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目的:建立分泌mRANTES的小鼠肝癌细胞,并观察其体内致瘤性。方法:mRANTEScDNA被克隆入pBabepuro逆转录病毒载体,构建的重组逆转录病毒载体pBabepuromRANTES转染包装细胞,嘌呤酶素抗性细胞培养上清感染小鼠肝癌细胞系Hepa1-6,免疫组化检测Hepa1-6、Hepa1-6mRANTES的mRANTES蛋白的表达。绘制Hepa1-6mRANTES与Hepa1-6的生长曲线观察细胞的生长。琼脂糖凝胶打孔法体外观察Hepa1-6mRANTES分泌蛋白对小鼠脾细胞的趋化作用。观察Hepa1-6mRANTES体内致瘤性。结果:构建了重组逆转录病毒载体pBabepuromRANTES,Hepa1-6不表达mRANTES,Hepa1-6mRANTES表达mRANTES蛋白。Hepa1-6mRANTES与Hepa1-6的生长曲线基本一样。Hepa1-6mRANTES分泌了对小鼠脾细胞有趋化作用的分子。Hepa1-6mRANTES体内致瘤性降低。结论:Hepa1-6mRANTES能产生mRANTES,mRANTES不改变细胞体外生长状况,Hepa1-6mRANTES产生的mRANTES有趋化活性,mRANTES能使Hepa1-6mRANTES致瘤性降低。
Objective: To establish murine hepatocellular carcinoma cells secreting mRANTES and observe its in vivo tumorigenicity. METHODS: The mRANTES cDNA was cloned into the pBabepuro retrovirus vector and the recombinant retroviral vector pBabepuromRANTES was constructed and transfected into the packaging cells. The purine resistant cell culture supernatant was used to infect the HepG1 hepatoma cell line Hepa1-6. The expression of Hepa1- 6, Hepa1-6mRANTES mRANTES protein expression. The growth curves of Hepa1-6mRANTES and Hepa1-6 were drawn to observe the cell growth. Chemotaxis effect of Hepa1-6mRANTES secreted protein on mouse spleen cells in vitro by agarose gel drilling. In vivo tumorigenicity of Hepa 1-6mRANTES was observed. Results: Recombinant retroviral vector pBabepuromRANTES was constructed, Hepa1-6 did not express mRANTES, and Hepa1-6mRANTES expressed mRANTES protein. Hepa1-6mRANTES and Hepa1-6 growth curve is basically the same. Hepa 1-6mRANTES secrete molecules that chemotact to mouse splenocytes. Hepa1-6mRANTES reduced in vivo tumorigenicity. Conclusion: Hepa1-6mRANTES can produce mRANTES, mRANTES does not change the cell growth in vitro, mRANTES produced by Hepa1-6mRANTES has chemotactic activity, and mRANTES can reduce the tumorigenicity of Hepa1-6mRANTES.