Regulation of the procine 1,25-dihydroxyvitamin D3-24-hydroxylase (CYP24) by 1,25-dihydroxyvitamin D3 and parathyroid hormone in AOK-B50 cells

The 24-hydroxylase is the enzyme responsible for the first step in the catabolism of 1,25-dihydroxyvitamin D3, the active form of vitamin D. This enzyme was shown to be upregulated by 1,25-dihydroxyvitamin D3 itself and downregulated by parathyroid hormone (PTH). Upregulation of 24-hydroxylase by 1,25-dihydroxyvitamin D3 has been characterized; however, the mechanism by which PTH acts to downregulate 24-hydroxylase expression remains unknown. Here we report the cloning of the porcine 24-hydroxylase, and show that 1,25-dihydroxyvitamin D3-stimulated 24-hydroxylase mRNA and activity are repressed by PTH in AOK-B50 cells, a porcine kidney proximal tubule cell line with stably transfected opossum PTH receptors. Forskolin mimicked the effects of PTH consistent with in vivo data, and suppression by PTH was not due to changes in VDR levels. The first 1400 bp of the 24-hydroxylase promoter were not able to mediate the effects of PTH on a reporter gene. In view of the above findings we concluded that AOK-B50 cells are a suitable model for further studying the mechanism of action of PTH on 24-hydroxylase mRNA.     

Characterization of the vitamin D endocrine system in human sebocytes in vitro

Sebocytes are sebum-producing cells that form the sebaceous glands. We investigated the role of sebocytes as target cells for vitamin D metabolites and the existence of an enzymatic machinery for the local synthesis and metabolism of 1,25-dihydroxyvitamin D₃ [1,25(OH)₂D₃, calcitriol], the biologically active vitamin D metabolite, in these cell types. Expression of vitamin D receptor (VDR), vitamin D-25-hydroxylase (25OHase), 25-hydroxyvitamin D-1α-hydroxylase (1αOHase), and 1,25-dihydroxyvitamin D-24-hydroxylase (24OHase) was detected in SZ95 sebocytes in vitro using real time quantitative polymerase chain reaction. Splice variants of 1αOHase were identified by nested touchdown polymerase chain reaction. We demonstrated that incubation of SZ95 sebocytes with 1,25(OH)₂D₃ resulted in a cell culture condition-, time-, and dose-dependent modulation of cell proliferation, cell cycle regulation, lipid content and interleukin-6/interleukin-8 secretion in vitro. RNA expression of VDR and 24OHase was upregulated along with vitamin D analogue treatment. Although several other splice variants of 1αOHase were detected, our findings indicate that the full length product represents the major 1αOHase gene product in SZ95 cells. In conclusion, SZ95 sebocytes express VDR and the enzymatic machinery to synthesize and metabolize biologically active vitamin D analogues. Sebocytes represent target cells for biologically active metabolites. Our findings indicate that the vitamin D endocrine system is of high importance for sebocyte function and physiology. We conclude that sebaceous glands represent potential targets for therapy with vitamin D analogues or for pharmacological modulation of 1,25(OH)₂D₃ synthesis/metabolism.