Fig. 8. Proliferation of PHA-stimulated (A) and Wa HRV-stimulated (B) sort-purified splenic CD4+ T cells co-cultured with each γδ T cell subset.

Cells collected from the co-cultures (see Fig. 7 legend) were stained with antibodies to CD3, CD4, and BrdU to measure proliferation of CD4+ αβ T cells by flow cytometry. Frequencies of proliferating cells were defined as BrdU+CD4+ T cells among total CD3+ T cells (Fig. 8A) (n = 6). Three γδ T cell subsets and CD4+ αβ T cells were also sort-purified from MNCs isolated from spleen of Gn pigs inoculated with two oral doses of attenuated HRV and challenged with virulent HRV. The co-cultures of CD4+ αβ T cells with each γδ T cell subset or without were stimulated with Wa HRV antigen for 5 days and the cells were collected for CD3, CD4, and BrdU staining to measure proliferation of CD4+ αβ T cells among CD3+ T cells (Fig. 8B) (n = 3). Data are presented as mean concentration ± standard error of the mean. Different letters on top of bars indicate significant differences in frequencies of proliferating CD4+ αβ T cells among the cell cultures (Kruskal-Wallis test, p < 0.05), while shared letters indicate no significant difference.