Studies on differentiation of Th2 functional subset among adult peripheral blood cells in vitro

Objective: To study expression of Th1/Th2 type cytokines in human peripheral αβT cells activated with CD3 monoclonal antibody(CD3mAb),cultured in neutral and Th2 type polarizing condition.Methods: Peripheral blood mononuclear cells(PBMC) were stimulated with CD3mAb to generate CD3mAb activated T cells(CD3AT);CD3AT cells were cultured in medium with IL-2 as neutral culturing condition,or with IL-4 plus anti-IL-12 neutralizing Ab as Th2 type polarizing condition for three weeks.Fresh isolation PBMC and cultured CD3AT for 7 to 21 days in neutral and Th2 type polarizing condition were stimulated with Phorbo 12-myristate 13-acetate(PMA),ionomycin and monensin.Intracellular cytokines of cell were stained with multicolor fluorescence mAb.The percentage of Th1 and Th2 expressing cell among fresh PBMC or CD3AT were measured by flow cytometry.Results: The percentages of IFN-γ and IL-4 producing T cells among fresh isolation PBMC were 23.6% and 2.4%,respectively.Compared to the neutral condition,in the culture of Th2 polarizing condition for 3 weeks,the percentages of IFN-γ⁺ producing cells among CD3AT sharply decreased(P<0.001) and that of IL-4 producing T cells had significant increase(P<0.001),and that of IFN-γ⁺IL-4⁺αβT cells modestly increased(3.6% to 6.1%).Conclusions: αβT cells in human peripheral blood could be developed as type-2 functional subset in Th2 polarizing condition in vitro.