人白细胞介素-16真核表达载体的构建及其对Th2型肿瘤细胞的促逆转作用

CONSTRUCTION OF HUMAN IL-16 EUKARYOTIC EXPRESSION VECTOR AND ITS FUNCTION OF SWITCHING Th2 TYPE TUMOR CELLS

目的 :构建人IL 16真核表达载体 ,分析其对Th2型肿瘤细胞的逆转作用。方法 :用RT PCR技术从人外周血单个核细胞中获取IL 16cDNA ,插入PUC 18T载体并测序 ,构建并鉴定真核表达载体PcDNA3 IL 16 ,转染KarpasT淋巴瘤细胞 ,分析阳性克隆中IFNγ ,IL 2 ,IL 4 ,IL 6 ,IL 13,TNFα和IL 16等细胞因子的表达状况。结果 :序列测定显示所克隆的人IL 16cDNA与基因库中所登录的序列完全一致 ,EcoRI和BamHI双酶切及PCR鉴定显示所构建的真核表达载体PcDNA3 IL 16完全正确 ,RT PCR显示转染PcDNA3 IL 16的KarpasT淋巴瘤细胞高表达IL 16mRNA ,同时 ,Th1类细胞因子IFNγ表达明显增高 ,Th2类细胞因子IL 4 ,IL 6 ,IL 13表达降低 ,MTT法显示KarpasT淋巴瘤细胞的增殖受到明显抑制。结论 :成功构建了真核表达载体PcDNA3 IL 16 ,转染KarpasT淋巴瘤细胞后使其细胞因子类型由Th2型向Th0型逆转 ,且生长受到明显抑制 ,表明向肿瘤细胞转染人IL 16是一种有用的肿瘤生物治疗方法。

Objective:To construct human IL 16 eukaryotic expression vector,and analyse its function of switching Th2 type tumor cells.Methods:The human IL 16 cDNA was amplified by RT PCR from peripheral blood mononuclear cells,and cloned into the PUC 18 T vector.Then the eukaryotic expression vector PcDNA3 IL 16 containing IL 16 cDNA was constructed and identified.The Karpas T lymphoma cells were transfected with PcDNA IL 16.Cytokines IFNγ,IL 2,IL 4,IL 6,IL 13,TNFα and IL 16 were detected by RT PCR from the positive cell clones.Results:The sequence of cloned human IL 16 cDNA completely matched with that of human IL 16 cDNA in the gene bank.RT PCR indicated the IL 16 mRNA was highly expressed in the Karpas cells after the recombinant vector PcDNA3 IL 16 was transfected into Karpas T lymphoma cells.The cells were switched from the expression of Th2 cytokines to Th0 type.The biological activity through MTT assay showed the proliferation of Karpas cells were sharply inhibited.Conclusion:The eukaryotic expression vector PcDNA3 IL 16 containing IL 16 cDNA was successfully constructed.The cytokine pattern of Karpas T lymphoma cells can be changed from Th2 type to Tho type by transfecting PcDNA IL 16.These results suggest that is a useful method to transfect IL 16 gene into tumor cells for tumor biotherapy.