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目的:探讨黑素瘤抗原(melanoma antigen gene,MAGE)-A11对雌激素受体(estrogen receptor,ER)介导的乳腺癌MCF-7细胞增殖的影响。方法:利用RT-PCR及Western blotting筛选出ER表达阳性的人乳腺癌MCF-7细胞作为模式细胞,采用基因转染、RT-PCR和Western blotting检测MAGE-A11对17β-雌二醇(17β-E)诱导的ER下游靶基因Efp表达的影响,采用免疫共沉淀法检测MCF-7细胞中MAGE-A11和ER蛋白的相互作用,采用MTT法和克隆形成实验分别检测MAGE-A11及17β-E处理对MCF-7细胞生存率和细胞克隆形成数的影响。结果:ER阳性MCF-7细胞经17β-E处理24 h后,下游靶基因Efp的mRNA(2.97±0.16 vs 1.71±0.09,P<0.05)和蛋白表达水平显著升高(2.65±0.12 vs 0.92±0.06,P<0.05);转染MAGE-A11的MCF-7细胞经17β-E 24 h处理后,其Efp的mRNA(4.01±0.19 vs 2.97±0.16,P<0.05)及蛋白表达(3.52±0.15 vs 2.65±0.12,P<0.05)更显著增加。免疫共沉淀结果显示,外源性MAGE-A11与ER之间存在相互作用。MCF-7细胞经17β-E处理后细胞增殖率显著增加[(152±6.7)%vs(108±4.8%),P<0.05],转染MAGE-A11的MCF-7细胞经17β-E处理后细胞增殖率更显著增加[(181±8.6)%vs(152±6.7)%,P<0.05];17β-E处理后MCF-7细胞克隆形成数显著增多[(77±5)vs(18±2)个,P<0.05],转染MAGE-A11的MCF-7细胞经17β-E处理后细胞的克隆形成数更显著增加[(125±6)vs(77±5)个,P<0.05)。结论:在ER阳性的乳腺癌MCF-7细胞中,MAGE-A11可通过与ER的相互作用增强ER介导的Efp的表达,从而促进细胞增殖,MAGE-A11可能成为ER阳性乳腺癌内分泌治疗耐药的靶基因。
Objective: To investigate the effect of melanoma antigen gene (MAGE) -A11 on the proliferation of breast cancer MCF-7 cells mediated by estrogen receptor (ER). Methods: Human breast cancer MCF-7 cells with positive expression of ER were screened by RT-PCR and Western blotting as model cells. The expression of 17β-estradiol (17β-estradiol) was detected by RT-PCR and Western blotting. E), and the interaction between MAGE-A11 and ER protein in MCF-7 cells was detected by co-immunoprecipitation. The expressions of MAGE-A11 and 17β-E were detected by MTT assay and clonogenic assay Effect of treatment on the survival rate of MCF-7 cells and the number of cell clone formation. Results: After treated with 17β-E for 24 h, the expression of Efp mRNA (2.97 ± 0.16 vs 1.71 ± 0.09, P <0.05) and protein level in the ER positive MCF-7 cells were significantly increased (2.65 ± 0.12 vs 0.92 ± 0.06, P <0.05). After treated with 17β-E for 24 h, the mRNA and protein expression of Efp (4.01 ± 0.19 vs 2.97 ± 0.16, P <0.05) in MCF-7 cells transfected with MAGE- vs 2.65 ± 0.12, P <0.05) increased more significantly. Co-immunoprecipitation showed that there was interaction between exogenous MAGE-A11 and ER. The proliferation of MCF-7 cells was significantly increased after treated with 17β-E [(152 ± 6.7)% vs (108 ± 4.8%), P <0.05] (181 ± 8.6)% vs (152 ± 6.7)%, P <0.05]. The number of MCF-7 cells after 17β-E treatment increased significantly [(77 ± 5) vs P <0.05]. The number of colony-forming cells in MCF-7 cells transfected with MAGE-A11 increased more than those in MAGE-A11 cells treated with 17β-E (125 ± 6 vs. 77 ± 5, P < 0.05). Conclusion: MAGE-A11 can enhance ER-mediated Efp expression and promote cell proliferation in ER-positive breast cancer MCF-7 cells by interacting with ER. MAGE-A11 may be an ER-positive breast cancer endocrine therapy The target gene for the drug.