Establishment of an osteoid preosteocyte-like cell MLO-A5 that spontaneously mineralizes in culture
Abstract
The mechanisms controlling the initiation of mineralization of bone matrix are not clear. To examine this process, we established a cell line called MLO-A5 that mineralizes in sheets, not nodules, within 3 days of culture in the presence of beta-glycerophosphate (beta-GP) and ascorbic acid and within 7 days in the absence of beta-GP and ascorbic acid. The mineral formed in both cases was shown to be bonelike apatite by Fourier transformed infrared (FTIR) spectroscopy. Mineral-to-matrix ratios (min/matrix) calculated from the FTIR data, which are related directly to ash weight, were approximately 0.4 in the absence of beta-GP and ascorbic acid and approximately 1.2 in the presence of beta-GP and ascorbic acid. By comparison, these ratios in fetal rat calvarial cells without beta-GP equal 0 and with beta-GP 1.9. This cell line and three others (MLO-A2, -D1, and -D6) were isolated from the long bones of transgenic mice expressing the large T-antigen driven by the osteocalcin promoter, the same mice from which the osteocyte-like cell line MLO-Y4 was isolated.(1) The cell lines were selected based on a dendritic or stellate morphology. MLO-A5 cells express high alkaline phosphatase, collagen type 1, parathyroid hormone/parathyroid hormone-related peptide (PTH/PTHrP) receptor, bone sialoprotein (BSP), and osteocalcin (767 ng/10(6) cells compared with <1-2.2 ng/10(6) cell for primary mouse osteoblasts and five osteoblast cell lines). The single unique feature of the MLO-A5 cells…
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