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Volume 270, Number 16, Issue of April 21, pp. 9420-9428, 1995
©1995 by The American Society for Biochemistry and Molecular Biology, Inc.

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Rapidly Forming Apatitic Mineral in an Osteoblastic Cell Line (UMR 10601 BSP)

Clark M. Stanford , Paul A. Jacobson , E. David Eanes , Lois A. Lembke , Ronald J. Midura

This study evaluated a rapid biomineralization phenomenon exhibited by an osteoblastic cell line, UMR 106-01 BSP, when treated with either organic phosphates [-glycerophosphate (-GP), Ser-P, or Thr-P], inorganic phosphate (P), or calcium. In a dose-dependent manner, these agents (2-10 m M) stimulated confluent cultures to deposit mineral in the cell layer (EDof 4.6 m M for -GP (30 ± 2 nmol Ca/µg DNA) and 3.8 m M (29 ± 2 nmol Ca/µg DNA) for P) with a plateau in mineral formation by 20 h (ET 12-15 h). -GP or Ptreatment yielded mineral crystals having an x-ray diffraction pattern similar to normal human bone. Alizarin red-S histology demonstrated calcium mineral deposition in the extracellular matrix and what appeared to be intracellular paranuclear staining. Electron microscopy revealed small, needle-like crystals associated with fibrillar, extracellular matrix deposits and intracellular spherical structures. Mineral formation was inhibited by levamisole (ED 250 µ M), pyrophosphate (ED 1-10 µ M), actinomycin C(500 ng/ml), cycloheximide (50 µg/ml), or brefeldin A (1 µg/ml). These results indicate that UMR 106-01 BSP cells form a bio-apatitic mineralized matrix upon addition of supplemental phosphate. This process involves alkaline phosphatase activity, on-going RNA and protein synthesis, as well as Golgi-mediated processing and secretion.

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