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Immune Regulation of 25-Hydroxyvitamin D-1-Hydroxylase in Human Monocytic THP1 Cells: Mechanisms of Interferon--Mediated Induction

Laboratory for Experimental Medicine and Endocrinology (L.O., K.S., A.V., R.B., C.M.) and Laboratory for Experimental Transplantation (M.W.), University Hospital Gasthuisberg, Catholic University of Leuven, B-3000 Leuven, Belgium

Address all correspondence and requests for reprints to: Chantal Mathieu, M.D., Ph.D., LEGENDO, UZ-Gasthuisberg, Onderwijs en Navorsing, Herestraat 49, B-3000 Leuven, Belgium. E-mail: chantal.mathieu{at}med.kuleuven.be.

Context: 25-Hydroxyvitamin D can be activated to 1,25-dihydroxyvitamin D₃ [1,25(OH)₂D₃] by the rate-limiting enzyme 1-hydroxylase in cells of the immune system under control of immune stimuli, such as interferon- (IFN). In pathological situations, such as sarcoidosis, this can lead to systemic excess of 1,25(OH)₂D₃ and hypercalcemia.

Objective: The aim of this study was to elucidate the intracellular pathways used by the immune system to tightly regulate 1,25(OH)₂D₃ production in monocytes and macrophages.

Design: Human monocytic THP1-cells were differentiated and activated by IFN and a secondary stimulus, such as lipopolysaccharide or phorbol myristate acetate. 1-Hydroxylase mRNA levels were quantified by real-time RT-PCR. The involvement of different signaling pathways in the regulation of this enzyme was investigated using specific pharmacological inhibitors, whereas phosphorylation of signal transducer and activator of transcription 1 and CCAAT/enhancer binding protein ß was investigated by Western blotting.

Results: In undifferentiated monocytic THP1 cells, IFN needs to be combined with a second stimulus, such as lipopolysaccharide, to induce 1-hydroxylase. In contrast, in phorbol myristate acetate-differentiated THP1 macrophages, IFN alone induces 1-hydroxylase and to much higher levels. Many different signaling pathways need to be activated concurrently to allow immune-mediated 1-hydroxylase up-regulation. We show involvement of the Janus kinase-signal transducer and activator of transcription, MAPK, and nuclear factor-B pathways, with a crucial role for the transcription factor CCAAT/enhancer binding protein ß. Furthermore, histone remodeling involving histone deacetylases and histone acetylase p300 is required.

Conclusion: The present findings indicate that IFN-mediated 1,25(OH)₂D₃ production, as observed in granulomatous diseases such as sarcoidosis, will take place only under conditions where the necessary other signaling pathways are also activated.