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目的:研究手霉素联合顺铂对人卵巢癌细胞3AO体外增殖及裸鼠移植瘤生长的抑制作用,并探讨其机制。方法:MTT法检测手霉素及手霉素联合顺铂对3AO细胞的体外增殖抑制作用;建立人卵巢癌裸鼠移植瘤模型,采用免疫组织化学SP法检测手霉素及手霉素联合顺铂对移植瘤组织survivin、NF-κB、血管内皮生长因子(vascular endothelial growth factor, VEGF)和caspase-3蛋白的表达,FCM检测移植瘤细胞周期和细胞凋亡率的变化。结果:手霉素对3AO细胞的增殖有明显抑制作用,且呈时间和剂量依赖效应(P<0.05)。手霉素可增强顺铂对3AO细胞的增殖抑制作用,随着手霉素浓度的增加(10～40 μmol/L),对3AO细胞的增殖抑制作用增强。手霉素或顺铂单药组以及联合用药组的裸鼠移植瘤体积均明显小于对照组,其中联合用药组裸鼠移植瘤体积明显小于手霉素或顺铂单药组(P<0.05)。各给药组裸鼠移植瘤组织中survivin、NF-κB和VEGF蛋白的表达水平均低于对照组(P<0.05),其中联合用药组又明显低于各单药组(P<0.05);各给药组caspase-3蛋白的表达水平均明显高于对照组(P<0.05)。手霉素单药组、顺铂单药组和手霉素联合顺铂组的细胞凋亡率依次升高,与对照组相比差异均有统计学意义(P<0.05);与对照组相比,G0/G1期细胞依次减少(P<0.05),G2/M期细胞依次增多(P<0.05),S期细胞变化不明显。结论:手霉素联合顺铂可明显抑制3AO细胞的体内外增殖并诱导其凋亡,其作用机制可能与下调survivin、NF-κB和VEGF以及上调caspase-3蛋白的表达有关。
OBJECTIVE: To study the inhibitory effect of cyclophosphamide combined with cisplatin on the proliferation of human ovarian cancer cell 3AO in vitro and the growth of xenografted tumor in nude mice, and to explore its mechanism. Methods: MTT assay was used to detect the in vitro proliferation of 3AO cells treated with 50μg / ml or 50μg / kg of cell viability. The human ovarian cancer xenografts were established by immunohistochemical SP method. The expression of survivin, NF-κB, VEGF and caspase-3 protein in the transplanted tumor tissues were detected by flow cytometry. The cell cycle and the apoptosis rate of the transplanted tumor were detected by FCM. Results: There was a significant inhibitory effect on the proliferation of 3AO cells in a dose and time dependent manner (P <0.05). Fucoxanthin could enhance the inhibitory effect of cisplatin on the proliferation of 3AO cells. With the increasing concentration of 10 ~ 40 μmol / L, the inhibitory effect on the proliferation of 3AO cells was enhanced. The volume of tumor xenografts in nude mice treated with monotherapy with or without cisplatin was significantly smaller than that of the control group. The volume of transplanted tumor in nude mice in combination group was significantly smaller than that in the control group (P <0.05) . The expression of survivin, NF-κB and VEGF protein in tumor xenografts of nude mice in each administration group was lower than that in control group (P <0.05), and the combination group was significantly lower than that of each single drug group (P <0.05). The expression of caspase-3 protein in each administration group was significantly higher than that in the control group (P <0.05). The rates of apoptosis in the monotherapy group, cisplatin group and cyclophosphamide group were higher than those in the control group (P <0.05). Compared with the control group Compared with the control group, the number of cells in G0 / G1 phase decreased (P <0.05), the number of G2 / M phase increased in turn (P <0.05), but there was no obvious change in S phase. CONCLUSION: The combination of cyclophosphamide and cisplatin can significantly inhibit the proliferation and induce the apoptosis of 3AO cells in vitro and in vivo. The mechanism may be related to the down-regulation of survivin, NF-κB, VEGF and the up-regulation of caspase-3 protein expression.