论文部分内容阅读
目的:研究串珠素反义cDNA质粒转染对喉癌细胞Hep-2增殖能力的影响。方法:利用阳离子脂质体作为载体,把重组真核表达载体串珠素反义cDNA质粒pAP转染喉癌细胞Hep-2,通过RT-PCR、Westernblot及MTT法检测转染后Hep-2细胞串珠素mRNA、蛋白质表达及细胞增殖水平,并观察转染后Hep-2细胞对碱性成纤维细胞生长因子(bFGF)的反应性。将Hep-2细胞分3组:①未转染的Hep-2细胞组(WT组);②空载体phβApr-neo1转染组(neo组);③串珠素反义cDNA质粒pAP转染组(pAP组)。结果:将质粒pAP成功地转入Hep-2细胞,获得了稳定表达串珠素反义cDNA基因的细胞系。串珠素mRNA和蛋白质水平在pAP组低表达,在WT组和neo组高表达,均差异有统计学意义(均P<0.01)。在0.1%FCS的RPMI1640培养基培养下,WT组、neo组和pAP组细胞的增殖率均降低,但pAP组细胞的增殖与WT组和neo组细胞相比明显减低;在0.1%FCS加1μg/LbFGF的RPMI1640培养液培养下,WT组和neo组细胞增殖接近正常,但pAP组细胞的增殖能力较弱。结论:串珠素反义cDNA质粒转染可以有效抑制喉癌细胞Hep-2的增殖能力。
Objective: To investigate the effect of antisense abacterial cDNA transfection on the proliferation of Hep-2 cells. Methods: The liposome was used as a vector to transfect the lacZ gene Hep-2 into the laryngeal carcinoma cell line pC-pAP. The expression of Hep-2 cells was detected by RT-PCR, Western blot and MTT assay MRNA, protein expression and cell proliferation, and observe the reactivity of Hep-2 cells to basic fibroblast growth factor (bFGF) after transfection. The Hep-2 cells were divided into 3 groups: ① untransfected Hep-2 cells (WT group); ② empty vector phβApr-neo1 transfected group (neo group); ③ perlecan antisense cDNA plasmid pAP transfected group pAP group). Results: Plasmid pAP was successfully transfected into Hep-2 cells and a cell line stably expressing the perlecan antisense cDNA gene was obtained. The mRNA and protein levels of beads were low in pAP group and high in WT group and neo group (all P <0.01). In 0.1% FCS RPMI1640 medium, the proliferation rate of WT, neo and pAP group cells were decreased, but the proliferation of pAP group cells was significantly reduced compared with WT and neo cells; in 0.1% FCS plus 1μg / LbFGF cultured in RPMI1640 medium, the proliferation of WT and neo cells was nearly normal, but the proliferation of pAP cells was weak. CONCLUSION: The antisense cDNA of perlecan may effectively inhibit the proliferation of laryngeal carcinoma Hep-2 cells.