Figure 4.

Impact of vitamin D on the peripheral immune response and expression of MHC class II and CIITA in the CNS of DA.PVGav1-Vra4 congenic rats. (A) qPCR performed on mRNA isolated from local draining lymph nodes and (B) Flow cytometry performed on d7 LN single cell suspension of the congenic rats subjected to either: (i) vitamin D supplemented, (ii) deprived, or (iii) regular diet (data based on 2 pooled independent experiments with similar outcome). (A) Graphs presenting mRNA levels of indicated targets (n = 8-12 per diet group). Vitamin D supplementation downregulates Th1 type response in the congenic strain. Relative expression was calculated in relation to the mean of housekeeping gene Rpl19 using 2-ΔΔCT method. (B) Graphs presenting total cell counts of CD4+ and CD8+ T cells as well as CD45RA+ B cells (n = 12 per diet group). Amount of CD8+ cells is significantly downregulated upon vitamin D supplementation. (A,B) 2 independent experiments with similar outcome pooled. (C) Immuno-co-targeting of MHC class II (green) and CIITA (red) in the same CNS tissue samples (n = 4-6 per diet group) as in the Figure 3. Vitamin D supplementation tends to downregulate both MHC class II and CIITA in the congenic strain as shown in the quantification in (D). Error bars represent SEM. Statistics were calculated using one-way ANOVA with Tukey correction for multiple testing (*p < 0.05), (**p < 0.001). Statistical significances refer to comparison between vitamin D deprived and supplemented DA.PVGav1-Vra4.