FIGURE 6.

Calcitriol-conditioned DC have a reduced ability to prime the Th17 response. A, Naive B10.RIII mice were gavaged with calcitriol or vehicle for 6 days and were immunized with a uveitogenic regimen of IRBP in CFA. After 48 h, the draining LN were extracted and the cytokine expression was quantitated by real-time PCR using the 2^−ΔΔCT method. Data were combined from two independent experiments. B, Naive B10.RIII were gavaged with calcitriol or vehicle for 6 days, but were not immunized. Splenic CD11c⁺ DC were stimulated with M. tuberculosis extract in the presence or absence of calcitriol overnight. Cytokines in the supernatant were assayed by Multiplex ELISA. C, BMDC conditioned or not with calcitriol were stimulated with M. tuberculosis extract for 5 h. The supernatant at 1/10 dilution was added to naive CD4⁺ T cells stimulated with anti-CD3/anti-CD28 mAbs. IL-17 in the supernatants was assayed after 48 h. RORγt expression was quantitated after 12 h. The relative expression levels of RORγt are normalized to GAPDH. D, BMDC conditioned or not with calcitriol were pulsed with IRBP p161–180 and M. tuberculosis extract for 5 h. One million cells were injected into footpads of B10.RIII mice. After 5 days, draining LN were extracted and recalled with p161–180. RORγt was quantitated after 6 h, and IL-17 production was measured after 48 h. *, p < 0.05. Data represent one of two independent experiments with similar results.