The aim of this study was to investigate the expression levels of vitamin D receptor (VDR) and NF-κB mRNAs in vitamin D (VD) supplemented multiple sclerosis (MS) patients. RRMS patients received 50,000 IU vitamin D3/week as an intra-muscular injection for 2 months. Blood samples were obtained from 30 MS patients before and after VD supplementation and 32 healthy individuals, and then VDR and NF-κB mRNA levels were measured by real time PCR method and analyzed with independent and paired t-tests. Moreover, some correlations were performed between the expression levels of selected genes and some clinical features of MS and control groups. Surprisingly, the expression level of VDR mRNA significantly decreased after 2 months supplementation with VD in our selected patients and in contrast, the level of serum 25(OH) D increased after supplementation. Although, we didn't find any significant difference in the expression level of NF-κB gene before and after treatment with VD, its expression significantly decreased in untreated MS cases compared with healthy controls. In conclusion, we found some new evidences from the molecular mechanism of vitamin D effectiveness in MS treatment. Also, we need more functional studies to find the effect of VD on the expression level of VDR mRNA.