PIN1反义核酸对乳腺癌MCF-7细胞增殖及周期的影响.pdfVIP

肿瘤防治研究2007年第34卷第12期 ·917· PIN1反义核酸对乳腺癌MCF一7细胞增殖及 周期的影响 周金华，朱 涛，李红雨，徐 钢，卢运萍，马 丁 Effect of PIN1 Antisense Nucleotide on Cell Proliferation and Cycle of Human Mammary Cancer Cell Line MCF-7 ZHOU Jin-hua，ZHU Tao，LI Hong-yu，XU Gang，LU Yun—ping，MA Ding Molecular Tumor Center，Tongji Medical College，Huazhong University of Science and Technology， Wuhan 430030，China Corresponding Author：MA Ding，E-mail：dma@tjh．tjmu．edu．cn Abstract：Objective To observe the effeCt of PIN1 antisense nucleotide on cell proliferation and cycle of human mammary cancer cell line MCF-7．Methods The eukaryotic vector named pPINas，expressing PIN1 antisense nucleotide，was constructed．MCF_7 was transfected with pPINas and selected in the cul— ture with G41 8．The selected clone MCF-7PINas was checked for expression of PIN1 bv RT PCR and Westem blot．The proliferation of cells and the change of cell cycle in clone MCF-7PINas were investiga— ted by～rrr and Flow Cytometry respectively．Results The expression of PIN1 at mRNA and protein level in MCF_7PINas clone was down-regulated remarkably．MTT showed the proliferation of MCF_ 7PINas was retarded obviously contrasting to M F_7(P0．05)．But the cell cycle distribution had no significant change(PO．05)．Conclusion Since blocking PIN1 expression with antisense nucleotide could depress the proliferation activity of MCF-7 remarkably，PIN1 may be a potential target of gene therapy for human ma mmary cancer． Key words：PIN1；Antisense nucleotide；Mammary cancer 摘 要：目的 本研究利用PIN1反义核酸阻断乳腺癌MCF-7细胞中PIN1表达，观察其对增殖及周期 的影响。方法 构建PIN1反义核酸真核表达质粒pPIN1as，用脂质体转染法将重组质粒转染MCF-7 细胞，G418筛选稳定表达重组质粒的克隆，RT_PcR检测PIN1基因表达水平，Western blot检测PIN1 蛋白的表达，MTT检测细胞增殖状况，流式细胞术检测细胞周期。结果 稳定表达PIN1反义核酸的 MCF-7细胞内PIN1基因表达在mRNA水平和蛋白水平都显著降低；MTT实验显示MCF-7PINas细 胞的增殖速度较 MCF一7细胞明显减慢(P0．05)，但F(：M显示MCF-7PINas细胞和MCF-7细胞的周 期分布差异无统计学意义(P0．05)。结论 阻断PIN1可以显著抑制乳腺癌MCF-7细胞的增殖活 性，提示PIN1可能成为乳腺癌基因治疗的新的靶点。 关键词：PIN1；反义核酸；乳腺癌 中图分类号：R737．9