MECHANISMS OF NEURONAL APOPTOSIS

神经元凋亡机制

• 批准号: 6187425
• 负责人: HERBERT M. GELLER
• 金额: $ 21.14万
• 依托单位: UNIV OF MED/DENT NJ-R W JOHNSON MED SCH
• 依托单位国家: 美国
• 财政年份: 1997
• 资助国家: 美国
• 起止时间: 1997-04-20 至 2002-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6187425
• 关键词: DNA damage; animal genetic material tag; apoptosis; camptothecin; chemical structure function; cyclin dependent kinase; cyclins; enzyme activity; gene induction /repression; gene targeting; genetically modified animals; guanine nucleotide binding protein; ionizing radiation; laboratory mouse; laboratory rat; neural degeneration; neurons; neuroprotectants; neurotoxicology; neurotoxins; p53 gene /protein; radiation genetics; regulatory gene; tissue /cell culture; tumor suppressor proteins

This research to investigate the underlying molecular mechanisms that trigger neuronal apoptosis after DNA damage. While apoptosis is a cardinal feature of programmed cell death during neural development , it has been increasingly recognized that neurons also undergo apoptosis in response to injury. DNA damage produces apoptotic death in cultured mammalian central nervous system (CNS) neurons. Cell death is inhibited by two drugs which inhibit cyclin-dependent kinases. These drugs also protect neurons against apoptotic cell death after trophic factor withdrawal. This suggest that apoptosis in neurons due to both toxic injury and lack of trophic support required enzymes that normally control the cell cycle. Investigating this hypothesis is the major focus of this application. The first two specific aims will determine the role of immediate early genes and cycle-related proteins in apoptosis of cultured rat CNS neurons after DNA damage by irradiation or treatment with camptothecin. We will first determine the changes in immediate early genes (IEGs) and cell cycle-related proteins (cyclin- dependent kinases, Cdks) following DNA damage. We will then determine whether apoptosis is reduced by microinjection of antibodies directed against the IEGs that are increased following DNA damage or by injection of inhibitors or dominant negative mutants the Cdks and also by treatment with antisense oligonucleotides against the cyclins and Cdks that are increased following DNA damage. We will also evaluate the role of the p53 protein in neuronal apoptosis by utilizing neurons from p53 knockout mice. We will also utilize a series of Cdk-inhibitor drugs to determine a structure/activity relationship between Cdk inhibitory activity and reduction of neuronal apoptosis due to DNA damage and also for their ability to produce apoptosis in dividing PC12 cells. Finally, since campotothecin toxicity is reduced by inhibition of transcription, we will evaluate whether inhibition of transcription reduces toxicity of campothecin by reducing 1)DNA damage by camptothecin or 2) the apoptotic process itself or both. These studies should provide significant information about the underlying mechanisms that trigger apoptosis after DNA damage. Moreover, drugs that simultaneously kill dividing cells and protect neurons against toxicity from cancer chemotherapeutic agents or radiation may be exceedingly useful in therapy of brain cancers, in which survivors normally suffer a significant intellectual impairment.

这项研究旨在探讨 DNA损伤后触发神经细胞凋亡。而细胞凋亡是一种 神经发育过程中细胞程序性死亡的基本特征 已经越来越多地认识到神经元也会在 对伤害的反应。DNA损伤导致培养细胞的凋亡死亡 哺乳动物中枢神经系统(CNS)神经元。细胞死亡受到抑制 通过两种抑制细胞周期蛋白依赖性激酶的药物。这些药物也 营养因子对神经细胞凋亡的保护作用 戒烟。这表明，神经细胞的凋亡是由于两种毒素 损伤和缺乏营养支持需要正常情况下 控制细胞周期。研究这一假说是主要的 此应用程序的焦点。前两个具体目标将决定 即刻早期基因和周期相关蛋白在细胞周期调控中的作用 体外培养的大鼠中枢神经系统神经元DNA损伤后的细胞凋亡 用喜树碱治疗。我们将首先确定 即刻早期基因(IEGs)和细胞周期相关蛋白(Cyclin- 依赖的激酶，CDK)在DNA损伤后。然后我们将确定 微量注射定向抗体是否可减少细胞凋亡 对抗DNA损伤后或注射后增加的IEG 抑制物或显性负突变体的CDKs，也通过 反义寡核苷酸对细胞周期蛋白和细胞周期蛋白依赖性蛋白激酶的作用 在DNA损伤后会增加。我们还将评估该角色 利用P53的神经元研究P53蛋白在神经元凋亡中的作用 基因敲除老鼠。我们还将利用一系列CDK抑制剂药物来 确定CDK抑制物之间的结构/活性关系 DNA损伤引起的神经细胞凋亡的活性和减少 因为它们在分裂PC12细胞时产生凋亡的能力。最后， 由于Campotothecin的毒性通过抑制转录而降低， 我们将评估抑制转录是否降低了 喜树碱通过减少1)喜树碱引起的DNA损伤或2)减少细胞凋亡 进程本身或两者都是。这些研究应该提供有意义的 有关触发细胞凋亡的潜在机制的信息 DNA损伤。此外，同时杀死分裂细胞和 保护神经元免受癌症化疗药物或 放射治疗在脑癌的治疗中可能非常有用， 幸存者通常会遭受严重的智力障碍。