Astrocyte-endothelial crosstalk after cerebral ischemia and hemorrhage

描述(申请人提供)：脑缺血和出血后的星形胶质细胞-内皮细胞串扰，反应性星形胶质细胞传统上被认为抑制了中枢神经系统损伤后的神经元可塑性。但现在出现的数据表明，反应性星形胶质细胞可能也有有益的作用。我们的初步数据提示：(I)反应性星形胶质细胞释放HMGB1可促进血管生成，(Ii)下调反应性星形胶质细胞释放HMGB1可能会加剧小鼠局灶性脑缺血后神经血管的恢复，(Iii)HMGB1可能上调脑内皮细胞上的RAGE受体，(Iv)内皮细胞RAGE可能通过β-2整合素增强内皮祖细胞的靶向黏附，(V)HMGB1可能促进内皮祖细胞的增殖、成熟和血管生成，从而促进卒中后的修复。基于这些先导数据，我们假设星形胶质细胞-内皮细胞串扰对于卒中后神经血管的恢复是必不可少的：反应性星形胶质细胞释放HMGB1，上调脑内皮细胞上的RAGE受体；RAGE结合循环内皮祖细胞上的β-2整合素，从而将它们拉入恢复中的大脑；一旦内皮祖细胞到达，HMGB1促进其增殖、成熟和血管生成。重要的是，我们认为这一途径可以促进缺血性或出血性中风后的恢复。我们将在三个目标上检验这一假设。在目标1中，我们探讨了来自刺激星形胶质细胞的HMGB1是如何上调对脑内皮细胞的RAGE并增强内皮祖细胞的靶向黏附的。在目标2中，我们剖析了HMGB1促进内皮祖细胞增殖、成熟和血管生成能力的机制。在目标3中，我们将使用局灶性脑缺血和脑出血的小鼠模型来证实这些星形胶质细胞-内皮-EPC机制，并表明它们实际上在体内介导神经血管的恢复。为了测试我们的途径，我们将结合使用细胞培养、体内小鼠模型、药物抑制剂、包括siRNA在内的分子技术、长期神经学结果和体内成像。这项研究应该明确一种新的机制，即反应性星形胶质细胞、脑内皮细胞和循环内皮祖细胞之间的相互作用是脑缺血和出血后神经血管恢复的基础。解剖这些细胞-细胞信号通路可能会导致新的治疗方法，促进缺血性和出血性中风患者的功能恢复。 公共卫生相关性：对于脑缺血和出血后脑星形胶质细胞-内皮细胞串扰的R01应用，中风研究的一个重要方向涉及研究新的治疗方法以促进康复。然而，中风后大脑如何反应和修复的潜在机制仍不清楚。在这里，我们将定义一种新的机制，涉及脑星形胶质细胞、内皮细胞和内皮祖细胞之间的新信号传递。我们的发现可能最终被用来开发新的治疗方法，以改善缺血性和出血性中风后的康复。

DESCRIPTION (provided by applicant): Astrocyte-endothelial crosstalk after cerebral ischemia and hemorrhage Reactive astrocytes were traditionally thought to inhibit neuronal plasticity after CNS injury. But emerging data now suggest that reactive astrocytes may also have beneficial actions. Our pilot data suggest that (i) reactive astrocytes release HMGB1 that can promote angiogenesis, (ii) downregulating the release of HMGB1 from reactive astrocytes may worsen neurovascular recovery after focal ischemia in mice, (iii) HMGB1 may upregulate RAGE receptors on cerebral endothelial cells, (iv) increased endothelial RAGE may enhance targeted adhesion of endothelial progenitor cells via beta-2 integrins, and (v) HMGB1 may increase proliferation, maturation and angiogenesis in endothelial progenitor cells, thus promoting repair after stroke. Based on these pilot data, we hypothesize that astrocyte-endothelial crosstalk is essential for neurovascular recovery after stroke: reactive astrocytes release HMGB1 that upregulates RAGE receptor on cerebral endothelium; RAGE binds beta-2 integrins on circulating endothelial progenitor cells thus pulling them into recovering brain; and once endothelial progenitors arrive, HMGB1 promotes their proliferation, maturation and angiogenesis. Importantly, we propose that this pathway can promote recovery after both ischemic or hemorrhagic strokes. We will test this hypothesis in three aims. In Aim 1, we ask how HMGB1 from stimulated astrocytes upregulate RAGE on cerebral endothelial cells and enhance the targeted adhesion of endothelial progenitor cells. In Aim 2, we dissect mechanisms that underlie the ability of HMGB1 to enhance proliferation, maturation and angiogenesis in endothelial progenitor cells. In Aim 3, we will use mouse models of focal cerebral ischemia and intracerebral hemorrhage to confirm these astrocyte-endothelium-EPC mechanisms and show that they actually mediate neurovascular recovery in vivo. To test our pathways, we will use a combination of cell culture, in vivo mouse models, pharmacologic inhibitors, molecular techniques including siRNA, long-term neurological outcomes, and in vivo imaging. This study should define a novel mechanism wherein crosstalk between reactive astrocytes, cerebral endothelium, and circulating endothelial progenitor cells underlie neurovascular recovery after cerebral ischemia and hemorrhage. Dissecting these cell-cell signaling pathways may lead to new therapeutic approaches for promoting functional recovery in patients after ischemic and hemorrhagic strokes. PUBLIC HEALTH RELEVANCE: For R01 application by Eng H. Lo Astrocyte-endothelial crosstalk after cerebral ischemia and hemorrhage An important direction for stroke research involves the investigation of new therapies to improve recovery. However, the underlying mechanisms of how brain responds to and repairs after stroke remain unclear. Here, we will define a new mechanism that involves novel signaling between brain astrocytes, endothelium and endothelial progenitor cells. Our findings may be eventually used to develop new therapies for improving recovery after both ischemic and hemorrhagic strokes.