分泌mMIP-1α趋化因子的小鼠肝癌细胞株的建立及其体内致瘤性

Establishment of a mouse hepatocellular carcinoma cell line producing mMIP-1α chemokines and the tomorigenicity of mMIP-1α transfected Hepa1-6

目的 :建立分泌 m MIP- 1α的小鼠肝癌细胞株 ,并观察其体内致瘤性。 方法 :m MIP- 1α c DN A被克隆入 p Babe puro逆转录病毒载体 ,构建的重组逆转录病毒载体 p Babe puro- m MIP- 1α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 ,RT- PCR和免疫组化分别检测 Hepa1- 6、Hepa1- 6 - m MIP- 1α的 m MIP- 1α m RNA和 m MIP- 1α蛋白的表达。绘制 Hepa1- 6 - m MIP- 1α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- 6 - m MIP- 1α分泌蛋白对小鼠脾细胞的趋化作用。观察 Hepa1- 6 - m MIP- 1α体内致瘤性。结果 :构建了重组逆转录病毒载体 p Babe puro- m MIP- 1α,Hep-a1- 6不表达 m MIP- 1α m RNA及蛋白 ,Hepa1- 6 - m MIP- 1α表达 m MIP- 1α m RNA和蛋白。 Hepa1- 6 - m MIP- 1α与 Hepa1- 6的生长曲线基本一样。 Hepa1- 6 - m MIP- 1α分泌了对小鼠脾细胞有趋化作用的分子。 Hepa1- 6 - m MIP- 1α体内致瘤性降低。 结论 :Hepa1- 6 - m MIP- 1α能产生 m MIP- 1α,m MIP- 1α不改变细胞体外生长状况 ,Hepa1- 6 - m MIP- 1α产生的 m MIP- 1α有趋化活性 ,m MIP- 1α能使 Hepa1- 6 - m MIP- 1α致瘤性降低。

Objective: To establish a mouse hepatocellular carcinoma cell line that can produce mMIP 1α and to evaluate the possibility of cancer gene therapy by mMIP 1α. Methods: mMIP 1α cDNA was cloned into retrovirus vector pBabe puro and pBabe puro mMIP 1α was constructed, then pBabe puro mMIP 1α was used to transfect packaging cells, anti puromycin cells was proliferated, the supernatant was used to infect hepa1 6, the anti puromycin clone (hepa1 6 mMIP 1α) and hepa1 6 were analysed for the expression of mMIP 1α mRNA and protein by RT PCR and immunohistochemistry respectively. The growth curve of hepa1 6 and hepa1 6 mMIP 1α was drawn. The chemotaxis of mMIP 1α produced by hepa1 6 mMIP 1α to mouse spleen cells was observed on agarose gel. C57B/L mouse was inoculated with the tumor cell and the tumorigenicity was studied. Results: Recombinant retrovirus vector pBabe puro mMIP 1α with mMIP 1α cDNA was constructed. Hepa1 6 did not produce mMIP 1α mRNA and protein, while hepa1 6 mMIP 1α could produce mMIP 1α mRNA and protein. The growth curve of hepa1 6 and hepa1 6 mMIP 1α showed no difference. The chemotaxis of mMIP 1α produced by hepa1 6 mMIP 1α to mouse spleen cells was observed. The tumorigenicity was reduced. Conclusion: A mouse hepatocellular carcinoma Hepa1 6 mMIP 1α is established and mMIP 1α can affect the tumorigenecity of hepa1 6.