Murley J S, Grdina D J
Center for Mechanistic Biology and Biotechnology, Argonne National Laboratory, IL 60439-4833, USA.
Carcinogenesis. 1995 Nov;16(11):2699-705. doi: 10.1093/carcin/16.11.2699.
The effects of cycloheximide (CHX) and 2-[(aminopropyl)-amino]ethanethiol (WR-1065), each alone or in combination, on radiation-induced mutation induction at the hypoxanthine-guanine phosphoribosyl transferase (hprt) locus and cell killing were investigated using a Chinese hamster ovary (CHO) AA8 cell system. Treatment with CHX, a potent inhibitor of protein synthesis, at a concentration of 10 micrograms/ml administered 30 min prior to irradiation with 7.5 Gy had no effect on cell survival but did reduce the radiation-induced mutation frequency (per 10(6) survivors) from 106.5 +/- 8.8 (SEM) to 36.2 +/- 5.6 (SEM). Exposure of cells to 4 mM WR-1065 reduced the mutation frequency to 44.8 +/- 4.2 (SEM), but the combination of agents afforded no additional protection, that is 41.1 +/- 3.3 (SEM). The mechanism of action attributed to CHX in reducing mutation frequency is its ability to prevent the induction of an error-prone repair system. Split-dose radiation experiments, that is 8 Gy versus 4 Gy + 4 Gy separated by 3 h, were performed to evaluate and contrast the relative abilities of CHX and WR-1065, each alone or in combination, in affecting cell survival. Cycloheximide administered to cells 30 min before the first radiation dose and present throughout the 3 h incubation time prior to the second dose inhibited split-dose repair as evidence by a reduction in surviving fraction by 60% as compared with the value obtained for non-CHX-treated cells that were exposed to two equal doses of 4 Gy. Cells exposed to 4 mM WR-1065 immediately following the first 4 Gy radiation dose and then washed free 2.5 h before exposure to a second Gy dose, which was also followed by a 30 min exposure to WR-1065, increased the surviving fraction by 80% over the value obtained for cells not exposed to WR-1065 during their split-dose radiation treatment. When CHX treatment was combined with WR-1065 was abolished, that is surviving cell fraction was again reduced by approximately 60% as compared with untreated control groups. These results indicate that protein synthesis is required for WR-1065 to affect split-dose related repair processes. Presumably, the inhibition of the induction of an error-phone repair system by CHX would account for its effects on both resultant decreases in mutation frequency and cell survival. In contrast, WR-1065 and/or its disulfide metabolite appear to facilitate the efficacy and fidelity of such a repair system once it is induced.(ABSTRACT TRUNCATED AT 400 WORDS)
使用中国仓鼠卵巢（CHO）AA8细胞系统，研究了放线菌酮（CHX）和2-[(氨丙基)-氨基]乙硫醇（WR-1065）单独或联合使用对次黄嘌呤-鸟嘌呤磷酸核糖转移酶（hprt）基因座辐射诱导的突变及细胞杀伤的影响。用蛋白质合成的强效抑制剂CHX，在7.5 Gy照射前30分钟以10微克/毫升的浓度处理，对细胞存活没有影响，但确实降低了辐射诱导的突变频率（每10^6个存活细胞），从106.5±8.8（标准误）降至36.2±5.6（标准误）。细胞暴露于4 mM的WR-1065使突变频率降至44.8±4.2（标准误），但两种药物联合使用并未提供额外的保护，即41.1±3.3（标准误）。归因于CHX降低突变频率的作用机制是其能够阻止易错修复系统的诱导。进行了分次剂量辐射实验，即8 Gy与4 Gy + 4 Gy间隔3小时，以评估和对比CHX和WR-1065单独或联合使用影响细胞存活的相对能力。在第一次辐射剂量前30分钟给细胞施用CHX，并在第二次剂量前的3小时孵育期间一直存在，抑制了分次剂量修复，与未用CHX处理的接受两个4 Gy等剂量照射的细胞相比，存活分数降低了60%。在第一次4 Gy辐射剂量后立即将细胞暴露于4 mM的WR-1065，然后在第二次4 Gy剂量照射前2.5小时冲洗掉，并且在第二次剂量照射后还进行30分钟的WR-1065暴露，与在分次剂量辐射处理期间未暴露于WR-1065的细胞相比，存活分数增加了80%。当CHX处理与WR-1065联合使用被取消时，即存活细胞分数与未处理的对照组相比再次降低了约60%。这些结果表明，蛋白质合成是WR-1065影响分次剂量相关修复过程所必需的。据推测，CHX对易错修复系统诱导的抑制作用将解释其对突变频率和细胞存活降低的影响。相比之下，WR-1065和/或其二硫化物代谢产物一旦被诱导，似乎会促进这种修复系统的功效和保真度。（摘要截短为400字）
