PIN1反义核酸转染抑制乳腺癌MCF.docVIP

PIN1反义核酸转染抑制乳腺癌MCF-7细胞的增殖 【摘要】? 　　目的 研究利用Pin1反义核酸阻断乳腺癌MCF-7细胞中Pin1的表达，并观察其对增殖的影响。 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。 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(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，这为乳腺癌的基因研究及治疗提供了新思路。 【关键词】? Pin1 反义核酸 乳腺癌 　　Effect of P1N1 antisense nucleotide on proliferation of human breast cancer cell line MCF-7. 　　HU Chun-xia, ZHOU Jin-hua, ZHU Tao, et al. 　　(Molecular Tumor Center, Tongji Medical College of Huazhong University of Science and Technology, Wuhan 430030, Hubei, P. R. China；Department of Obstetrics and Gynecology, Affiliated Hospital of? Hainan Medical College,Haikou,570102, P.R. China)??? 　　Abstract：Objective? To observe the effect of Pin1 antisense nucleotide on cell proliferation of human breast 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 in clone MCF-7PINAS were investigated by MTT.? 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 significantly retarded contrasting to MCF-7 (P0.05).? Conclusion? Blocking the expression of Pin1 with antisense nucleotide can signficntly depress the proliferation activity of MCF-7 to have offered new insight in gene research and treatment of human breast cancer.?? 　　Key words：Pin1;? Antisense nucleotide;? Breast cancer 　　PIN1(Peptidyl-Prolyl Cis/Trans Isomerase,NIMA-interating,1)是一种肽脯氨酰顺反异构酶，能够特异性地识别磷酸化后的丝/苏脯氨酰基序，而蛋白序列中丝/苏脯氨酰基序的磷酸化是一个重要的细胞内信号机制，它主要关系到细胞的增殖和转化。Pin1特异性识别并催化磷酸化的丝/苏脯氨酰，使蛋白质发生顺反异构反应，从而调节蛋白的功能。这种复杂信号机制的失调可导致细胞转化和肿瘤发生［1］。近年来发现Pin1在人类多种肿瘤组织中存在Pin1基因过度表达［2］。在乳腺癌、前列腺癌中高表达并提示预后不良［3,