Objective: To investigate the effect of the chemoprotectant tempol on the anti-tumor activity of cisplatin (DDP).
Methods: The cellular toxicity of tempol in human colon cancer SW480 cells and mouse colon cancer CT26 cells were evaluated using MTT and cell counting kit-8 assays. CalcuSyn software analysis was used to determine the interaction between tempol and DDP in inhibition of the cell viability. A subcutaneous homograft mouse model of colon cancer was established. The mice were randomly divided into control group, tempol group, cisplatin group and tempol + DDP treatment group with intraperitoneal injections of the indicated agents. The tumor size, body weight and lifespan of the mice were measured, and HE staining was used to analyze the cytotoxic effect of the agents on the kidney and liver. Immunohistochemistry and Western blotting were performed to detect the expression of Bax and Bcl2 in the tumor tissue, and TUNEL staining was used to analyze the tumor cell apoptosis. The level of reactive oxygen species (ROS) in the tumor tissue was determined using flow cytometry.
Results: Tempol showed inhibitory effects on the viability of SW480 and CT26 cells. CalcuSyn software analysis showed that tempol had a synergistic anti-tumor effect with DDP (CI < 1). In the homograft mouse model, tempol treatment alone did not produce obvious anti-tumor effect. HE staining showed that the combined use of tempol and DDP alleviated DDP-induced fibrogenesis in the kidneys, but tempol also reduced the anti-tumor activity of DDP. Compared with the mice treated with DDP alone, the mice treated with both tempol and DDP had a significantly larger tumor size (P < 0.01) and a shorter lifespan (P < 0.05). Tempol significantly reversed DDP-induced expression of Bax and Bcl2 in the tumor tissue and tumor cell apoptosis (P < 0.001), and obviously reduced the elevation of ROS level in the tumor tissue induced by DDP treatment (P < 0.05).
Conclusions: Tempol can attenuate the anti-tumor effect of DDP while reducing the side effects of DDP. Caution must be taken and the risks and benefits should be carefully weighed when considering the use of tempol as an anti-oxidant to reduce the toxicities of DDP.
目的: 探讨化疗保护剂tempol对顺铂抗肿瘤效果的影响。
方法: 细胞学水平中, 运用MTT及CCK8方法检测tempol对人大肠癌细胞SW480和小鼠结肠癌CT26生存能力的影响; 通过CalcuSyn软件分析tempol与顺铂对细胞生存抑制的相互作用关系; 动物水平中, 构建小鼠皮下结肠癌模型, 将小鼠随机分为对照组、tempol组、顺铂组、tempol联合顺铂组, 分别给药处理, 观测各组小鼠肿瘤生长、小鼠生存期及小鼠体质量指标; 剥取小鼠肾脏及肝脏做HE染色观测药物的毒副作用; 免疫组化及Western blot检测肿瘤组织中凋亡相关蛋白Bax及Bcl2表达水平; 通过肿瘤组织TUNEL染色, 将凋亡细胞数目进行量化分析; 流式细胞仪检测肿瘤组中活性氧水平。
结果: 研究显示tempol可抑制人大肠癌细胞SW480和小鼠结肠癌CT26细胞生存; CalcuSyn软件分析显示tempol与化疗药物顺铂具有协同抗肿瘤的作用(CI < 1);但在小鼠体内实验中, tempol无明显的抗肿瘤作用; 虽然肾脏HE染色显示tempol与顺铂联合运用逆转顺铂造成的肾脏纤维化, 但同时tempol减弱顺铂对小鼠肿瘤生长的抑制作用(P < 0.01), 缩短小鼠生存期(P < 0.05), 明显拮抗顺铂的抗肿瘤效果; tempol逆转顺铂诱导的肿瘤细胞凋亡Bax及Bcl2表达, 同时肿瘤组织TUNEL染色显示顺铂明显增加肿瘤细胞的凋亡比率, 而与tempol联合运用则凋亡比率明显下降(P < 0.001);此外tempol降低顺铂引起的活性氧升高(P < 0.05)。
结论: tempol虽然可以降低化疗药物顺铂对其他器官的毒性, 但同时也减弱顺铂的抗肿瘤效果, 提示当将tempol作为化疗保护剂用于临床以减弱通过升高活性氧杀伤肿瘤的药物毒副作用时, 应衡量利弊, 谨慎运用。
Keywords: anti-oxidants; chemotherapy; cisplatin; drug toxicity; reactive oxygen species; tempol.