Abstract
BACKGROUND: Multipotential precursor cell lines derived from human bone marrow, capable of differentiating into cartilage or bone, may provide a useful tissue development model for studying the regulation and metabolism of putative growth and differentiation factors necessary for tissue regeneration. In mammals, the process of bone development depends on the proliferation and differentiation of osteoblast lineage cells, and the subsequent synthesis and mineralization of bone extracellular matrix (ECM). Vitamin D metabolites play a pivotal role in bone and mineral homeostasis, and are positive factors on bone development. Recently, it was demonstrated that a human-derived engineered osteoblast precursor cell line (OPC1), derived from human bone marrow, can metabolize the parental precursor vitamin D(3) (vitaD(3)) to the active steroid 1α,25-dihydroxyvitamin D(3) (1,25OH(2)D(3)), and elicit an osteogenic response that results in the decrease in proliferation and increase in ECM synthesis during early bone development. The aim in this study is to characterize gene expression, matrix production and mineralization within a bone development model. METHODS: We investigated whether vitaD(3) influences bone ECM mineralization in the same manner as 1,25OH(2)D(3) in confluent cultures of OPC1s. In addition, we explored the influence of vitamin D metabolites, in combination with other commonly used osteogenic factors, ascorbic acid, β-glycerophosphate, dexamethasone (dex) and recombinant human bone morphogenetic protein-2 (rhBMP-2) on the osteoinduction of OPC1. RESULTS: It was demonstrated that OPC1 expresses the mRNA for the enzymatic equipment necessary to convert vitaD(3) to 1,25OH(2)D(3), as well as the mRNA expression of the catabolic enzyme known to regulate the concentration of active 1,25OH(2)D(3). It was also demonstrated that mRNA expression for the vitamin D receptor (VDR) was influenced by both vitaD(3) and 1,25OH(2)D(3). Differential results using vitamin D metabolites in combination with ascorbic acid, β-glycerophosphate, dex and/or rhBMP-2 were observed in alkaline phosphatase (ALP) activity and calcium deposition, and mRNA expression of procollagen type I (proColI), osteocalcin (OC) and osteopontin (OP). CONCLUSIONS: Overall it was demonstrated that vitamin D in combination with osteogenic factors influences the temporal bone development sequence in a positive manner.