1Alpha,25-(OH)2D3 acts in the early phase of osteoblast differentiation to enhance mineralization via accelerated production of mature matrix vesicles.

1Alpha,25-dihydroxyitamin D(3) (1,25D3) deficiency leads to impaired bone mineralization. We used the human pre-osteoblastic cell line SV-HFO, which forms within 19 days of culture an extracellular matrix that starts to mineralize around day 12, to examine the mechanism by which 1,25D3 regulates osteoblasts and directly stimulates mineralization. Time phase studies showed that 1,25D3 treatment prior to the onset of mineralization, rather than during mineralization led to accelerated and enhanced mineralization. This is supported by the observation of unaltered stimulation by 1,25D3 even when osteoblasts were devitalized just prior to onset of mineralization and after 1,25D3 treatment. Gene Chip expression profiling identified the pre-mineralization and mineralization phase as two strongly distinctive transcriptional periods with only 0.6% overlap of genes regulated by 1,25D3. In neither phase 1,25D3 significantly altered expression of extracellular matrix genes. 1,25D3 significantly accelerated the production of mature matrix vesicles (MVs) in the pre-mineralization. Duration rather than timing determined the extent of the 1,25D3 effect. We propose the concept that besides indirect effects via intestinal calcium uptake 1,25D3 directly accelerates osteoblast-mediated mineralization via increased production of mature MVs in the period prior to mineralization. The accelerated deposition of mature MVs leads to an earlier onset and higher rate of mineralization. These effects are independent of changes in extracellular matrix protein composition. These data on 1,25D3, mineralization, and MV biology add new insights into the role of 1,25D3 in bone metabolism and emphasize the importance of MVs in bone and maintaining bone health and strength by optimal mineralization status.

Osteoclastogenic capacity of peripheral blood mononuclear cells is not different between women with and without osteoporosis.

INTRODUCTION: Peripheral Blood Mononuclear Cells (PBMCs) have been extensively used as a culture model to generate osteoclasts in vitro. The aim of this study was to assess the osteoclastogenic potential of PBMCs derived from post-menopausal women with longstanding osteoporosis and compare this with PBMCs from healthy controls. MATERIAL AND METHODS: We selected from the population-based Rotterdam Study 82 participants of which 43 were diagnosed with osteoporosis (T-score below -2.5 at the lumbar spine) and the presence of at least 1 fracture and 29 healthy controls (T-score above 1; no fracture). PBMCs were differentiated into osteoclasts, and both differentiation capacity and activity were measured. Total RNA was obtained to assess gene expression of osteoclast markers. Deoxypyridinoline (DPD) was measured in plasma as a marker for bone resorption, in vivo. RESULTS: Neither the number of osteoclasts nor cathepsin K (CTSK) and dendritic cell-specific transmembrane protein (TM7SF4) gene expression was significantly different between both groups. There was also no significant difference in resorption pit area and plasma DPD levels. Stratification by fracture type into a group with vertebral, non-vertebral and both vertebral and non-vertebral fractures showed no difference in osteoclast formation or osteoclastic bone resorption. However, plasma DPD, but not the RNA expression markers, was significantly lower in the group of subjects with vertebral fracture group and those with vertebral and non-vertebral fractures compared to the healthy controls. No differences in osteoclastogenesis, osteoclastic resorption and plasma DPD levels were detected also after exclusion of past or present users of bisphosphonates and glucocorticoids. Stratification into high and low DPD levels showed higher osteoclastogenesis and more osteoclastic bone resorption in the high DPD group compared to the low DPD levels within the group of osteoporotic subjects. CONCLUSION: This study showed no difference in PBMC osteoclastogenic capacity and activity between women with and without osteoporosis and at least one previous fracture, who were on average 29.5years after menopause, suggesting that there is no difference in circulating osteoclast precursors. Although we cannot exclude that circulating precursors may behave differently at the bone site, it is possible that long after menopause a more stable phase of bone turnover is reached compared to earlier after the start of menopause in which differences in circulating osteoclast precursors and osteoclastogenic potential are more prominent.