背景与目的:有研究发现,肝癌缺失基因1(DLC1)在多种肿瘤中低表达或不表达,其通过调节黏着斑激酶(FAK)、促分裂原活化蛋白激酶(MAPK)等影响肿瘤细胞的凋亡.而对于DLC1在卵巢癌中的表达及作用等的研究甚少,本实验采用DLC1基因转染该基因表达缺失的人卵巢癌多药耐药细胞系OVCAR-3,观察转染前后该细胞对顺铂耐药性及FAK、p38MAPK的变化.方法:将OVCAR-3细胞分为3组,空白组为未经处理的OVCAR-3细胞,阴性对照组转染空质粒pEGFP-C3,实验组转染重组质粒pEGFP-C3-DLC1.RT-PCR和Western blot检测各组细胞中DLC1基因和蛋白的表达,四甲基偶氮唑蓝(MTT)法测定各组细胞对顺铂的半数抑制浓度(IC50),Western blot观察FAK、p38蛋白及其磷酸化水平的变化,流式细胞仪测定顺铂处理前后各组细胞凋亡率及周期分布改变.结果:DLC1基因和蛋白在实验组表达而空白对照组和阴性对照组均未见表达,实验组细胞与其他两组细胞相比,对顺铂的IC50较低(4.02 vs 4.99/4.90 μmol/L,P＜0.01),p-p38蛋白表达上升而p-FAK蛋白表达下降(3.02 vs 1.52/1.61,3.13 vs 9.03/8.99,P＜0.01),顺铂处理前后实验组与其他两组细胞相比凋亡率增加(8.97% vs 1.81%/1.95%,30.68% vs 18.03%/20.33%,P＜0.01),G1期细胞比例增加(65.80% vs60.82%/59.80%,66.48% vs 55.42%/53.94%,P＜0.05).结论:转染DLC1基因可使OVCAR-3G1期细胞比例及凋亡率升高,对顺铂敏感性增加,该作用可能依赖于细胞内外源性DLC1基因的表达、p-FAK表达降低和p-p38表达增强.%Background and purpose: The deleted in liver cancer gene1 (DLC1) is lowly or negatively expressed in different cancers and have an effect on the apoptosis of cancer cells duc to the regulation of focal adhesion kinase (FAK) and mitogen-actived protein kinase (MAPK). However, the expression level and role of the DLC 1 gene in ovarian cancer has been rarely studied. This study aimed to transfect the DLC1 gene into the OVCAR-3 cells, where the DLC1 gene was once negatively expressed, so as to investigate the effect of the DLC1 gene on chemoresistance as well as a variation of FAK and p38MAPK. Methods: Ovarian cancer cell line OVCAR-3 was treated with none(blank group), empty plasmid pEGFP-C3(negative control group) and pEGFP-C3-DLC 1 (experimental group). The expression of DLC1 mRNA and protein were detected separately using RT-PCR and Western blot. The IC50 of cisplatin was determined using the Methyl thiazolyl tetrazolium test (MTT). The changes of FAK and p38 protein expression and their phosphorylated status were determined by Western blot. Apoptosis and cell cycle distribution were detected by flow cytometry. Results: DLC1 gene and protein were only observed in the experiment group rather than in the other groups. The IC50 of the experiment group was lower than in the other groups (4.02 vs 4.99/4.90 μmol/L, P＜0.01).There was a higher expression of p-p38 but lower expression of p-FAK (3.02 vs 1.52/1.61, 3.13 vs 9.03/8.99, P＜0.01)after the treatment of cisplatin in experimental group. Apoptosis was significantly higher and G1 arrest was oberved only in the experimental group after the treatment of cisplatin, there were statistically differences among the groups (P＜0.01). Conclusion: The OVCAR-3 cells were more sensitive to cisplatin after transient transfection of DLC1 gene in terms of the induction of apoptosis and G 1 arrest, which might be through the upregulation of p-FAK expression and downregulation of p-p38 expression.
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