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目的:探讨甲基转移酶抑制剂5-氮杂-2’-脱氧胞苷(5-aza-2’-deoxycytidine,5-Aza-dC)对卵巢癌细胞系OVCAR3中FANCF(Fanconi anemia complementation group F)基因表达的影响,及其对化疗药物紫杉醇敏感性的影响,以了解5-Aza-dC的抗肿瘤效应,寻找卵巢癌治疗的新靶点。方法:采用5-Aza-dC处理FANCF基因甲基化的卵巢癌细胞OVCAR3和FANCF基因非甲基化的卵巢癌细胞A2780。应用甲基化特异性PCR(methylation speci-c-PCR,MSP)、RT-PCR和蛋白质印迹法分析这2种细胞经5-Aza-dC处理前后FANCF基因的甲基化状态以及FANCF mRNA和蛋白的表达情况。MTT法检测卵巢癌细胞的增殖情况。FCM检测紫杉醇对卵巢癌细胞凋亡的影响。建立OVCAR3和A2780细胞裸鼠移植瘤模型,观察5-Aza-dC对裸鼠移植瘤生长的影响,并采用免疫组织化学法检测移植瘤中FANCF蛋白的表达情况。结果:体外实验显示,5-Aza-dC处理后,OVCAR3细胞中FANCF基因发生去甲基化,FANCF mRNA和蛋白的表达水平增加,细胞生长缓慢。5-Aza-dC处理组OVCAR3细胞裸鼠移植瘤体积较未处理组明显缩小,质量也明显降低,FANCF蛋白表达增加;而A2780细胞裸鼠移植瘤组中未观察到上述变化。结论:甲基转移酶抑制剂5-Aza-dC可能通过抑制卵巢癌细胞增殖而成为潜在的卵巢癌治疗药物,但同时也会增加紫杉醇的耐药风险。
AIM: To investigate the effect of 5-aza-2’-deoxycytidine (5-Aza-dC) on the expression of FANCF in ovarian cancer cell line OVCAR3 (Fanconi anemia complementation group F ) Gene expression and its impact on paclitaxel chemosensitivity in order to understand the antitumor effect of 5-Aza-dC and to find a new target for the treatment of ovarian cancer. Methods: The ovarian cancer cells OVCAR3 with FANCF gene and non-methylated ovarian cancer cell A2780 with FANCF gene were treated with 5-Aza-dC. The methylation status of FANCF gene and the expression of FANCF mRNA and protein in these two cell lines before and after treatment with 5-Aza-dC were analyzed by methylation-specific PCR (MSP), RT-PCR and Western blotting. The expression of the situation. MTT assay ovarian cancer cell proliferation. Effect of paclitaxel on the apoptosis of ovarian cancer cells by. The effects of 5-Aza-dC on the growth of transplanted xenografts in nude mice were observed. The expression of FANCF protein in the transplanted tumors was detected by immunohistochemistry. Results: In vitro, the FANCF gene was demethylated in OVCAR3 cells after 5-Aza-dC treatment. The expression of FANCF mRNA and protein was increased and the cell growth was slow. The transplanted tumor volume of OVCAR3 cells in nude mice treated with 5-Aza-dC treatment was significantly smaller than that of untreated mice, the quality was significantly decreased, and the expression of FANCF protein was increased. However, no changes were observed in nude mice transplanted with A2780 cells. CONCLUSION: Methyltransferase 5-Aza-dC may be a potential therapeutic agent for ovarian cancer by inhibiting the proliferation of ovarian cancer cells, but it may also increase the risk of paclitaxel resistance.