The Roles of Granule Exocytosis and Fas/Fas Ligand Pathway in the Cytotoxicity of Mycobacterium Tuberculosis Antigen Activated Human ??T Cells to Tumor Cell Lines / 中国肿瘤生物治疗杂志

ABSTRACT

Objective: To investigate the mechanisms of anti-tumor cytotoxicity of human ??T cells activated by Mycobacterium tuberculosis antigen (Mtb-Ag) . Methods: The peripheral blood mononuclear cells (PBMC) of healthy donors were stimulated by Mtb-Ag in vitro to activate preferentially ??T cells that were expanded in IL-2 contained medium.The high purified ??T cells were isolated by the positive selection on MACS separator. The expression of mRNA of perforin, granzyme B and Fas Ligand in purified ??T cells were measured by RT-PCR technique. Fas Ligand on ??T cells and Fas molecule on tumor cells were detected by flow cytometry. The MTT assay was used to measureanti-tumor cytotoxicity of ??T cells. The inhibitory effects of concanamycin A (CMA) for perforin/granzyme pathway, and brefeldin A (BFA) for Fas/Fas ligand pathway on cytotoxicity of ??T cells were observed. Results: The mRNA of perforin and granzyme B and Fas ligand were highly expressed in purified ??T cells. The cytotoxicity of ??T cells against K562 and PG that expressed low level of Fas molecule (6.5% and 7.8%) was remarkably inhibited by pretreatment of the ??T cells with CMA (200 nmol/L) by 72% and 76%, respectively, while the BFA(30 ?mol/L) showed less inhibitory effects by 32% and 25%, respectively; However, it was showed that the BFA (30 ?mol/L) preferentially inhibited the cytotoxicity of ??T cells against tumor cell lines Raji and A375 that expressed high level of Fas molecule on cell surface (98.5% and 70.6%, respectively).Conclusion: Preferentially, granule exocytosis was a main mechanism by which the Mtb-Ag activated ??T cells showed cytotoxicity aganist tumor cells that expressed low level of Fas molecule, whereas both Fas/Fas ligand pathway and granule exocytosis were involved in the cytotoxicicty of Mtb-Ag activated ??T cells against the tumor cells that expressed high level of Fas molecule.