Shinomiya N, Shinomiya M, Wakiyama H, Katsura Y, Rokutanda M
Department of Biology, National Defense Medical College, Saitama, Japan.
Exp Cell Res. 1994 Feb;210(2):236-42. doi: 10.1006/excr.1994.1035.
The ability of caffeine, an agent that suppresses cell replication by inhibiting deoxyribonucleic acids (DNA) repair, to modulate the cytotoxicity of cis-diamminedichloroplatinum (CDDP) was investigated in murine lymphoma cell line EL-4. EL-4 cells were precultured with or without 20 micrograms/ml CDDP for 1 h and then cultured in the presence of 5 mM caffeine up to 48 h after reseeding. In CDDP-pretreated cells, suppression of cell growth and decrease in cell viability from 24 h were observed. Cell cycle arrest in G2 + M phase and a concomitant increase in both rhodamine 123 (R123) uptake and cell size (forward scatter) were observed in these cells. Treatment with caffeine alone suppressed growth rate, R123 uptake, cell size, and frequency of S phase fraction in the cell cycle. Combination of the two agents, CDDP+caffeine, strongly suppressed not only cell viability but also R123 uptake and cell size, compared with CDDP pretreatment alone. DNA histogram analysis by flow cytometry revealed that cultivation with caffeine hastened G2 + M arrest in CDDP-pretreated cells by reduction in the time of passing through S phase. DNA fragmentation was observed following incubation of CDDP-pretreated cells with caffeine for 16 h when marked accumulation in G2 + M phase was observed. The intensity of these ladder fragments increased in a time-related manner. These results demonstrate that enhancement of cytotoxic activity against CDDP-treated cells by caffeine is characterized by an acceleration of DNA degradation in G2 + M phase, namely apoptotic cell death. The fact that induction of DNA fragmentation during G2 + M phase by caffeine modulates the cytotoxicity of CDDP may give rise to a new combination regime of chemotherapy against malignant tumor cells.
咖啡因是一种通过抑制脱氧核糖核酸（DNA）修复来抑制细胞复制的物质，其对顺二氨二氯铂（CDDP）细胞毒性的调节作用在小鼠淋巴瘤细胞系EL-4中进行了研究。EL-4细胞在有或无20微克/毫升CDDP的情况下预培养1小时，然后在重新接种后于5毫摩尔咖啡因存在的条件下培养长达48小时。在预先用CDDP处理的细胞中，从24小时起观察到细胞生长受到抑制且细胞活力下降。在这些细胞中观察到细胞周期停滞于G2 + M期，同时罗丹明123（R123）摄取和细胞大小（前向散射）均增加。单独用咖啡因处理可抑制细胞周期中的生长速率、R123摄取、细胞大小以及S期分数频率。与单独的CDDP预处理相比，两种药物联合使用（CDDP + 咖啡因）不仅强烈抑制细胞活力，还抑制R123摄取和细胞大小。通过流式细胞术进行的DNA直方图分析显示，用咖啡因培养可通过缩短通过S期的时间加速CDDP预处理细胞中的G2 + M期停滞。在用咖啡因孵育预先用CDDP处理的细胞16小时后，当观察到G2 + M期有明显积累时，观察到DNA片段化。这些梯形片段的强度呈时间依赖性增加。这些结果表明，咖啡因增强对CDDP处理细胞的细胞毒性作用的特征是加速G2 + M期的DNA降解，即凋亡性细胞死亡。咖啡因在G2 + M期诱导DNA片段化从而调节CDDP的细胞毒性这一事实可能会产生一种针对恶性肿瘤细胞的新的联合化疗方案。
