Complement C5a synergizes with TLR4 to produce serum factors that drive Th17-cell differentiation. (A) Mouse CD4⁺ T cells were activated by plate-bound anti-CD3/CD28 in the presence of 5% serum from WT mice treated with PBS, LPS, CVF, or LPS+CVF. Culture medium and recombinant IL-6 + TGF-β were used as negative and positive controls, respectively, for Th17-cell differentiation. (B) CD4⁺ T cells were activated as in panel A in the presence of 5% serum from C3^−/−, C3aR^−/−, and C5aR^−/− mice treated with LPS + CVF. (C) CD4⁺ T cells were activated as in panel A in the presence of 5% serum from WT mice treated with LPS, C5a, and LPS + C5a or from C5aR^−/− mice treated with LPS + C5a. Cells were cultured for 3 days after activation, and IFN-γ and IL-17–producing cells were detected by flow cytometry after intracellular staining. Data in panels A and B were from the same experiment, whereas data in panel C were from a separate experiment.