Measurements of the Vitamin D Metabolome in the Calgary Vitamin D Study: Relationship of Vitamin D Metabolites to Bone Loss

In a 36-month randomized controlled trial examining the effect of high-dose vitamin D₃ on radial and tibial total bone mineral density (TtBMD), measured by high-resolution peripheral quantitative tomography (HR-pQCT), participants (311 healthy males and females aged 55-70 years with dual-energy X-ray absorptiometry T-scores > -2.5 without vitamin D deficiency) were randomized to receive 400 IU (N = 109), 4000 IU (N = 100), or 10,000 IU (N = 102) daily. Participants had HR-pQCT radius and tibia scans and blood sampling at baseline, 6, 12, 24, and 36 months. This secondary analysis examined the effect of vitamin D dose on plasma measurements of the vitamin D metabolome by liquid chromatography-tandem mass spectrometry (LC-MS/MS), exploring whether the observed decline in TtBMD was associated with changes in four key metabolites [25-(OH)D₃ ; 24,25-(OH)₂ D₃ ; 1,25-(OH)₂ D₃ ; and 1,24,25-(OH)₃ D₃ ]. The relationship between peak values in vitamin D metabolites and changes in TtBMD over 36 months was assessed using linear regression, controlling for sex. Increasing vitamin D dose was associated with a marked increase in 25-(OH)D₃ , 24,25-(OH)₂ D₃ and 1,24,25-(OH)₃ D₃ , but no dose-related change in plasma 1,25-(OH)₂ D₃ was observed. There was a significant negative slope for radius TtBMD and 1,24,25-(OH)₃ D₃ (-0.05, 95% confidence interval [CI] -0.08, -0.03, p < 0.001) after controlling for sex. A significant interaction between TtBMD and sex was seen for 25-(OH)D₃ (female: -0.01, 95% CI -0.12, -0.07; male: -0.04, 95% CI -0.06, -0.01, p = 0.001) and 24,25-(OH)₂ D₃ (female: -0.75, 95% CI -0.98, -0.52; male: -0.35, 95% CI -0.59, -0.11, p < 0.001). For the tibia there was a significant negative slope for 25-(OH)D₃ (-0.03, 95% CI -0.05, -0.01, p < 0.001), 24,25-(OH)₂ D₃ (-0.30, 95% CI -0.44, -0.16, p < 0.001), and 1,24,25-(OH)₃ D₃ (-0.03, 95% CI -0.05, -0.01, p = 0.01) after controlling for sex. These results suggest vitamin D metabolites other than 1,25-(OH)₂ D₃ may be responsible for the bone loss seen in the Calgary Vitamin D Study. Although plasma 1,25-(OH)₂ D₃ did not change with vitamin D dose, it is possible rapid catabolism to 1,24,25-(OH)₃ D₃ prevented the detection of a dose-related rise in plasma 1,25-(OH)₂ D₃ . © 2023 The Authors. Journal of Bone and Mineral Research published by Wiley Periodicals LLC on behalf of American Society for Bone and Mineral Research (ASBMR).