title: Recombinant human osteogenic protein-1 (hOP-1) induces new bone formation in vivo with a specific activity comparable with natural bovine osteogenic protein and stimulates osteoblast proliferation and differentiation in vitro.
abstract: We reported previously that a 32-36-kDa osteogenic protein purified from bovine bone matrix is composed of dimers of two members of the transforming growth factor (TGF)-beta superfamily: the bovine equivalent of human osteogenic protein-1 (OP-1) and bone morphogenetic protein-2a, BMP-2a (BMP-2). In the present study, we produced the recombinant human OP-1 (hOP-1) in mammalian cells as a processed mature disulfide-linked homodimer with an apparent molecular weight of 36,000. Examination of hOP-1 in the rat subcutaneous bone induction model demonstrated that hOP-1 was capable of inducing new bone formation with a specific activity comparable with that exhibited by highly purified bovine osteogenic protein preparations. The half-maximal bone-inducing activity of hOP-1 in combination with a rat collagen matrix preparation was 50-100 ng/25 mg of matrix as determined by the calcium content of day 12 implants. Evaluation of hOP-1 effects on cell growth and collagen synthesis in rat osteoblast-enriched bone cell cultures showed that both cell proliferation and collagen synthesis were stimulated in a dose-dependent manner and increased 3-fold in response to 40 ng of hOP-1/ml. Examination of the expression of markers characteristic of the osteoblast phenotype showed that hOP-1 specifically stimulated the induction of alkaline phosphatase (4-fold increase at 40 ng of hOP-1/ml), parathyroid hormone-mediated intracellular cAMP production (4-fold increase at 40 ng of hOP-1/ml), and osteocalcin synthesis (5-fold increase at 25 ng of hOP-1/ml). In long-term (11-17 day) cultures of osteoblasts in the presence of beta-glycerophosphate and L(+)-ascorbate, hOP-1 markedly increased the rate of mineralization as measured by the number of mineral nodules per well (20-fold increase at 20 ng of hOP-1/ml). Direct comparison of TGF-beta 1 and hOP-1 in these bone cell cultures indicated that, although both hOP-1 and TGF-beta 1 promoted cell proliferation and collagen synthesis, only hOP-1 was effective in specifically stimulating markers of the osteoblast phenotype.
authors.affiliation: Creative BioMolecules Inc., Hopkinton, Massachusetts 01748.
authors.forename: T K
authors.initials: TK
authors.lastname: Sampath
authors.forename: J C
authors.initials: JC
authors.lastname: Maliakal
authors.forename: P V
authors.initials: PV
authors.lastname: Hauschka
authors.forename: W K
authors.initials: WK
authors.lastname: Jones
authors.forename: H
authors.initials: H
authors.lastname: Sasak
authors.forename: R F
authors.initials: RF
authors.lastname: Tucker
authors.forename: K H
authors.initials: KH
authors.lastname: White
authors.forename: J E
authors.initials: JE
authors.lastname: Coughlin
authors.forename: M M
authors.initials: MM
authors.lastname: Tucker
authors.forename: R H
authors.initials: RH
authors.lastname: Pang
mesh_terms: D000469:Alkaline Phosphatase / Q000096:biosynthesis; D000595:Amino Acid Sequence; D000818:Animals; D015153:Blotting, Western; D055419:Bone Morphogenetic Protein 7; D019485:Bone Morphogenetic Proteins; D016466:CHO Cells; D002417:Cattle; D006224:Cricetinae; D000242:Cyclic AMP / Q000096:biosynthesis; D004591:Electrophoresis, Polyacrylamide Gel; D004790:Enzyme Induction; D006801:Humans; D008969:Molecular Sequence Data; D010006:Osteoblasts / Q000166:cytology / Q000187:drug effects*; D015675:Osteocalcin / Q000096:biosynthesis; D010012:Osteogenesis / Q000187:drug effects*; D010281:Parathyroid Hormone / Q000502:physiology; D011506:Proteins / Q000494:pharmacology*; D051381:Rats; D011994:Recombinant Proteins / Q000494:pharmacology; D016212:Transforming Growth Factor beta / Q000235:genetics
chemical_list: C525819:BMP7 protein, human; C525821:Bmp7 protein, rat; D055419:Bone Morphogenetic Protein 7; D019485:Bone Morphogenetic Proteins; D010281:Parathyroid Hormone; D011506:Proteins; D011994:Recombinant Proteins; D016212:Transforming Growth Factor beta; D015675:Osteocalcin; D000242:Cyclic AMP; D000469:Alkaline Phosphatase