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OBJECTIVE To determine the serum level of free 1,25-dihydroxyvitamin D [1,25-(OH)2D] in patients with vitamin D toxicity and to assess the in vitro effect of differing concentrations of vitamin D metabolites on the free serum levels of 1,25-(OH)2D. METHODS 1) A case study of patients hospitalized with vitamin D toxicity after accidentally ingesting a veterinary vitamin D concentrate and 2) an in vitro experiment in which vitamin D metabolites in various concentrations were added to normal serum and their effect was noted on percentage of free 1,25-(OH)2D. METHODS 11 patients (age range, 8 to 69 years) were studied 10 to 40 days after hospitalization for hypercalcemia. METHODS Serum total 25-hydroxyvitamin D (25-OHD) and 1,25-(OH)2D levels were measured by radioreceptor assays. The percentage of free 1,25-(OH)2D was measured by centrifugal ultrafiltration isodialysis and was used to calculate actual free 1,25-(OH)2D levels. In the in vitro studies, vitamin D metabolites [25-OHD; 24,25-(OH)2D; 25,26-(OH)2D; and 25-OHD-26,23 lactone] were added to normal serum in concentrations expected to occur with vitamin D toxicity. The percentage of free 1,25-(OH)2D was measured by isodialysis. RESULTS All patients presented with marked hypercalcemia (mean calcium level, 3.99 +/- 0.33 mmol/L). Serum 25-OHD levels ranged from 847 to 1652 nmol/L, and total 1,25-(OH)2D levels (mean, 106 +/- 86 pmol/L) were elevated in only three patients. The percentage of free 1,25-(OH)2D (mean, 1.023% +/- 0.366%) was elevated in all nine patients in whom it was measured. Actual free 1,25-(OH)2D levels (mean, 856 +/- 600 fmol/L) were elevated in six of the nine patients. Total 1,25-(OH)2D levels were correlated with 25-OHD levels (r = 0.66; P = 0.03), whereas total and free 1,25-(OH)2D levels were highly correlated (r = 0.957; P < 0.001). In the in vitro studies, the percentage of free 1,25-(OH)2D increased after 25-OHD or 24,25-(OH)2D was added. CONCLUSIONS Although the patients had normal or near-normal total 1,25-(OH)2D values, most patients had elevated free 1,25-(OH)2D levels. These findings suggest that elevated free 1,25-(OH)2D levels might play a role in the pathogenesis of hypercalcemia in vitamin D toxicity.
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