Divergent Pathways of Cell Death after Brain Injury

描述（由申请人提供）：现在已经确定，在实验模型和人体中，程序性细胞死亡导致创伤性脑损伤TBI后继发性神经元死亡;然而，最近的数据表明存在多种细胞死亡途径。具体而言，涉及线粒体蛋白质凋亡诱导因子AIF的细胞死亡的替代/另外的途径，其产生大规模DNA片段化和细胞死亡，其被bcl-2和hsp 7 O抑制，但不被半胱天冬酶抑制剂抑制，似乎有助于体外亚硝化应激模型中的神经元细胞死亡，以及体内TBI后的神经元细胞死亡。本研究的目的是确定急性脑损伤后不同的和平行的细胞死亡途径是否有助于神经元死亡。假设在实验模型和TBI后的人类中发生由AIF介导的非半胱天冬酶依赖性神经元死亡，并且抑制核转位和/或AIF的活化减少TBI后的继发性神经病理损伤。具体目标的缩写如下：1）确认在暴露于亚硝化/氧化应激的原代皮层神经元中发生半胱天冬酶非依赖性、AIF介导的程序性细胞死亡，并使用关键调节剂和选择性抑制剂确立AIF的作用。2)在大鼠和bcl-2过表达小鼠中表征TBI后AIF的亚细胞再分布。确定AIF易位到细胞核和大规模DNA片段化的时间关联，AIF发生核易位的脑区域和细胞类型，并比较TBI后AlF介导和caspase-3介导的细胞死亡和坏死之间的相对贡献和区域差异。3)在大鼠和bcl-2过表达小鼠中建立TBI后AIF介导的细胞死亡的作用。使用标准和严格的范例测试AIF的这些关键调节剂对AIF介导的细胞死亡的生化足迹的影响以及它们对组织学和功能结果的影响。4)确定TBI后人类是否发生AIF介导的细胞死亡。TBI是成人和儿童发病率和死亡率的主要原因。延迟性细胞死亡导致发病率和死亡率，目前尚无特异性治疗方法成功地从实验研究过渡到床边。初步研究表明，其他非坏死性半胱天冬酶非依赖性细胞死亡途径参与TBI后的整体神经元死亡。该提案解决了关键问题：AIF介导的程序性细胞死亡是否有助于TBI后的迟发性神经元死亡和认知缺陷？如果AIF易位的抑制减少了体内TBI后的神经元死亡并改善了神经学结果，并且在人类中证实了AIF介导的细胞死亡，则靶向多个细胞死亡途径的新型临床相关策略将可用于治疗急性脑损伤。

DESCRIPTION (provided by applicant): It is now established that programmed-cell death contributes to secondary neuronal death after traumatic brain injury TBI in experimental models and in humans; however, recent data suggest that multiple cell death pathways exist. Specifically, an alternate/additional pathway of cell death involving the mitochondrial protein apoptosis-inducing factor AIF about that produces large scale DNA fragmentation and cell death that it is inhibited by bcl-2 and hsp7O, but not by caspase inhibitors, appears to contribute to neuronal cell death in models of nitrosative stress in vitro, and after TBI in vivo. The objective of this research is to determine whether divergent and parallel cell death pathways contribute to neuronal demise after acute brain injury. The HYPOTHESIS is that caspase-independent neuronal death, mediated by AIF occurs in experimental models and in humans after TBI and that inhibiting nuclear translocation and/or activation of AIF reduces secondary neuropathologic damage after TBI. Abbreviated SPECIFIC AIMS are: 1) Confirm that caspase-independent, AIF-mediated programmed cell death occurs in primary cortical neurons exposed to nitrosative/oxidative stress and establish a role for AIF using both key regulators and selective inhibitors. 2) Characterize the subcellular redistribution of AIF after TBI in rats and bcl-2 over-expressing mice. Determine the temporal association of AIF translocation into nuclei and large scale DNA fragmentation, the regions of brain and cell-types where nuclear translocation of AIF occurs, and compare the relative contributions of and regional differences between AlF-mediated and caspase-3-mediated cell death and necrosis after TBI. 3) Establish a role for AIF-mediated cell death after TBI in rats and bcl-2 over-expressing mice. Test the effects of these key regulators of AIF on biochemical footprints of AIF-mediated cell death and for their effects on histological and functional outcome using a standard and stringent paradigm. 4) Determine whether AIF-mediated cell death occurs in humans after TBI. TBI is a major cause of morbidity and mortality in adults and children. Delayed cell death contributes to morbidity and mortality and currently no specific therapies have successfully transitioned from experimental studies to the bedside. Preliminary studies suggest that other non-necrotic caspase-independent cell death pathways participate in overall neuronal death after TBI. This proposal addresses the key question: Does AIF-mediated programmed cell death contribute to delayed neuronal death and cognitive deficits after TBI? If inhibition of AIF translocation reduces neuronal death and improves neurological outcome after TBI in vivo and AIF-mediated cell death is confirmed in humans, a novel, clinically-relevant strategy targeting multiple cell death pathways will be available for the treatment of acute brain injury.