1,25-dihydroxyvitamin D resistance, rickets, and alopecia: analysis of receptors and bioresponse in cultured fibroblasts from patients and parents.

To investigate further the cellular defects of vitamin D-dependent rickets type II with alopecia, we studied 1,25-dihydroxyvitamin D3 [1,25-(OH)2D3] receptors and the response to 1,25-(OH)2D3 in cultured skin fibroblasts from rachitic patients. Our studies included cells from four affected patients from three kindreds and their parents and cells from five normal subjects. We measured total 1,25-(OH)2D3 receptor binding in cell extracts and the capacity of 1,25-(OH)2D3 to induce the enzyme 25-hydroxyvitamin D3-24-hydroxylase (24-hydroxylase) as a marker of functional response. In normal fibroblasts, the 1,25-(OH)2D3 maximal binding capacity was 52 +/- 5 fmol/100 micrograms DNA (mean +/- SE), and the apparent dissociation constant (Kd) was 0.05 +/- 0.01 nM. The maximal induced 24-hydroxylase activity after 1,25-(OH)2D3 treatment was 11.5 +/- 1 fmol/10(6) cells X 30 min, and the dose of 1,25-(OH)2D3 that achieved half-maximal induction was 2.3 +/- 0.3 nM. Fibroblasts from all four rachitic patients had the same defect: no measurable 1,25-(OH)2D3 receptor binding and no detectable response above basal activity even after high doses of 1,25-(OH)2D3. Cells from all parents except one had normal 1,25-(OH)2D3 binding characteristics and normal 24-hydroxylase bioresponse to 1,25-(OH)2D3. One parent despite a normal phenotype had only half the normal level of binding sites and only half the normal bioresponse. In summary, the cultured fibroblasts from four affected children representing three different kindreds with 1,25-(OH)2D3 resistance failed to exhibit detectable 1,25-(OH)2D3 receptors. We postulate that this biochemical defect produced both the inability to respond to 1,25-(OH)2D3 in vitro and the 1,25-(OH)2D3 resistance in vivo. The obligate heterozygotic parents were normal, except for one who had both half the normal number of receptors and half the normal response to 1,25-(OH)2D3. The data confirm the critical role of the receptor in 1,25-(OH)2D3 action and the close coupling of receptor content and functional responsiveness.