Analysis by pulsed-field gel electrophoresis of DNA double-strand breaks induced by heat and/or X-irradiation in bulk and replicating DNA of CHO cells.
Analysis by pulsed-field gel electrophoresis of DNA double-strand breaks induced by heat and/or X-irradiation in bulk and replicating DNA of CHO cells.
复制标题
通过脉冲场凝胶电泳分析由热和/或 X 辐射诱导的 DNA 双链断裂，以及 CHO 细胞的复制 DNA。
DOI：
10.1080/09553009514551041
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发表时间：
1995
影响因子：
2.6
通讯作者：
Dewey,WC
中科院分区：
文献类型：
--
作者：
Wong,RS;Dynlacht,JR;Cedervall,B;Dewey,WC
对于给定量的细胞杀伤，与X射线相比，单独加热（10-80分钟，45·5°C）诱导非常少的双链断裂（dsb）。此外，在45·5°C下10分钟后立即进行X射线照射，仅导致X射线诱导的dsb剂量-反应曲线的斜率增加1·3倍，即与未加热的对照细胞的0·53 ± 0·005相比，加热细胞的dsb/100 Mbp/戈伊为0·67 ± 0·006（95%置信度）。然而，同样的热处理对由60-戈伊X射线诱导的dsb的修复速率造成> 5倍的抑制，其中在耐热（TT）细胞中的抑制程度比在非耐热（NT）细胞中的抑制程度小得多。这减少了对TT细胞修复的抑制，这与从TT细胞分离的核中去除过量核蛋白的速度比从NT细胞分离的核中去除过量核蛋白的速度更快相关。这些结果加上热诱导的放射增敏和修复dsb的热抑制的TT比为2-3与热放射增敏主要是由于核蛋白的热聚集干扰修复酶进入DNA dsb的假设一致。然而，S期细胞的选择性热辐射增敏可能是由于在复制区或复制区附近辐射诱导的dsb增加所致。例如，在单独的热疗（10-30分钟，45·5°C）或联合处理（60戈伊前10分钟，45·5 ° C）后，发现复制DNA中的dsb与大量DNA相比优先增加。在45·5°C下处理30分钟，在复制的DNA中诱导的dsbs相当于100戈伊，而在本体DNA中诱导的dsbs相当于3-5戈伊。当细胞在照射前立即加热时，60戈伊在复制DNA中诱导的dsb增加相当于200戈伊。我们假设，加热后观察到的复制DNA中单链区域增加2倍，导致辐射选择性地在复制叉处或其附近诱导dsb，其中热诱导的单链DNA增加应该发生。因此，这种优先增加双链断裂在复制DNA的热单独，特别是当热与辐射相结合，可以解释至少部分，高敏感性的S期细胞的热杀死和热辐射增敏。
For a given amount of cell killing, heat alone (10–80 min, 45·5°C) induced very few double-strand breaks (dsbs) compared with X-rays. Furthermore, 10 min at 45·5°C immediately prior to X-rays caused only a 1·3-fold increase in the slope of the X-ray-induced dsb dose—response curve, i.e. 0·67 ± 0·006 (95% confidence) dsbs/100Mbp/Gy for heated cells compared with 0·53 ± 0·005 for unheated control cells. However, this same heat treatment caused > 5-fold inhibition in the rate of repair of dsbs induced by 60-Gy X-rays, with the degree of inhibition being much less in thermotolerant (TT) cells than in non-tolerant (NT) cells. This reduced inhibition of repair in TT cells correlated with the more rapid removal of excess nuclear protein from nuclei isolated from TT cells than from NT cells. These results plus a TT ratio of 2–3 for both heat-induced radiosensitization and heat-inhibition of repairing dsbs are consistent with the hypothesis that heat radiosensitization results primarily from heat aggregation of nuclear protein interfering with access of repair enzymes to DNA dsbs. The selective heat-radiosensitization of S-phase cells, however, may result from an increase in radiation-induced dsbs in or near replicating regions. For example, a preferential increase in dsbs in replicating DNA compared with bulk DNA was found following either hyperthermia alone (10–30 min, 45·5°C) or a combined treatment (10 min, 45·5°C before 60 Gy). A 30-min treatment at 45·5°C induced dsbs equivalent to ∼ 10 Gy in replicating DNA compared with 3–5 Gy in bulk DNA. When cells were heated immediately before irradiation, the increase in dsbs induced in the replicating DNA by 60 Gy was equivalent to 200 Gy. We hypothesize that the observed 2-fold increase in single-stranded regions in replicating DNA after heat resulted in radiation selectively inducing dsbs at or near the replication fork where the heat-induced increase in single-stranded DNA should occur. Thus, this preferential increase in dsbs in the replicating DNA by heat alone and especially when heat was combined with radiation may explain at least in part, the high sensitivity of S-phase cells to heat killing and heat radiosensitization.