脆性组胺酸三联体基因对人肺腺癌A549细胞恶性表型的影响  

Ectogenous fragile histidine triad gene inhibits the malignant phenotype of human lung cancer cell line A549 in vitro and in vivo

作　　者：张立群[1] 汪蕙[1] 赖百塘[1] 岳文涛[1] 湛秀萍[1] 杨学惠[1] 张春燕[1] 张同梅[1] 

机构地区：[1]北京市结核病胸部肿瘤研究所细胞生物学实验室,101149

出　　处：《中华结核和呼吸杂志》2005年第5期333-336,共4页Chinese Journal of Tuberculosis and Respiratory Diseases

摘　　要：目的研究外源性脆性组胺酸三联体(FHIT)基因在体内外对人肺腺癌A549细胞恶性表型的影响。方法用脂质体介导外源FHIT基因转染A549细胞,建立单克隆细胞系FHITA-549和PEGFP-A549。逆转录聚合酶链反应(RTPCR)、免疫组化方法检测外源FHIT基因在A549细胞的表达状况。采用细胞生长曲线、集落形成试验、流式细胞仪及裸鼠移植瘤试验研究外源FHIT基因对A549细胞体外增殖、凋亡、细胞周期和体内成瘤性的影响。结果RTPCR试验证实FHIT-A549中有FHITmRNA表达,免疫组织化学染色显示FHIT-A549细胞FHIT蛋白表达强阳性,而A549细胞和转空载体的PEGFPA549细胞FHIT基因和蛋白表达均阴性。FHIT-A549细胞的集落形成率为2.6%,显著低于A549细胞的50.1%和转染空载体PEGFPA549细胞的53.6%,三者相比差异有统计学意义(P<0.01)。流式细胞仪分析显示FHIT-A549细胞95.8%阻滞在G2期。FHITA549细胞移植瘤的瘤重为(0.04±0.03)g,显著低于A549细胞的(0.24±0.11)g和转染空载体PEGFPA549细胞的(0.25±0.07)g,三者差异有统计学意义(P<0.01)。结论A549细胞内外源FHIT基因的转导并表达能显著抑制其恶性增殖和分裂,诱导其凋亡,调节其细胞周期、抑制成瘤性。Objective To investigate the inhibition effects of fragile histidine triad (FHIT) gene on the malignant growth of A549 cell line. Methods A mammalian expression vector PEGFP-FHIT was constructed and transfected into the A549 cell line by lipofectamine. Then the transfected cell line was screened by G418. Individual G418-resistant colonies were isolated by limited dilution. The moloclonal transfected cell line was screened by RT-PCR and immunochemical staining. The inhibition growth efficacy of extraneous FHIT was evaluated by clonogenic survival assay ,flow cytometry and heteroplastic transplant on nude mice. Results Presence of extraneous FHIT gene in FHIT-A549 cell was proved by RT-PCR. Immunochemical stain demonstrated that the expression of extraneous FHIT protein was positive in FHIT-A549 cell and negative in PEGFP-A549 cell and A549 cell. The clonal formation rate of FHIT-A549(2.6%) was significantly lower than that of A549 cell (50.1%) and PEGFP-A549 cell (53.6%, P<0.01). FHIT-A549 cell (95.8%) was blocked in G 2 phage. Tumorigenicity of A549 cells in nude mice was greatly inhibited by expression of ectogenous FHIT gene. The weight of tumor was significantly lower in FHIT-A549 cell (0.04±0.03) than in A549 cell (0.24±0.11) and PEGFP-A549 cell (0.25±0.07,P<0.01). Conclusions Reintroduction of the expression of ectogeneous FHIT gene can obviously suppress the proliferation and tumorigenicity in human lung cancer cell line A549 and induce apoptosis. The data demonstrate oncosuppressive properties of FHIT gene.

关 键 词：脆性组胺酸三联体 细胞恶性表型 人肺腺癌 逆转录-聚合酶链反应(RT-PCR) A549细胞 FHIT基因 免疫组织化学染色 流式细胞仪分析 免疫组化方法 细胞生长曲线 集落形成试验 细胞体外增殖 mRNA表达 细胞周期 蛋白表达 脂质体介导 

分 类 号：R734.2[医药卫生—肿瘤] R737.9[医药卫生—临床医学]