肌动蛋白样分子参与TNFRⅡ介导的TM-TNF-α杀瘤活性的研究

The co-action of actin-like protein with cytotoxicity of TM-TNF-α mediated by TNF receptor Ⅱ

目的 寻找并研究参与TM TNF α杀瘤的协同分子 ,进一步揭示TM TNF α发挥生物学作用的特征 ,阐明其与S TNF α功能差异的分子机制。方法 利用免疫共沉淀法、质谱分析、Westernblot及细胞毒实验探索参与TM TNF α杀瘤功能的协同作用分子并确定其性质。结果 TM TNF α免疫共沉淀产物中发现一相对分子质量 (Mr)为 4 3× 10 3蛋白分子 ,用质谱分析及Westernblot证实该分子属于肌动蛋白家族 ;用抗肌动蛋白抗体封闭效应细胞和 或靶细胞后 ,TM TNF α对主要表达TNFRⅡ的肿瘤细胞HL 6 0的杀伤作用显著下降 ,而对主要表达TNFRⅠ的肿瘤细胞MCF 7则无影响。结论 Mr为 4 3× 10 3的肌动蛋白样分子可能参与并增强TM TNF α通过TNFRⅡ介导的杀瘤功能。

Objective The co-action of actin-like protein with the cytotoxicity of TM-TNF-α was studied to discover the characteristic functions of TM-TNF-α and to illustrate the molecular mechanisms to explain the differences of biological effects between the two types of TNF-α. Methods Immunoprecipitation, peptide spectrum analysis, Western blot and cytotoxicity bioassay were used to substantiate and define the co-action molecules to explain biological function of TM-TNF-α. Results A 43kD protein was found in the immunoprecipitate of target cell stimulated by TM-TNF-α. It was identified as a protein of actin family by peptide spectrum analysis and Western blot. The preincubation of effect cells and/or target cells with anti-actin antibody resulted in significant inhibition of the cytotoxicity of TM-TNF-α on HL-60 cell line expressing TNFRⅡ, but had no effect on the cytotoxicity of TM-TNF-α on MCF-7 cell line which expressing mainly TNFRⅠ. Conclusion The results suggested that the 43kD actin-like protein may costimulate TNFRⅡ and enhance the cytotoxicity of TM-TNF-α.