Steroidogenesis in the human skin: 21-hydroxylation in cultured keratinocytes

We have evaluated the metabolism of radiolabeled progesterone (P) by the microsomal fraction isolated from HaCaT keratinocytes. P was widely metabolized to different compounds that included DOC (5–7% conversion) thus demonstrated 21-hydroxylase (21-OHase) activity, a key step in adrenal synthesis of gluco- and mineralocorticoids. However, RT-PCR amplification for the CYPc21 transcript of the corresponding gene showed no evidence for gene expression in HaCaT cells suggesting that the 21-OHase enzyme present in keratinocytes is different from that described in adrenal gland. Further characterization showed that whereas estradiol stimulated markedly P metabolism by HaCaT microsomes, with generation of new unidentified compounds, Lineweaver-Burk analysis of keratinocyte 21-OHase activity showed that the K m and V max were unaffected by estrogen. The apparent K m was 0.6 μM without estradiol and 0.7 μM with estradiol, while the respective V max values were 60 and 76 nmol/l/min. To conclude, we found extensive metabolism of P in human keratinocytes, we also provide the first demonstration of 21-OHase activity in this cell system and further showed that it is coded by a gene different from the adrenal CYPc21.