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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Ca~(2+)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Ca~(2+)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。
Objective To study the inhibitory and apoptotic effects of 5-Fu combined with resveratrol on tumor cells A431 and TE-1, as well as their therapeutic effects on the skin cancer in mice. Methods MTT assay, growth curve assay, Chou-Talalay assay and lactate dehydrogenase (LDH) spill assay were used to evaluate the antitumor effects of the combination therapy in vitro. Light microscopy, DNA Ladder and confocal determination of intracellular Ca ~ (2+) changes in drug-induced apoptosis and its mechanism. Skin cancer was induced by two stages of cancer promotion, and the expression of activated caspase-3 in mouse skin was detected by immunohistochemistry. Results Combination therapy can significantly reduce the median concentration of 5-Fu or resveratrol A431. The combination of the two drugs has a synergistic effect on the apoptosis of tumor cells, and more typical morphological changes of apoptosis and significant DNA fragmentation can be observed. The mechanism may be related to the increase of Ca 2+ intensity in cells. The number of mouse epidermal tumors decreased significantly after combined administration, and the activation of caspase-3 in epidermal cells was significantly increased compared with that of single drug. Conclusions The combination therapy has synergistic anti-tumor effect both in vitro and in vivo.