γ干扰素cDNA在胃癌细胞中的表达研究

IFN-Y cDNA genes expressed in stomach carcinoma cells and preliminary studies in tumor gene therapy

实验用构建的正向连入人ＩＦＮ－γｃＤＮＡ片段的重组表达载体ｐＭＡＭｎｅｏ－γ－ＩＦＮ，以Ｌｉｐｏ－ｆｅｃｔｉｎ介导法将重组载体转染入胃癌细胞７９０１中，挑选出Ｇ４１８抗性克隆，地塞米松诱导其分泌ＩＦｔＩ－γ经活性测定筛选出高分泌ＩＦＮ－γ的细胞株ＲＰＭ７９０１－７（１７２５ＩＵ／ｍｌ），Ｓｏｕｔｈｅｒｎｂｌｏｔ证实ＩＦＮ－γｃＤＮＡ确已导入该细胞株中。体外培养过程中，ＲｐＭ７９０１细胞与转染了对照质粒ｐＭＡＭｎｅｏ的细胞ｐＭ７９０１及野生型７９０１细胞在形态，生长能力等方面无明显差别，而经地塞米松诱导的ＲpＭ７９０１细胞则与之有显著差异。裸鼠体内接种显示，经诱导的ＲｐＭ７９０１细胞在致瘤性上显著低于野生型７９０１细胞及ｐＭ７９０１细胞。经基因转移后可高分泌ＩＦＮ－γ的肿瘤细胞为肿瘤的基因治疗提供了资料。

A recombinant expression vector pMAMneo-Y-IFN carrying forward-linking human IFN-Y cDNA genes was first constructed , then it was transfected into 7901 stomach carcinoma cells by lipofectin . After G418 resistance .election and limiting dilution ,the positive clones were induced by dexamethasone to secret INF-Y. The clone RpM790l-7 which secreted the highest level of IFN-Y ( 1725IU/ml ) was selected by assay of IFN-Y activity in culture medium,and then identified by Southern blot. The characteristics of uninduced RpM7901 cells remained unremarkable different from that of wild-type tumor cells 7901 and pM7901 cells which had been transfected by pMAMneo in vitro, but the induced RpM7901 became very different. The result . of injected in nude mice with different kind of cells showed that the carcinogenicity of , high level IFN-Y-secreting RpM7901 cells was remarkably lower than that of wild-type 7901 cells and pM7901 cells. All above demonstrated that the high level IFN-Y-secreting tumor cells could possibly be used in tumor gene therapy.