摘要
目的研究外源FHIT基因对不同FHIT基因背景肺癌细胞系恶性增殖的影响并探讨抑癌机理。方法用Lipofectamine介导PEGFP-FHIT真核表达质粒转染受体细胞系并建立稳定转染细胞系。RT-PCR、免疫组织化学及Western blot方法鉴定转染细胞中外源FHIT基因和蛋白的表达。细胞集落形成实验研究外源FHIT基因对肺癌细胞恶性增殖的影响,流式细胞仪分析细胞周期和凋亡。Western blot检测母系及转染细胞系中p21Waf/cip蛋白的表达。结果3种受体细胞转染外源FHIT基因后均检测到FHIT mRNA和蛋白的表达。集落形成实验801D-FHIT(46.3%)、A2-FHIT(43.7%)、A549-FHIT(32.7%)细胞集落形成率比801D(58.2%)、A2(60.3%)、A549(52.8%)的低,差异有显著性(P<0.01)。FACS分析细胞周期显示:801D-FHIT、A2-FHIT和A549-FHIT细胞分别出现了G1、S和G2/M期阻滞。转染FHIT基因的801D-FHIT、A2-FHIT和A549-FHIT细胞p21Waf/cip蛋白表达增加。结论外源FHIT基因能够抑制不同FHIT基因背景的肺癌细胞的恶性增殖,并引起细胞周期改变,不同FHIT基因背景的细胞产生细胞周期阻滞的时间不同。FHIT基因通过促进p21Waf/cip蛋白的表达来调节细胞周期。
Objective To investigate the effects of FHIT gene on the malignant growth of lung cancer cells with different gene background and its mechanism of cancer inhibition. Methods A mammalian expression vector PEGFP-FHIT was transfected into the lung cancer lines by lipofectamine. We tested the transfected cancer cell lines by RT-PCR, immunochemical stain and Western blot. The inhibition growth efficacy of extraneous FHIT was evaluated by clonogenic survival assay and flow cytometry. The expression of p21^waf/cip protein was examined by Western blot. Results All of the transfected cancer cell lines had the expression of FHIT gene and protein. The clonal formation rate of 801D-FHIT(46.3% ), A2-FHIT(43.7% ), A549-FHIT (32.7 % ) was lower than that of 801D (58.2%), A2 (60.3 % ), A549 (52.8 % ) ( P 〈 0.01 ) . Cell cycle analysis by FACS showed that 801D-FHIT, A2-FHIT and A549-FHIT cells were blocked at G1, S and G2 phase, respectively. A significant increase in p21^waf/cip protein expression was noticed in FHIT expression clones 801D-FHIT, A2-FHIT and A549-FHIT compared with that of the control parental cell lines. Conclusion Extrogenous FHIT gene can suppress the proliferation in different FHIT gene status and regulate the cell cycle. There is a different cell cycle arrest in three sorts of lung cancer cell lines with different expression of FHIT gene. Our data suggests that observed growth-inhibitory effect in FHIT-reexpressioning cells could be related to cell cycle arrest and linked to the increased p21^waf/cip protein expression.
出处
《解剖学报》
CAS
CSCD
北大核心
2005年第5期513-518,共6页
Acta Anatomica Sinica