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Experimental allergic encephalomyelitis (EAE) is a Th1 cell‐mediated autoimmune disease model of multiple sclerosis (MS). Vitamin D deficiency is commonly observed in MS patients and vitamin D supplements reduce the clinical symptoms of EAE and MS. Earlier studies have shown that in vivo treatment with vitamin D analogs ameliorates EAE in association with the inhibition of IL‐12 production and Th1 differentiation. The mechanisms in the regulation of Th1 response by vitamin D in EAE/MS are, however, not known. We show that in vivo treatment of C57BL/6 and SJL/J mice (i.p.) with 100 ng of 1,25 dihydroxyvitamin D3, on every other day from Day 0–30, ameliorates EAE in association with the inhibition of IL‐12 production and neural antigen‐specific Th1 response. In vitro treatment with 1,25(OH)2D3 inhibited IFNγ‐induced tyrosine phosphorylation of STAT1, without affecting JAK2, in EOC‐20 microglial …