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目的为搞清 GAM在 gp130介导的信号传导中所起的作用。方法选择在 IL - 6刺激下出现分化和增殖抑制的M1细胞系 ,分别用正向和反向插入 GAM c DNA的真核表达载体 p EF- BOS与 p SV2 - neo质粒共转染 ,经 G418选择培养筛选得到稳定转染的细胞 ,并用半套式 PCR方法证实了 GAM基因与这两种细胞基因组 DNA的整合。结果流式细胞仪分析表明正向 GAM c DNA转染的 M1细胞中 GAM的表达水平明显升高 ,3H掺入实验表明 GAM表达水平升高使 M1细胞对 IL - 6的反应性下降。结论 GAM高表达 M1细胞的建立为深入研究 GAM在 gp130介导的信号传导中的作用提供了良好的实验模型
The purpose is to elucidate the role of GAM in gp130-mediated signaling. Methods The M1 cell line with differentiation and proliferation inhibition stimulated by IL - 6 was selected and transfected with the eukaryotic expression vector p EF-BOS and p SV2 - neo respectively forward and reverse GAM c DNA. After G418 The cells that were stably transfected were selected and cultured, and the integration of the GAM gene with the genomic DNA of both cells was confirmed by the semi-nested PCR method. Results Flow cytometry analysis showed that the expression level of GAM in M1 cells transfected with GAM c DNA was significantly increased. 3H - incorporation assay showed that the increased expression of GAM decreased the reactivity of M1 cells to IL - 6. Conclusion The establishment of GAM-overexpressing M1 cells provides a good experimental model for further study on the role of GAM in gp130-mediated signaling