SENSING HYPOXIA IN THE CNS USING HERPES VECTORS

脑的缺血性损伤导致脑缺血期间的大量发病率。 围产期以及生命后期的死亡率。 在完整动物和体外的研究已经确定， 中枢神经系统缺血时发生的极端生理扰动 引发一种依赖于新基因的迟发性神经元死亡 缺氧引发的基因反应先于 延迟凋亡细胞死亡的激活。 一种方法， 确定新的治疗策略， 针对这种形式的神经元细胞死亡是通过检查 在CNS中指导低氧应答基因表达的机制。 遗传上保守的缺氧反应表现在 哺乳动物系统通过转录激活和后 转录mRNA稳定。 在外周，转录 已知事件是通过缺氧诱导的 一种转录因子HIF-1 α，它与细胞内的同源顺式元件结合， 限制性基因组的启动子区，从而模拟 基因转录速率。关于利用 这种缺氧反应机制在各种细胞内， CNS的区室（神经元、星形胶质细胞和小胶质细胞）及其 然而，缺乏与凋亡神经元损失的关系。 大体上， 我们假设在缺血后早期， 基因表达在细胞内表现出异质性 这种反应会触发一个序列， 直接或间接诱发迟发性神经元死亡。 我们计划 利用低氧应答HSV病毒载体来绘制区域和 缺氧信号传导的时间演变的隔室内的 缺血性小鼠CNS。后续研究将利用HIF神经元 同种型特异性抗体以表征HIF同种型诱导， 细胞定位，并通过EMSA确认DNA结合反应性 所有的超移测定都在缺氧条件下进行。 我们假设 这些实验将表征异质缺氧反应， 将定义缺血诱导的CNS转录中的离散因子 activation. 我们的长期目标是确定早期应对措施， 缺血性脑，并随后确定在缺氧调节节点 选择性调节缺氧基因激活信号级联反应。 这些发现将突出新的治疗策略， 抗缺氧诱导的迟发性神经元死亡。

Ischemic damage to the brain results in substantial morbidity during the perinatal period as well as mortality in the later decades of life. Studies in the intact animal as well as in vitro have established that the extreme physiologic perturbations which occur during CNS ischemia trigger a delayed form of neuronal death which is dependent on new gene transcription and that hypoxia triggered gene responses precede activation of delayed apoptotic cell death. One approach to the identification of novel therapeutic strategies which would protect against this form of neuronal cell death is through examination of the mechanisms which direct hypoxia responsive gene expression in the CNS. The phylogenetically conserved hypoxia response is manifest in the mammalian systems through transcriptional activation and post- transcriptional mRNA stabilization. In the periphery, transcriptional events are known to be mediated through the hypoxia inducible transcription factor, HIF-1alpha, which binds cognate cis elements in the promoter region of a restricted set of genes thereby simulating the rate of gene transcription. Information regarding the utilization of this hypoxia responsive mechanism within the various cellular compartments of the CNS (neuronal, astrocytic and microglial) and their relationship to apoptotic neuronal loss is lacking, however. Broadly, we hypothesize that early in the post-ischemic CNS hypoxic-regulated gene expression exhibits heterogeneity within the cellular compartments in the CNS and that this response triggers a sequence which either directly or indirectly elicits delayed neuronal death. We plan to exploit hypoxia responsive HSV viral vectors to map the regional and temporal evolution of hypoxic signaling within the compartments of the ischemic murine CNS. Subsequent studies will utilize HIF neuronal isoform specific antibodies to characterize HIF isoform induction, cellular localization and confirm DNA binding reactivity through EMSA supershift assays all under hypoxic conditions. We hypothesize that these experiments will characterize heterogeneous hypoxic response and will define discrete factors in ischemia induced CNS transcriptional activation. Our long-term goals are to identify early responses in the ischemic brain and subsequently identify regulatory nodes in the hypoxic signal cascade which can selectively modulate hypoxia gene activation. Such findings will highlight novel therapeutic strategies directed against hypoxia induced delayed neuronal death.