PIN1 反义核酸对乳腺癌MCF-7 细胞增殖及周期的影响

 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pPINlas，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-7细胞明显减慢（P〈0．05），但FCM显示MCF-7PINas细胞和MCF-7细胞的周期分布差异无统计学意义（P〉0．05）。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。

Effect of PIN1 Antisense Nucleotide on Cell Proliferation and Cycle of Human Mammary Cancer Cell Line MCF-7

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 culture with G418.The selected clone MCF-7PINas was checked for expression of PIN1 by RT-PCR and Western blot.The proliferation of cells and the change of cell cycle in clone MCF-7PINas were investigated by MTT and Flow Cytometry respectively.Results The expression of PIN1 at mRNA and protein level in MCF-7PINas clone was downregulated remarkably.MTT showed the proliferation of MCF-7PINas was retarded obviously contrasting to MCF-7(P<0.05).But the cell cycle distribution had no significant change(P>0.05).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 mammary cancer.