FOS样抗原1对乳腺癌细胞MDA-MB-231增殖、侵袭和PRDM10基因甲基化的影响

Effects of FOSL1 on cell proliferation, cell invasiveness and the methylation of PRDM10 gene in breast cancer cell line MDA-MB-231

目的构建FOS样抗原1(FOSL1)基因沉默的载体,研究FOSL1对人乳腺癌细胞系MDA-MB-231增殖和侵袭的影响,以及对此细胞中PRDM10基因的甲基化状态的影响。方法 FOSL1沉默的pLVX-sh RNA-FOSL1-sh RNA干扰质粒载体,采用慢病毒感染法分别用p LVX-sh RNA-FOSL1-sh RNA载体以及pLVX-sh RNA空载的两种病毒感染靶细胞MDA-MB-231,分别记作转染组和空载组。以未转染的MDA-MB-231作为对照组。RT-PCR验证FOSL1表达后,分别采用MTT实验和Transwell实验检测MDA-MB-231的细胞增殖能力和侵袭能力。采用MSP法检测PRDM10基因的甲基化状态。Q-PCR和Western blot分别检测PRDM10的mRNA和蛋白质表达水平。结果MTT实验显示,24 h、48 h和72 h时,转染组MDA-MB-231的OD值低于对照组(均P<0.05);48 h和72 h时,转染组MDA-MB-231的OD值低于空载组(均P<0.05)。Transwell实验显示,与空载组和对照组相比,转染组的MDA-MB-231细胞侵袭能力降低(均P<0.05)。MSP实验显示,与空载组和对照组相比,转染组的MDA-MB-231细胞中PRDM10基因甲基化水平降低。Q-PCR和Western blot实验显示,与空载组和对照组相比,转染组PRDM10的mRNA和蛋白质表达水平均升高。结论 FOSL1基因沉默抑制MDA-MB-231细胞的增殖和侵袭,可能与该细胞中PRDM10基因的去甲基化有关。

Objective To construct the silencing vector of FOS like antigen 1 （FOSL1） gene, and study the effects of FOSL1 on cell proliferation, cell invasiveness and the methylation level of PRDM10 gene in breast cancer cell line MDAMB-231. Methods The FOSL1 silencing vector of gene pLVX-shRNA-FOSL1-shRNA was purchased. The FOSL1 silencing vector and the empty vector were separately transfected into MDA-MB-231, which were regarded as transfection group and empty group, respectively. Untransfected MDA-MB-231 was used as control group. FOSL1 was verified by PCR in MDA-MB-231. The cell proliferation ability and cell invasion ability of MDA-MB-231 were detected by MTT and Transwell assay, respectively. MSP was used to detect the methylation status of PRDM10 gene. The mRNA and protein expression levels of PRDM10 gene were detected by Q-PCR and Western-blot assay. Results MTT results showed that the optical density （OD） values were significantly lower in transfection group compared with those of control group at 24 h, 48 h and 72 h （all P〈0.05）, and the same as those compared with empty group at 48 h and 72 h （both P〈0.05）. Compared with empty group and control group, Transwell assays showed that the cell invasive abilities of MDA-MB-231 were decreased in transfection group （both P〈0.05）, and MSP assay showed that the methylation of PRDM10 gene was decreased in MDA-MB- 231, and Q-PCR and Western-blot tests showed that the expressions of PRDM10 gene were increased in mRNA level and in protein level. Conclusion Silencing of FOSL1 gene inhibits the proliferation and invasion of MDA-MB-231 cells, which might be related to the demethylation of PRDM10 gene in the cells.