Endocrine Abstracts

Recently, evidence has emerged that active 1,25dihydroxyvitamin D₃ (1,25D₃) is involved in modulating the immune response. Additionally, vitamin D₃ insufficiency has been proposed as a predisposing factor for autoimmune diseases, which correlate with circulating inactive 25-hydroxyvitamin D₃ (25D₃). Substrate 25D₃ is converted to 1,25D₃ by the activating enzyme 1alpha-hydroxylase. We and others have previously shown that during DC differentiation from circulating monocyte precursors, DCs upregulate 1alpha-hydroxylase and have an increased capacity to produce 1,25D₃. Conversely, we previously demonstrated that increased capacity to produce 1,25D₃ coincides with downregulation of VDR expression. This implies 1,25D₃ produced locally by mature DCs may signal in a paracrine fashion to regulate or inhibit development of circulating monocyte precursors which express abundant VDR. Here we extend these observations and demonstrate that macrophages and DCs show similar 1alpha-hydroxylase kinetics. To assess the functional significance of circulating 25D₃ levels we cultured DCs and macrophages from monocyte precursors under conditions that mimic vitamin D₃ deficiency, insufficiency and sufficiency. Cells were cultured with 5% human serum alone or with 50 or 150 nM 25D₃ added during culture in the presence or absence of ketoconazole (1alpha-hydroxylase inhibitor). Analysis of supernatants by radioimmunoassay showed dose-dependent accumulation of active 1,25D₃ in DCs and macrophages which was inhibited by ketoconazole. In parallel, increasing doses of 1,25D₃ and 25D₃ resulted in lower expression of the antigen presenting MHC class II and co-stimulatory molecules. Conversely, 25D₃ and 1,25D₃ enhanced expression of the monocyte marker CD14. Responses to 25D₃ but not 1,25D₃ were blocked by ketoconazole. Together, data suggest local synthesis of 1,25D₃ may have immunomodulatory effects on DC and macrophage phenotype, furthermore the ability of macrophages and DCs to synthesise 1,25D₃ is highly dependent on substrate 25D₃ levels, which maybe a pivotal factor in mediating the impact of vitamin D₃ deficiency and insufficiency on autoimmune disease.