Kinetics of in vitro mineralization by an osteogenic clonal cell line (C1) derived from mouse teratocarcinoma

Isolation and Characterization of Human Clonogenic Osteoblast Progenitors Immunoselected from Fetal Bone Marrow Stroma Using STRO-1 Monoclonal Antibody

“…This suggests that some form of homotypic cell–cell interaction is required to induce the cascade of events in F/STRO‐1 + A cells which eventually results in the expression of the mature osteoblast phenotype. A similar requirement for cell–cell contacts and aggregates to induce commitment to the osteogenic lineage has been reported for clonal mesodermal cells (40,60) . However, these data do not indicate whether F/STRO‐1 + A cells are osteogenic lineage‐restricted progenitors.…”

“…Although F/STRO‐1 + A cells form multilayers in very confluent cultures, they do not form mineralized nodules as seen in cultures of rat BM stromal cells. Accordingly, we cultured the cells as 3D aggregates in conditions that induce osteogenesis in immortalized clonal mesenchymal cells (40) . Under these conditions, an extracellular matrix was progressively deposited and mineralized.…”

“…Media were changed every 2 days, and [ 3 H]thymidine (1 μCi/ml) was added to the media in some dishes 24 h before the aggregates were harvested. Calcium levels in the matrix formed in aggregates was determined at several time points as described (40) . The amount of OC and carboxy‐terminal propeptide of type I procollagen (PICP) secreted were measured in conditioned media harvested at various time points.…”

Multi-Channelled Collagen–Calcium Phosphate Scaffolds: Their Physical Properties and Human Cell Response

“…Alkaline phosphatase and collagen I levels were high, a common finding after the addition of ascorbic acid. 34,35 A significant increase in metabolic activity values at 14 days was observed on all collagencalcium phosphate scaffolds but not on the glass surface controls (Fig. 7).…”

Up‐regulation of bone morphogenetic protein receptor IB by growth factors enhances BMP‐2‐induced human bone cell functions

“…Exponentially‐growing cells were obtained by culture with 10% FCS‐DMEM in a 96‐well plate for 48 h. After incubating with 1% FCS‐DMEM for another 24 h, TGF‐β1 (0.1 ng/ml), FGF‐2 (0.1 ng/ml), and PDGF‐AB (1 ng/ml) in 1% FCS‐DMEM, and in some experiments BMP‐2 (300 ng/ml), was added for 24–72 h. For the determination of ALP activity, p ‐nitrophenyl phosphate (Sigma) was used as a substrate. Since ALP is a tightly membrane‐bound enzyme, the cell monolayer was directly overlaid with the substrate solution as described previously (Gerstenfeld et al, 1987; Chentoufi et al, 1993). Briefly, ALP activity of the cell layer was measured in triplicate wells by rinsing twice with PBS and then incubating the cells with 200 µl of 5 mM p ‐nitrophenyl phosphate in 50 mM glycine, 1 mM MgCl 2 and 150 mM 2‐amino‐2‐methlyl‐1‐propanol buffer, pH 10.5 at 37°C for 1 h. A 50 µl aliquot of 3 M NaOH was added to stop the enzymatic reaction and the absorption at 405 nm was measured spectrophotometrically using a plate reader (Titertek Multiskan® Plus MKII; Labsystems, Helsinki, Finland).…”