CHROMOSOME CHANGES, GENOME INSTABILITY, BYSTANDER EFFECT

描述（由申请人提供）：辐射诱导的旁观者效应是一种广泛描述的现象，它挑战了公认的辐射作用范例，但在机制水平上仍然知之甚少。基因组的不稳定性可能被认为是癌症发展的驱动力，电离辐射已被证明是这种潜在反应的有效诱导剂。这两种非靶向效应之间的关系尚未确定。据推测，辐射诱导的旁观者反应和基因组不稳定性反应是通过反应性自由基介导的途径机械连接。此外，我们假设，旁观者响应细胞将显示基因组不稳定性的水平等于直接辐射损伤的细胞所示。这一发现可能会增加对低/极低剂量辐射影响的关切。我们已经明确显示了一个旁观者效应与精确的带电粒子微束，其中核击中细胞有意识地从非击中旁观者细胞区分。我们将确定是否旁观者信号或非核[胞质]照射可以诱导哺乳动物细胞的基因组不稳定性。将使用特异性化学抑制剂和基因特异性siRNA测试旁观者和不稳定反应途径中的反应性自由基物质需求 接近。将评估染色体畸变频率随时间的变化。 染色单体/染色体畸变分析加/减端粒肽核酸探针， 应用多重荧光原位杂交技术（m-FISH）进行人类染色体核型分析 成纤维细胞，并在人-仓鼠杂交AL细胞中进行人染色体11 m-BAND分析。此外，我们将通过研究人支气管上皮细胞作为2D单层和作为3D气道样组织来比较2D与3D反应，因为更接近地反映体内细胞行为。四个具体目标和辅助假设将有助于定义旁观者反应和基因组不稳定性之间的机制关系。每个项目的目标之间的相互作用将确保在定义辐射诱导的旁观者效应的机制方面实现本P01的中心假设方面取得重大进展。

DESCRIPTION (provided by applicant): The radiation-induced bystander effect is a widely described phenomenon which challenges the accepted paradigm of radiation action, but remains poorly understood at a mechanistic level. Genomic instability may be considered to be a driving force in the development of cancer, and ionizing radiations have been shown to be efficient inducers of this latent response. The relationship between these two nontargeted effects has not been established. It is hypothesized that the radiation-induced bystander response and the genomic instability response are mechanistically linked through reactive radical species mediated pathways. Furthermore, we hypothesize that bystander responding cells will show genomic instability at levels equal to that shown by directly radiation damaged cells. Such a finding could augment concerns about the effects of low/very low doses of radiation. We have definitively shown a bystander effect with a precision charged particle microbeam, where nuclear hit cells were knowingly discriminated from non-hit bystander cells. We will determine whether bystander signaling or non-nuclear [cytoplasmic] irradiation can induce genomic instability in mammalian cells. Reactive radical species requirement in bystander and instability response pathways will be tested using specific chemical inhibitors and gene specific siRNA approaches. Frequencies of chromosomal aberrations will be assessed as a function of time. Chromatid/chromosome aberration analysis plus/minus a telomere peptide nucleic acid probe and complete karyotype analyses by multiplex fluorescence in-situ hybridization [m-FISH] in human fibroblast cells, and human chromosome 11 m-BAND analysis in human-hamster hybrid AL cells will be undertaken. Further, we will compare 2D versus 3D responses by studying human bronchial epithelial cells both as 2D monolayers and as 3D airway like tissues, as more closely reflecting cellular behavior in vivo. Four Specific Aims and subsidiary hypotheses will aid in the definition of the mechanistic relationship between the bystander response and genomic instability. Interaction between the Aims of each project will ensure significant progress to fulfillment of the central hypothesis of this P01 in defining the mechanism/s of the radiation-induced bystander effect.