目的 研究氟尿嘧啶(5-Fu)联合白藜芦醇对肿瘤细胞A431、TE-1的抑制和致凋亡作用及机制,以及两药联合对小鼠皮肤癌的治疗作用。方法 四甲基偶氮唑蓝(MTT)法、生长曲线实验、Chou-Talalay法和乳酸脱氢酶(LDH)溢出实验对联合用药抗肿瘤作用进行体外评价。光镜观察、DNA Ladder和共聚焦测定胞内Ca2+变化研究药物诱导凋亡的作用和机制。两阶段促癌法诱发小鼠皮肤癌,免疫组化法检测鼠皮中活化caspase-3的表达。结果 联合用药可以大大降低氟尿嘧啶(5-Fu)或白藜芦醇对A431的中效浓度。两药联用对肿瘤细胞的凋亡有协同作用,可以观察到更多典型凋亡形态学变化及显著的DNA片段化现象,其机制可能与大量增加细胞中Ca2+强度有关。联合给药后小鼠表皮肿瘤数量明显减少,表皮细胞中caspase-3的活化程度与单药相比明显增加。结论 联合给药在体内外均有协同抗肿瘤作用。
Abstract
OBJECTIVE To study the effects of 5-Fu combined with resveratrol (Res) on the growth and apoptosis of A431 and TE-1 cell lines and the underlying mechanism.To study the therapeutic effects of 5-Fu/Res combination on mouse skin papilloma chemically induced by DMBA/TPA. METHODS The effects of 5-Fu/Res combination on the viability of cancer cells were evaluated by MTT assay, growth curves assay and LDH releasing assay. The inhibitory effect of combination of the two drugs was analyzed by the method of Chou and Talalay. Apoptoses of A431 and TE-1 were determined by inverted microscope and gel electrophoresis of DNA fragment analysis. Intracellular Ca2+ concentration was determined to study the underlying mechanisms of 5-Fu and Res on the apoptosis of cancer cells. The mouse skin papilloma model was established by DMBA and TPA,the expression of actived-caspase-3 in mouse skin was examined by immunohistochemistry. RESULTS Combination of 5-Fu and Res decreased the median inhibitory concentration (IC50) to cancer cells remarkably. 5-Fu/Res combination showed a synergistic effect on apoptosis of A431 and TE-1 cells. Much more typical morphological changes of apoptosis and amount of fragmented DNA were observed in the cells treated with 5-Fu and resveratrol in combination than that in the cells treated with the agents alone. The increase of [Ca2+]i induced by 5-Fu/Res combination might be involved in the apoptotic induction. There was a significant decrease of the number of tumors after treatment with 5-Fu and Res in the tumor-bearing mice model. Active caspase-3 in the cells of mice skin was generated by 5-Fu in combination resveratrol was more effective than either alone. CONCLUSION The 5-Fu/Res combination shows synergistic anti-tumor effects both in vitro and in vivo.
关键词
氟尿嘧啶 /
白藜芦醇 /
TE-1 /
A431 /
协同效应 /
凋亡
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Key words
5-fluorouracil /
resveratrol /
TE-1 /
A431 /
synergistic effect /
apoptosis
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中图分类号：
R965
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参考文献
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脚注
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基金
河北省卫生厅资助项目(20130455)
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