title: Identification of the chondrogenic-inducing activity from bovine dentin (bCIA) as a low-molecular-mass amelogenin polypeptide.
abstract: Dentin extracellular matrix has been shown to contain components capable of inducing chondrogenesis and osteogenesis at ectopic sites when implanted in vivo, and chondrogenesis in cultures of embryonic muscle-derived fibroblasts (EMF) in vitro. The polypeptide responsible, called the chondrogenic-inducing agent (CIA), has been isolated from a 4.0-M guanidinium hydrochloride extract of demineralized bovine dentin matrix. Following Sephacryl S-100 chromatography, CIA activity was identified in fractions by assay for uptake of [35S]-SO4 into proteoglycan by the EMF after 24 hrs in culture. The active fraction induced the EMF to produce type II collagen mRNA and decrease production of type I collagen mRNA after 5 days in culture. The EMF + CIA, cultured for 4 to 7 wks, formed toluidine-blue- and alizarin-red-stainable nodules, indicative of chondrogenic induction. In vivo implants in rat muscle with collagen carrier produced ectopic bone after 7 wks. The CIA was brought to near-homogeneity by reverse-phase high-performance liquid chromatography, tested at each step by EMF [35S]-SO4-incorporation assays. The CIA components had masses in the ranges of 6000 to 10,000 Da by both mass spectroscopy and gel electrophoresis. The CIA amino acid composition, NH2-terminal, and internal amino acid sequences were determined. These data showed unequivocally that the CIA peptides were derived from bovine amelogenin. The peptides contain the amino-terminal portion of the bovine amelogenin. The presence of these chondrogenic/osteogenic amelogenin-polypeptides in dentin matrix leads us to hypothesize that they may be involved in epithelial-mesenchymal signaling during tooth development interactions-the first time a function has been indicated for these molecules.
authors.affiliation: Department of Basic and Behavioral Sciences, Northwestern University Dental School, Chicago, Illinois 60611, USA.
authors.forename: D R
authors.initials: DR
authors.lastname: Nebgen
authors.forename: H
authors.initials: H
authors.lastname: Inoue
authors.forename: B
authors.initials: B
authors.lastname: Sabsay
authors.forename: K
authors.initials: K
authors.lastname: Wei
authors.forename: C S
authors.initials: CS
authors.lastname: Ho
authors.forename: A
authors.initials: A
authors.lastname: Veis
mesh_terms: D053523:Amelogenin; D000595:Amino Acid Sequence; D000818:Animals; D001681:Biological Assay; D002417:Cattle; D002478:Cells, Cultured; D005591:Chemical Fractionation / Q000379:methods; D020219:Chondrogenesis; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D004274:DNA, Recombinant / Q000096:biosynthesis; D003746:Dental Enamel Proteins / Q000737:chemistry* / Q000235:genetics; D003804:Dentin / Q000737:chemistry*; D016326:Extracellular Matrix Proteins / Q000302:isolation & purification; D005347:Fibroblasts / Q000378:metabolism; D018507:Gene Expression Regulation, Developmental; D006133:Growth Substances / Q000737:chemistry* / Q000302:isolation & purification; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010455:Peptides / Q000737:chemistry / Q000302:isolation & purification; D011506:Proteins / Q000737:chemistry* / Q000302:isolation & purification; D051381:Rats; D017207:Rats, Sprague-Dawley; D009418:S100 Proteins / Q000032:analysis
chemical_list: D053523:Amelogenin; C507201:Amelx protein, rat; D004274:DNA, Recombinant; D003746:Dental Enamel Proteins; D016326:Extracellular Matrix Proteins; D006133:Growth Substances; D010455:Peptides; D011506:Proteins; D009418:S100 Proteins