ascorbic-acid and alpha-glycerophosphoric-acid

Previous studies showed that B7-H3 (CD276), a cytokine involved in the activation of T lymphocytes, regulates murine bone formation. However, the role of B7-H3 in bone formation is barely understood. Herewith, we report, that stimulation of B7-H3 promotes the differentiation of human marrow stromal cells (hMSCs) to osteoblasts. With 4H7, a new monoclonal antibody against human B7-H3, we have identified B7-H3 is located on the surface of human marrow stromal cells. Evermore, we have found that increase of B7-H3 levels are correlated with the differentiation course of hMSCs. Stimulation of B7-H3 with 4H7 antibody considerably increases the numbers of osteoblasts generated from the hMSCs in the presence of inducing medium containing dexamethasone, sodium β-glycerophosphate and l-ascorbic acid. 4H7 treatments significantly increase osteoblast markers including alkaline phosphatase (ALP), and osteocalcin (OC) after day 7 and day 14 of the inducing hMSCs differentiation. The numbers of mineralized nodules of osteoblasts have been remarkly increased after 21 days of induced differentiation of hMSCs. However, stimulation effects of 4H7 antibody on membrane B7-H3 has been eliminated by addition of B7-H3Fc fusion protein. These results indicate 4H7 antibody specifically stimulates the membrane B7-H3 and directs the differentiation of hMSCs. Furthermore, our study also shows that stimulation of B7-H3 increases the expression of osteoprotein (OPG), and decreases the expression of its cognate ligand, the receptor activator of nuclear factor kappaB ligand (RANKL).

Topics: Antibodies, Monoclonal; Ascorbic Acid; B7 Antigens; Biomarkers; Bone Marrow Cells; Cell Differentiation; Cells, Cultured; Dexamethasone; Glycerophosphates; Humans; Osteoblasts; Osteocalcin; Osteogenesis; Stromal Cells; Up-Regulation

Effects of ascorbic acid and 5,6-O-isopropylidene-2,3-O-dimethylascorbic acid on final product formation in radiolysis of ethanol, aqueous solutions of ethanol, ethylene glycol, alpha-methylglycoside, maltose, alpha-glycerophosphate, and alpha-glucose phosphate were studied. It was found that ascorbic acid is able to suppress reactions involving various alpha-hydroxyl-containing carbon-centered radicals and depending on the experimental conditions can either oxidize or reduce alpha-hydroxyethyl radicals.

Topics: Antioxidants; Ascorbic Acid; Carbon; Drug Design; Ethanol; Ethylene Glycol; Free Radicals; Glucosephosphates; Glycerophosphates; Humans; Hydrogen-Ion Concentration; Hydroxyl Radical; Maltose; Methylglycosides; Models, Chemical

Cells released by sequential enzymatic digestion of 18-day chick calvariae were cultured over a 4-5 week period in Alpha modified Eagles medium. In some cultures the medium was supplemented with ascorbate and/or Na-beta-glycerophosphate. Microscopic examination of these cultures showed both polygonal and spindle-shaped cells. The biochemical nature of these cells was investigated by incubating the cultures with radiolabelled proline and subsequently analysing the medium and cell layer proteins by SDS/PAGE and fluorography. Osteoblast and chondrocyte-containing cultures were clearly distinguished in this way as the former cells secreted type I collagen while the latter secreted types II and X collagens as the major medium macromolecules. Type X collagen synthesis occurred after 14 days, but only in cultures supplemented with both ascorbate and Na-beta-glycerophosphate, and was maintained for the duration of the culture period. Unsupplemented cultures and those containing either ascorbate alone or Na-beta-glycerophosphate alone failed to synthesize type X collagen after 28 days. Isolated cells pulsed with radiolabelled proline at confluence and organ cultures of embryonic chick calvaria synthesized types I and V collagens only. These data demonstrate that the expression of phenotype by heterogeneous populations of bone cells could be modulated by a combination of culture conditions including the length of time in culture and conditions favourable for the formation of a mineralized matrix.

Topics: Animals; Ascorbic Acid; Cartilage; Cells, Cultured; Chick Embryo; Collagen; Culture Techniques; Glycerophosphates; Hydroxyproline; Molecular Weight; Osteogenesis; Peptide Mapping; Phenotype; Proline; Protein Biosynthesis; Proteins