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HEAT-INDUCED RADIOSENSITIZATION

热致辐射增敏

基本信息

批准号：
3188173
负责人：
RAYMOND L WARTERS
金额：
$ 14.06万
依托单位：
UNIVERSITY OF UTAH
依托单位国家：
美国
项目类别：
财政年份：
1987
资助国家：
美国
起止时间：
1987-09-30 至 1993-05-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/3188173
关键词：
DNA damage DNA repair chromatin chromosome disorders conformation gel electrophoresis heat nucleic acid sequence nucleic acid structure radiation genetics radiation sensitivity radiobiology

项目摘要

The overall objective of the proposal is to determine the effect(s) of radiosensitizing hyperthermic treatments on cell nuclear structure and function. In particular we will determine the effects(s) of : 1) acute heating (AH; 43 to 45 degrees); 2) step-down-heating (SDH; AH followed by chronic exposure to 41 degrees); 3) chronic heating at 41 degrees (tolerance development) before x-irradiation; and 4) chronic heating at 41 degrees subsequent to x-irradiation on the induction and/or repair of radiation- induced DNA double strand breaks (dsb), apurinic (AP) sites, DNA protein crosslinks (DPC) and chromosome aberration induction in CHO cells grown in monolayer. The hypothesis we will test is that a hyperthermia-induced increase in the induction frequency, and/or inhibition of repair, of one or more of these radiation-induced DNA lesions correlates with radiosensitization at the cellular level. To date, this hypothesis has not proven generally true in the case of DNA single strand breaks (ssb). The induction frequency and repairability (rate and extent) of radiation- induced lesions (ssb abd dsb) in specific DNA sequences will be measure in control and heated cells. Bulk DNA will be isolated by alkaline/neutral filter elution or alkaline sucrose gradient sedimentation from irradiated cells, and cells during the repair of DNA lesions. Bulk DNA collected onto nitrocellulose filters will be probed by filter hybridization against nucleic acid probes for transcriptionally active and/or inert DNA sequences. The possible role of heat-induced alterations of nuclear structure and/or composition in the inhibition of DNA damage removal will also be assessed. The kinetics of increase, and removal, of polypeptides in nucleosomal and nonnculesomal chromatin DNA and in cell nuclear matrices will be determined by exogenous nuclease digestion of DNA, centrifugal isolation of nuclear components and polypeptide quantitation and characterization of polyacrylamide gel electrophoresis. The extent to which hyperthermic exposure alters the number and/or conformation of nuclear matrix DNA binding sites will be determined by assessing the DNA "domain" size and frequency in control and heated cells.

该提案的总体目标是确定以下措施的效果：放射增敏热疗对细胞核结构和功能。我们将特别确定以下影响：1) 急性加热（AH；43 至 45 度）； 2) 降压加热（SDH；AH 随后长期暴露于41度）； 3）41度慢性加热（可耐受 X射线照射前的发育）； 4) 41度长期加热在X射线照射诱导和/或修复辐射之后- 诱导 DNA 双链断裂 (dsb)、无嘌呤 (AP) 位点、DNA 蛋白 CHO 细胞中的交联 (DPC) 和染色体畸变诱导单层。我们要测试的假设是，高热引起的一种或多种的诱导频率增加和/或修复抑制更多这些由辐射引起的 DNA 损伤与细胞水平的放射增敏作用。迄今为止，这一假说还没有已证明在 DNA 单链断裂 (ssb) 的情况下通常是正确的。辐射的感应频率和可修复性（速率和程度）特定 DNA 序列中诱导的损伤 (ssb abd dsb) 将在控制和加热单元。大量 DNA 将通过碱性/中性分离辐照后的过滤洗脱或碱性蔗糖梯度沉淀细胞，以及 DNA 损伤修复过程中的细胞。大量 DNA 收集到硝化纤维滤膜将通过滤膜杂交进行探测用于转录活性和/或惰性DNA序列的核酸探针。热引起的核结构改变和/或的可能作用还将评估组合物对DNA损伤去除的抑制作用。核小体和核小体中多肽增加和去除的动力学将测定非核体染色质 DNA 和细胞核基质通过外源核酸酶消化DNA，离心分离核成分和多肽的定量和表征聚丙烯酰胺凝胶电泳。高温的程度暴露会改变核基质 DNA 结合的数量和/或构象位点将通过评估DNA“结构域”的大小和频率来确定控制和加热单元。