CYTOGENETICS--ALPHA PARTICLE INDUCED CHROMOSOMAL CHANGES

描述：(申请人描述)该计划项目的总体研究目标是了解辐射损伤的识别、处理和后果的机制基础，特别强调氡子体阿尔法粒子及其对人类健康的影响。核DNA的变化显著影响人类健康，与细胞死亡、突变和致癌变化有明确的关系。然而，非核靶产生的应力对长期有害影响的潜在贡献尚不确定。显然，膜、细胞质和细胞核之间的分子相互作用有助于损伤识别、信号和反应途径。通过使用微束可以有效地了解细胞成分在辐射响应中的作用。在这个项目中，我们将使用定义的影响(降到1)或100keV/um的阿尔法粒子来照射一个群体中的细胞核、细胞质或已知细胞。非HIT细胞(旁观者)可能受到活性氧的刺激和/或受到诱导的可移动细胞因子的影响。来自基因敲除小鼠、野生型、P53-和ATM的小鼠胚胎成纤维细胞将作为单个群体和成对群体进行照射。响应将分别在HIT和非HIT单元中进行评估。我们将提出这样的问题：粒子间距是否会影响基于原子核的反应？细胞质辐射是否会引发生物反应？在非HIT细胞中，细胞特异性辐射是否在基因依赖的基础上启动反应？我们将通过蛋白表达和定位来检测微核的发生率，通过RT-PCR来检测细胞凋亡、细胞生长和发育、细胞周期延迟和基因表达。与该计划项目的其他项目一致，终端和精确的现场特定微束照射的组合以解决明确的问题，将建立机制参数，并将定义低剂量下的阿尔法粒子响应性，重点是旁观者效应的贡献。

DESCRIPTION: (Applicant's Description) The overall research objectives of this program project are directed toward understanding the mechanistic bases for the recognition, processing and consequences of radiation induced lesions, with a particular emphasis on radon progeny alpha particles and their impact on human health. Nuclear DNA changes significantly impact human health with clear relationships to cell death, mutation and oncogenic changes. There is uncertainty however as to the potential contributions of stresses induced in non-nuclear targets to long term deleterious effects. It is clear that molecular interplay between membrane, cytoplasm and nucleus contributes to damage recognition, signaling and response pathways. An understanding of the role of cellular components in radiation responses can efficiently be obtained by use of a microbeam. In this project we will used defined influences (down to 1) or 100 keV/um alpha-particles to irradiate cell nucleus, cytoplasm or known cells in a population. Non-hit cells (bystanders) may be stressed by reactive oxygen species and/or affected by induced mobile cytokines. Mouse embryo fibroblasts derived from knockout mice, wild-type, p53-, and atm- will be irradiated both as single and as pair wise populations. Responses will be separately assessed in hit versus non-hit cells. We will ask the questions: Does particle spacing influence nucleus based responses? Does cytoplasmic irradiation initiate a biological response? Does cell specific irradiation initiate responses in non-hit cells on a genotype dependent basis? We will examine incidence of micronuclei; apoptosis, cell growth and development; cell cycle delay; and gene expression both by protein expression and localization, and mRNA by RT-PCR. In concert with the other projects of this program project, the combination of endpoints and precise site specific microbeam irradiation to address definitive question will establish mechanistic parameters and will define alpha-particle responsiveness at low doses with emphasis on the contribution of the bystander effect.