Effect of Staphylococcal Enterotoxin A on the Distribution and Clonal Expansion of TCR Vβ Subfamilies and the Cytotoxicity of T Cells Stimulated by PML-RARα Peptid.

Bacterial superantigens (SAgs) are potent T cell activators. Superantigens stimulate some population of T cells, based on Sag binding to specific T cell receptor variable region β chains, a property that distinguishes SAgs from conventional T cell mitogens. The biologic properties of SAgs make them attractive for use in immunotherapy. However, the T cell stimulatory effects of SAgs cannot be confined to specific antigens, its important role might be to enhance the immune effect for tumor antigens. PML-RARα peptide is a specific antigen associated with acute promyelocytic leukemia, which can induce specific proliferation of T cells from healthy human donor in vitro. But the immunogenicity of PML-RARα peptide is weak based on our previous study. In order to investigate the differentiation and proliferation of T cells induced by PML-RARα peptide that combined with Staphyloccucal enterotoxins A(SEA), T cell phenotype, TCR Vβ repertoire usage, clonality and cytotoxicity of T cells were analyzed. Peripheral blood mononuclear cells from healthy donors were cultured with PML-RARα peptide and SEA for 20 days. After induction, the distribution and clonal expansion of TCR Vβ subfamilies of T cells were examined by RT-PCR and GeneScan. The cytotoxicity of T cells induced against NB4 cell lines were examined by Cell Counting Kit-8 (CCK-8). The CD4 and CD8 surface marker on the harvested CD3⁺ T cells were detected by flow cytometry (FCM). And cytokine levels of IFN-γ, IL-4 and IL-10 in the cultured supernatants were measured by Enzyme-Linked Immunosorbent Assay (ELISA) as well. The results showed that restricted expression of Vβ repertoire (11 Vβ subfamilies) was found, which were Vβ1, Vβ2, Vβ3, Vβ6, Vβ7, Vβ8, Vβ13, Vβ14, Vβ16, Vβ17, Vβ19 subfamilies. Clonal expanded T cells could be identified in some Vβ subfamilies, which were preferentially used in Vβ13 and Vβ14. The specific cytotoxicity of T cells induced by PML-RARα peptide with SEA was higher than that from PML-RARα peptide alone induction group against NB4 cells. The FCM assay showed that the activation of CD8⁺ T cells was in priority induced by PML-RARα peptide whether with SEA or not, while it came out to be an equal between CD4⁺ and CD8⁺ T cells induced only by SEA. And the IFN-γ level of Th1-type cytokine was higher than that of the Th2-type cytokines (IL-4 and IL-8) in both cultured supernatants of T cells induced by PML-RARα peptide with or without SEA. In conclusion, PML-RARα peptide can induce specific T cells with clonal expansions of TCR Vβ subfamilies. SEA could enhance the induction of the specific PML-RARα peptide. The T cells induced by combination of PML-RARα peptide and SEA have higher specific antileukemia effect.