We found that na ve CD4 T cells express no or very low selleck bio levels of CYP27B1. Inhibitors,Modulators,Libraries However, the cells started to express CYP27B1 mRNA shortly after stimulation, and the expression peaked after 2 3 days of stimulation. These results suggested that acti vated human CD4 T cells have the capacity to convert 25 D3 to 1,25 Inhibitors,Modulators,Libraries 2D3. To validate this, we stimulated purified CD4 T cells in the presence of 100 nM 25 D3 corresponding to physiological concentrations of 25 D3 in serum and then measured the production of 1,25 2D3. We found that activated CD4 T cells pro duced 1,25 2D3 with a kinetic similar to the kinetics of CYP27B1 expression in the cells, and that the produc tion peaked after 3 days of stimulation. Finally, to determine whether the cells expressed the receptor for 1,25 2D3, we determined the expression of the VDR in CD4 T cells stimulated for 0 5 days.

We found that VDR expression peaked simultaneously with the peak pro duction of 1,25 2D3 at day 3. Taken to gether, these experiments demonstrated that activated human CD4 T cells express CYP27B1, that they have the capacity to convert 25 D3 at physiological concentra tions to the active 1,25 2D3, and that they express the receptor for 1,25 2D3. Activated T cells have the ability to Inhibitors,Modulators,Libraries produce 1,25 2D3 in sufficiently high concentrations to affect vitamin D responsive genes Having demonstrated that activated CD4 T cells have the capacity to produce the active form of vitamin D in a dose dependent manner. In addition, we found that 25 D3 also induced CD38 up regulation, suggesting that activated T cells can pro duce sufficiently high concentrations of 1,25 2D3 to affect vitamin D responsive genes.

We noted that whereas 25 D3 at a physiological concentration of 100 nM induced maximum CD38 up regulation, 1,25 2D3 was required at a 100 fold higher concentra tion than physiological levels to induce the same effect. That a supra physiological con centration of 1,25 2D3 was required to obtain an ef fect in our study is in good line with previous studies. However, in Inhibitors,Modulators,Libraries contrast to our observations that 25 D3 at 100 nM had a clear effect on T cells, most previous studies on 25 D3 required at least a 100 fold higher concentration of 25 D3 than found in serum to induce an effect. One explanation for this discrepancy could be that whereas our experiments were performed in serum free medium most previous studies have been performed in medium supplemented with serum.

To study whether the presence of serum could explain the discrepancy between the concentra tions of 25 D3 required to obtain an effect on T cells, we activated CD4 T cells in the presence of in creasing concentrations of 25 D3 Inhibitors,Modulators,Libraries and 1,25 2D3 in medium supplemented with 5% fetal bovine serum selleck products and measured CD38 expression at day 3. We found that the presence of serum significantly shifted the con centration of 25 D3 required to induce CD38 expres sion.