Targeting CNTF to increase adult forebrain neurogenesis

描述（由申请人提供）：前脑室下区（SVZ）的纤毛神经营养因子（CNTF）促进成年小鼠的神经发生。我们发现CNTF通常通过细胞接触在中枢神经系统中受到抑制，并在中风诱导的神经元丢失后大幅增加。这有助于我们发现一种新的局灶黏附激酶(FAK)-JNK信号通路，该信号通路通过与胶质细胞中的DvE5整合素结合的配体激活，并有效抑制其CNTF的表达。重要的是，药理学FAK抑制剂的全身治疗刺激SVZ CNTF和神经发生。我们的数据显示，卒中诱导的神经发生完全由卒中诱导的CNTF介导，这可能被卒中诱导的LIF和炎症在第一周抵消。在体外，玻璃体连接蛋白抑制CNTF并刺激LIF，可能代表了神经发生的血管来源调节剂。在这里，我们将在体内定义这些途径，使用可用的转基因小鼠和成年小鼠的药理学药物的组合。具体来说，Aim 1a将确定正常情况下SVZ星形胶质细胞中vitronectin是否通过DvE5 integrin-FAK途径抑制CNTF表达。这将通过在玻璃体连接素-/-小鼠的脑内拯救实验来完成，注射针对Dv和E5整合素亚基的整合素阻断抗体，使用E5整合素-/-和诱导星形胶质FAK-/-小鼠，并确定对祖细胞和神经干细胞（NSCs）的影响。Aim 1b将通过脑内注射药理学抑制剂和JNK-/-小鼠来检测这是否由JNK和/或其他途径介导，同时测量对CNTF表达和神经发生的影响。Aim 2a将确定中风引起的血源性玻璃体连接蛋白渗漏是否会导致LIF表达增加，从而刺激NSC自我更新。这将在大脑中动脉闭塞（MCAO）模型中通过静脉注射玻璃体连接蛋白-/-来完成

DESCRIPTION (provided by applicant): Ciliary neurotrophic factor (CNTF) in the subventricular zone (SVZ) of the forebrain promotes neurogenesis in adult mice. We found that CNTF is normally repressed in the CNS by cell contact and increases greatly after stroke-induced neuron loss. This helped us to discover a new focal adhesion kinase (FAK)-JNK signaling pathway that is activated by ligand binding to DvE5 integrin in glia and potently inhibit their CNTF expression. Importantly, systemic treatment with pharmacological FAK inhibitors stimulates SVZ CNTF and neurogenesis. Our data show that stroke-induced neurogenesis is mediated entirely by stroke-induced CNTF, which may be counteracted by stroke-induced LIF and inflammation over the first week. In vitro, vitronectin represses CNTF and stimulates LIF, perhaps representing a vascular-derived regulator of neurogenesis. Here we will define these pathways in vivo, using a combination of available transgenic mice and pharmacological drugs in adult mice. Specifically, Aim 1a will determine whether vitronectin represses CNTF expression through the DvE5 integrin-FAK pathway in SVZ astrocytes under normal conditions. This will be done by intracerebral rescue experiments in vitronectin-/- mice, injecting integrin blocking antibodies against Dv and E5 integrin subunits, using E5 integrin-/- and inducible astroglial FAK-/- mice and defining effects on progenitors vs. neural stem cells (NSCs). Aim 1b will test whether this is mediated by JNK and/or other pathways, using intracerebral injections of pharmacological inhibitors and JNK-/- mice, while measuring effects on CNTF expression and neurogenesis. Aim 2a will determine whether stroke-induced leakage of blood-derived vitronectin causes increased LIF expression to stimulate NSC self-renewal. This will be done in a middle cerebral artery occlusion (MCAO) model by i.v. injections of vitronectin in vitronectin-/- mice. Aim 2b will determine whether the same integrin-FAK pathways play a role after MCAO, and whether CNTF and LIF can be regulated by FAK inhibitors. Aim 2c will define whether LIF is involved in repressing neurogenesis or is preconditioning the SVZ for later CNTF-induced neurogenesis. Aim 3a. Wwill test whether systemic FAK inhibitor treatment is more effective in inducing SVZ CNTF and neurogenesis when started after inflammation has resolved following MCAO, or Aim 3b when given intermittently. FAK acts via a different mechanism than the D2 dopamine receptor, which we showed increases SVZ neurogenesis by increasing astroglial CNTF. Therefore, Aim 3c will combine FAK inhibitor and D2 agonist to maximize CNTF expression and neurogenesis after MCAO, and determine whether this allows more new neurons to populate the injured tissue. The potential effects of these treatments on sensorimotor function will be assessed by behavioral tests.