Analysis of cytokine patterns produced by individual CD4(+) lymph node cells during experimental murine leishmaniasis in resistant and susceptible mice

The concept of subdividing CD4(+) T cells into T(h)0, T(h)1 and T(h)2 cells is based on the cytokine pattern produced by long-term in vitro cultured T cell lines, However, there exists uncertainty whether this classification can also be applied to CD4(+) T cells in vivo. Herein it was investigated w hether and at which frequency T(h)0, T(h)1 and T(h)2 cells are induced in v ivo during an infection of mice with Leishmania major. Cytokine co-producti on in single IFN-gamma(+) or IL-4(+) CD4(+) T cells as well as the frequenc y of such cells were assessed in the lymph nodes (LN) of infected mice, For this purpose, T cells derived from the draining LN were activated by phorb ol myristate acetate (PMA)/ionomycin, and the intracellular cytokines IL-2, IL-4, IL-5, IL-10 and IFN-gamma were analyzed by immunofluorescence, One w eek after infection, a strong, but comparable increase of IFN-gamma(+) CD4( +) and IL-4(+) CD4(+) cells (up to 7% of all CD4(+) cells) in the LN was ob served in resistant C57BL/6 mice and susceptible BALB/c mice. IFN-gamma and IL-4 were not co-produced by single cells ('T(h)0 cells'). At rater stages of the infection, the number of IL-4(+) CD4(+) cells decreased in C57BL/6, but not in BALB/c mice. All IL-4(+) CD4(+) cells showed an unexpected phen otype, because at least half of these cells co-produced IL-2, and the major ity of the IL-4(+) CD4(+) did not co-produce the T(h)2 cytokines IL-5 and I L-10, Similar cytokine profiles were obtained when the CD4(+) T cells were stimulated by Leishmania major-antigen instead of PMA/ionomycin, This study demonstrates that 'classical' T(h)1 cells (IFN-gamma(+) IL-2(+)), but no ' classical' T(h)2 cells (IL-4(+)IL-5(+)IL-10(+)) and no T(h)0 cells (IFN-gam ma(+)IL-4(+)) are generated during L. major infection of mice in vivo.