MECHANISMS OF RESISTANCE TO EXCITOTOXIC CELL DEATH

描述（由申请人提供）：本申请是我们先前授权的直接延续，该授权针对控制C57 BL/6 J和FVB/NJ小鼠近交系中的尿素诱导细胞死亡易感性的QTL的遗传作图，这两种小鼠近交系对尿素诱导细胞死亡的易感性显著不同。在该项目的初始资助周期中，我们通过与C57 BL/6 J和FVB/NJ小鼠的异交鉴定了该复杂性状的3个易感基因座（Sicd 1 -3）。在最近的资助周期中，这些基因座已经使用相互同源菌株和使用间隔特异性同源菌株进行了确认，我们已经成功地将我们的Sicd 1基因座缩小到3.66 Mb间隔。在本申请中，我们提出使用已建立的同源菌株：1）鉴定Sicd 1的数量性状基因，并确定我们的候选基因在Sicd 1中的等位基因差异是否可以控制小鼠中的尿素诱导的细胞死亡易感性; 2）使用外显子表达谱来定义和表征Sicd 2易感性基因座的特定候选基因的作用;研究Sicd 1和Sicd 2 QTL间的上位性互作效应对油菜对油菜素诱导细胞死亡敏感性的影响。在目标1中，我们将确定潜在的候选基因的Sicd 1基因座，并确定是否在我们的候选基因的表达差异可以导致不同的易感性，通过几种不同类型的转基因小鼠，以减少诱导细胞死亡。在目标2中，我们将使用现有的同源品系或来自新的、高度信息化的杂交的小鼠，通过重组方法结合转录组分析来进一步定位和鉴定负责映射到Sicd 2的基因。最后，在目标3中，我们将确定来自Sicd 1和来自Sicd 2的基因座是否以互补的方式作用以改变对顺铂诱导的细胞死亡的易感性。总之，这些实验将阐明癫痫中海马神经元存活的关键途径，并有助于识别小鼠中候选的癫痫诱导细胞死亡修饰基因。了解这种疾病的分子病理生理学对于合理设计治疗干预措施至关重要。同样，癫痫细胞死亡途径的表征可能为了解兴奋性毒性起核心作用的其他神经退行性疾病的机制提供了依据。 公共卫生相关性：我们的研究将确定和表征负责调节小鼠品系中癫痫发作诱导的兴奋性毒性细胞死亡的易感性的基因。癫痫修饰基因的鉴定将为癫痫的发病机制提供深入的了解，并有助于开发治疗人类癫痫的新靶点。

DESCRIPTION (provided by applicant): This application is a direct continuation of our previous grant directed at the genetic mapping of QTL controlling seizure-induced cell death susceptibility in the C57BL/6J and FVB/NJ mouse inbred strains, which differ markedly in their susceptibility to seizure-induced cell death. During the initial funding cycle of this program, we identified 3 susceptibility loci for this complex trait (Sicd1-3) through outcross to C57BL/6J and FVB/NJ mice. In the most recent funding cycle, these loci have been confirmed using reciprocal congenic strains and using interval-specific congenic strains, we have successfully narrowed down our Sicd1 locus to a 3.66 Mb interval. In this application, we propose to use the established congenic strains to: 1) identify quantitative trait genes for Sicd1 and determine if allelic differences in our candidate gene in Sicd1 can control seizure-induced cell death susceptibility in mice; 2) to define and characterize the role of specific candidate genes for the Sicd2 susceptibility locus using exon expression profiling; and 3) to investigate the epistatic interaction between Sicd1 and Sicd2 QTLs influencing susceptibility to seizure-induced cell death. In Aim 1, we will identify prospective candidate genes for the Sicd1 locus and determine whether differences in expression of our candidate gene can result in differential susceptibility to seizure-induced cell death by making several different types of transgenic mice. In Aim 2, we will use existing congenic strains or mice from new, highly informative crosses to further localize and identify the genes responsible for mapping to Sicd2 by recombinational methods combined with transcriptome analysis. Lastly, in Aim 3, we will determine if loci from Sicd1 and from Sicd2 act in a complementary fashion to alter susceptibility to seizure-induced cell death. Taken together, these experiments will elucidate pathways critical for the survival of hippocampal neurons in epilepsy and aid in the identification of candidate seizure-induced cell death modifier genes in the mouse. An understanding of the molecular pathophysiology of this disease is essential to the rational design of therapeutic interventions. As well, the characterization of cell death pathways in epilepsy may provide insights into mechanisms involved in other neurodegenerative disorders in which excitotoxicity plays a central role. PUBLIC HEALTH RELEVANCE: Our studies will identify and characterize the genes responsible for modulating susceptibility to seizure- induced excitotoxic cell death in murine strains. Identification of epilepsy modifier genes will provide insight into the pathogenesis of epilepsy and aid in the development of novel therapeutic targets for the treatment of human epilepsy.