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目的:检测特异性封闭高度恶性人卵巢癌细胞株Sw626、A2780中畸胎瘤源性生长因子(PC -cell -devivedGrowthFactor,PCDGF)的表达,并观察其对增殖能力的抑制效应。方法:用RT -PCR技术扩增PCDGF基因cDNA保守序列,经pGEM -TEasy载体克隆后双酶切,反向插入哺乳动物真核表达载体pcDNA3 1 ( +)构建反义PCDGF核酸载体。并通过脂质体转染法转染人卵巢癌细胞株Sw626、A2780,用RT- PCR和Westernblot技术检测转染前后PCDGFmRNA和蛋白的表达,MTT实验检测其增殖能力的变化。结果:PCDGF反义核酸载体转染株的mRNA和蛋白水平明显低于对照组,Sw626的抑制率分别为50%、72%;A2780分别为53%、70%。同时增殖能力亦明显降低,其抑制率Sw626及A2780分别为71%、73%。结论:PCDGF反义核酸在高度恶性人卵巢癌细胞株Sw626及A2780中起了特异性封闭作用。为进一步研究PCDGF的分子生物学特性、致瘤机理及今后利用反义PCDGF核酸载体治疗卵巢癌提供了实验和理论模型。
OBJECTIVE: To detect the expression of PCDGF in Sw626 and A2780 cells, and to observe its inhibitory effect on the proliferation of human ovarian cancer cell lines. Methods: The cDNA sequence of PCDGF gene was amplified by RT-PCR and cloned by pGEM-TEasy vector. The antisense PCDGF DNA vector was constructed by reverse insertion of mammalian eukaryotic expression vector pcDNA3 1 (+). The transfected human ovarian cancer cell lines Sw626 and A2780 were transfected by lipofectamine. The expression of PCDGF mRNA and protein was detected by RT-PCR and Western blot. The proliferation of PCDGF was detected by MTT assay. Results: The mRNA and protein levels of PCDGF antisense vector were significantly lower than those of control group. The inhibition rate of Sw626 was 50% and 72%, respectively. The A2780 was 53% and 70% respectively. At the same time, the proliferation ability was also significantly reduced. The inhibition rates of Sw626 and A2780 were 71% and 73% respectively. CONCLUSION: PCDGF antisense nucleic acid plays a specific blocking role in the highly malignant human ovarian cancer cell lines Sw626 and A2780. It provides experimental and theoretical models for further study of the molecular biological characteristics of PCDGF, tumorigenic mechanism and the future use of antisense PCDGF nucleic acid vector in the treatment of ovarian cancer.