Skin is an autonomous organ in synthesis, two-step activation and degradation of vitamin D-3: CYP27 in epidermis completes the set of essential vitamin D-3-hydroxylases

The current understanding of the vitamin D-3 system shows skin as the uniqu e site of vitamin D-3 production and liver is thought to be the main site o f conversion to 25(OH)D-3. Skin is capable of activating 25(OH)D-3 via 1 al pha -hydroxylation and the resulting 1 alpha ,25(OH)(2)D-3 plays a role in epidermal homeostasis in normal and diseased skin. It also rapidly up-regul ates the major vitamin D-3 metabolizing enzyme 24-hydroxylase at the mRNA l evel, which is an established indicator for 1 alpha ,25(OH)(2)D-3-presence. We investigated the capability of primary human keratinocytes to produce 2 5(OH)D-3 and subsequent metabolites from vitamin D-3. Thus, by orchestratin g the entire system of production, activation and inactivation, skin could be independent of other organs in supply of hormonally active vitamin D-3. First, we demonstrated substantial conversion of H-3-D-3 to H-3-25(OH)D-3 i n primary human keratinocytes. 25-Hydroxylation was slow, followed first or der rate kinetics and was not saturable under our experimental conditions. Then we showed expression of 25-hydroxylase mRNA and compared it to levels of 1 alpha -hydroxylase and 24-hydroxylase. Pre-incubation with vitamin D-3 resulted in dose and time dependent up-regulation of 24-hydroxylase mRNA, whereas neither 1 alpha -hydroxylase nor 25-hydroxylase expression was affe cted. Since both, D-3 and 25(OH)D-3 are lacking intrinsic 24-hydroxylase-in ducing capacity, up-regulation had to be the consequence of a two-step acti vation process via 25-hydroxylation and subsequent 1 alpha -hydroxylation. 24-Hydroxylase-activities closely followed the corresponding mRNA levels. W hen 1 alpha ,25(OH)(2)D-3 itself or its precursor 25(OH)D-3 were used as in ducing agents, 24-hydroxylase mRNA and enzyme activity followed a transient time course. In contrast, induction observed with physiological doses of D -3 remained high, even after a 20 h-time period. These differing characteri stics may be explained by the slow but constant formation of 1 alpha ,25(OH )(2)D-3 from a large reservoir of D-3 in the target cell, providing constan t supplies for induction. (C) 2001 Elsevier Science Inc. All rights reserve d.