Identification of 25-hydroxyvitamin D3 1α-hydroxylase gene expression in macrophages

Background The 25-hydroxyvitamin D-3 1 alpha-hydroxylase (1 alpha-hydroxylase) is almost exclusively expressed in the kidney. However, 1 alpha-hydroxylase activities have been observed in some extrarenal tissues, including inflammatory cells of the monocyte/macrophage lineage. In sarcoidosis, macrophage 1 alpha-hydroxylase causes overproduction of 1,25-(OH)(2)D-3, resulting in hypercalcemia. In this study, we investigated the regulation of macrophage 1 alpha-hydroxylase at a molecular level. Methods. We used the human monocytic cell line THP-1, which can be differentiated into macrophage-like cells by treatment with phorbol ester. The expression of 1 alpha-hydroxylase in THP-1 cells was examined by Northern blotting and immunoblotting using an antibody raised against a synthetic peptide corresponding to the 14 C-terminal amino acids of 1 alpha-hydroxylase. We investigated the regulation of 1 alpha-hydroxylase mRNA expression by RNase protection assay. Results. Northern blot and immunoblot analyses confirmed the expression of 1 alpha-hydroxylase in THP-1 cells at the mRNA and protein levels. Although parathyroid hormone and calcitonin, known stimulators of renal 1 alpha-hydroxylase, did not affect the expression of 1 alpha-hydroxylase mRNA, 8-Br-cAMP (5 x 10(-4) mol/L) increased the expression of 1 alpha-hydroxylase mRNA in THP-1 cells (198 +/- 9%). 1,25-(OH)(2)D-3, known as a suppressor of renal 1 alpha-hydroxylase, did not affect the expression of la-hydroxylase mRNA. By contrast, 1,25-(OH)(2)D-3 markedly increased the expression of 25-hydroxyvitamin D-3 24-hydroxylase mRNA. Interferon-gamma (2000 IU/mL) increased the expression of 1 alpha-hydroxylase mRNA in differentiated THP-1 cells (922 +/- 25%). Conclusions. The present results suggest that 1 alpha-hydroxylase activity in macrophages is mediated by the same enzyme as in kidney. Interferon-gamma treatment increases macrophage 1 alpha-hydroxylase levels via directly increasing gene expression of this enzyme.