Novel insight into the role of the vitamin D receptor in the development and function of the immune system.

ABSTRACT

Immune cells express the vitamin D receptor (VDR) and are therefore vitamin D targets. The Vdr protein can be readily measured in the kidney using antibodies to the Vdr and western blot. It is much more difficult to measure Vdr protein in the spleen because of the low level of VDR expression in resting immune cells. In order to more sensitively measure VDR expression, the Cre enzyme was inserted in the 3rd exon of the VDR gene and a reporter mouse that irreversibly expresses tdTomato was made. Mice that express one copy of the VDRCre gene were confirmed to be VDR +/- and mice that express two copies were confirmed to be VDR -/-. Initial characterization of the immune cells from the VDR +/-/VDRtdTomato+ mice, compared to VDR+/+ wildtype (WT) littermates, showed no effect of being hemizygous for the VDR on immune cell frequencies. High tdTomato expression was shown to be present in the bone marrow (BM) and thymus immune cell precursors. In the periphery, monocytes, neutrophils and macrophages had very high tdTomato+ (88-98%) expression while lymphocytes ranged from 60% to 70% tdTomato+. Tissue resident innate lymphoid cell (ILC) 1 and 3 cells were about 60-80% tdTomoto+, while ILC2 cells had very low tdTomato expression. Stimulation of VDRtdTomato+ splenocytes showed that the tdTomato- CD4+ and CD8+ T cells proliferated more than their tdTomato+ counterparts. T cells were sorted for tdTomato+ and tdTomato- and then activated for 72 h. Sorted tdTomato+ T cells expressed the VDR protein only after 72 h post-activation. The sorted tdTomato- T cells proliferated more than the sorted tdTomato+ T cells. Interestingly, activation of the tdTomato- T cells failed to induce new tdTomato expression. The data suggest that an early immune precursor expresses the VDR. In the periphery, neutrophils and monocytes are almost all tdTomato+, while some immune cells (ILC2 and some T cells) may never express the VDR.