Stem cell mobilization with G-CSF enhances IL-17A production by splenic CD4⁺ and CD8⁺ T cells. (A) Th1 (IFN-γ and TNF), Th2 (IL-4, IL-5), and IL-10 and (B) IL-17A in culture supernatants generated by splenocytes from naive or G-CSF–mobilized WT B6 or BALB/c mice stimulated with CD3 for 72 hours. Histograms represent mean ± SEM (n = 6-12 individual animals/group) of data from 2 or 3 replicate experiments. *P < .05. **P < .01. ***P < .001. (C) IL-17A generation from splenocytes stimulated with CD3 or concanavalin A for 72 hours from G-CSF–mobilized B6 mice FACS sort depletion of CD3⁺ or CD4⁺ and CD8⁺ T-cell populations. Whole spleen and sorted cell populations were plated in triplicate, and histograms represent mean ± SEM from one of 2 replicate experiments. ND indicates not detected. (D) IL-17F in culture supernatants were generated as in panel C. (E) Flow cytometric analysis of intracellular IL-17A and IFN-γ production in CD4⁺ and CD8⁺ T cells after culture of B6 splenocytes for 48 hours in the presence of CD3 and IL-23. Numbers within quadrants represent frequency of IL-17A– or IFN-γ–producing cells within CD4 or CD8 compartments. (F) IL-17A generation by splenocytes stimulated with CD3 for 72 hours from G-CSF–mobilized B6 mice ± FACS sort depletion of CD3⁺, CD4⁺, or CD8⁺ T-cell populations. Sorted cell populations were plated in triplicate, and histograms represent mean ± SEM from one of 2 identical experiments. (G) Time course of IL-17A and IL-17F generation in vitro. Splenocytes from nonmobilized or G-CSF–mobilized donors (4 per group) were stimulated as in panel B and IL-17A and IL-17F determined in culture supernatants taken at the time points indicated. (H) Donors (4 per group) were mobilized with a range of G-CSF doses (micrograms per day) as indicated and IL-17A determined in culture supernatants 72 hours later.