, and quantify the impact of HHDPs on the vascular reactivity of mouse cerebral blood vessels in vivo. Using high resolution Magnetic Resonance Imaging (9.4 T MRI), we will identify the contribution of pial and/or penetrating resistance arterioles as well as large vessels to HHDP mediated ischemic insults. Applying 2-photon imaging of identified arterioles and capillaries in vivo, we will refine our analysis to the modulatory effect of pericytes, the main regulatory unit for capillary blood flow. In a translational approach we will initiate a transfer of knowledge to patients suffering from subarachnoid hemorrhage. We will investigate the correlation between BOXes and other HHDPs in the cerebrospinal fluid of patients and vasospasm or secondary ischemia and elaborate the parameters for a comprehensive clinical study. The proposed investigations will contribute to the development of innovative treatment strategies in patients with subarachnoid hemorrhage.