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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 murine hepatoma cell line secreting m MIP-1α and observe its tumorigenicity in vivo. METHODS: m MIP-1α c DN A was cloned into p Babe puro retroviral vector and the constructed recombinant retroviral vector p Babe puro- m MIP-1α was transfected into packaging cells and puromycin-resistant cell culture supernatant was infected. Hepatocellular carcinoma cell line Hepa1-6, RT-PCR and immunohistochemistry were used to detect the expression of m MIP-1α m RNA and m MIP-1α protein of Hepa1-6, Hepa1- 6-m MIP-1α, respectively. The cell growth was observed by plotting the growth curves of Hepa1-6-m MIP-1a and Hepa1-6. The chemotaxis of mouse spleen cells by Hepa1-6-m MIP-1alpha secretory protein was observed in vitro by agarose gel punching. Hepa1-6-m MIP-1alpha in vivo tumorigenicity was observed. RESULTS: A recombinant retroviral vector p Babe puro- m MIP-1 alpha was constructed. Hep-a1-6 did not express m MIP-1 alpha m RNA and protein, and Hepa1-6-m MIP-1 alpha expressed m MIP-1 a m RNA and protein. The growth curve of Hepa1-6-m MIP-1a and Hepa1-6 is basically the same. Hepa1-6-m MIP-1α secretes molecules that have a chemotactic effect on mouse spleen cells. Hepa1-6-m MIP-1α has reduced tumorigenicity in vivo. Conclusion: Hep1 - 6 - m MIP-1α can produce m MIP-1 α, m MIP-1 α does not change the growth of cells in vitro, and M MIP-1 α produced by Hepa1- 6 - m MIP-1 α has chemotaxis activity, m MIP- 1α can reduce the tumorigenicity of Hepa1-6-m MIP-1α.