gms | German Medical Science

The three main enzymes of vitamin D metabolism, vitamin D3-25-hydroxylase (25-OHase), 25-hydroxyvitamin D3-1α-hydroxylase (1α-OHase) and 25-hydroxyvitamin D3-24-hydroxylase (24-OHase) have been described in malignant breast tissue. This in vitro study aimed to obtain more information the regulation of these enzymes in the human breast cancer cell line MCF-7.Cells were cultured in cell culture flasks, then sowed in Petri dishes and stimulated with the vitamin D metabolites vitamin D3 (calciferol), 25-calciferol and calcitriol for 24, 48 and 72 hours in physiological (10-9 M) and supraphysiological (10-7 M) concentrations. Before stimulation the abundance of the vitamin D- receptor was proven by conventional PCR. After stimulation, RNA was extracted and the expression of the enzymes was assessed by real time PCR. Results were normalized to the “house-keeping gene” HPRT as an internal standard for each sample. Expression of 25-OHase could not be influenced by substrates. The expression 1α-OHase could be slightly induced after stimulation with 10-7 M calcitriol after 72 h. However, after stimulation with supraphysiological concentrations of calcitriol the expression of 24-OHase was increased 144fold after 24 h, 32fold after 48 h and still 10fold after 72 h. These results could not be seen at physiological calcitriol doses. Our results demonstrate that MCF-7 cells not only possess 24-OHase, but also are able to regulate the expression of this calcitriol inactivating enzyme. This might be a mechanism for these tumor cells to protect themselves against the antiproliferative and apoptosis- inducing effects of calcitriol.