The temporal expression patterns and activity distributions of enamelysin and EMSP1, which are the major proteinases in immature enamel, were characterized. Extracellular matrix fractions from developing porcine incisors, individually comprised of predentin, dentin, and four secretory-stage enamel samples, including the highly mineralized enamel (HME) at the enamel-dentin junction (EDJ), were isolated, and their resident proteinases were identified by zymography. Soft-tissue fractions, which included cells from the extension site of enamel formation (ESEF), secretory- and maturation-stage ameloblasts, and odontoblasts, were characterized histologically and by RT-PCR for their expression of enamelysin and EMSP1. A significant finding was that EMSP1, expressed by odontoblasts, concentrates in the HME, but is not detected in predentin or dentin. We conclude that odontoblasts deposit EMSP1 via their cell processes into the deepest enamel layer, which facilitates the hardening of this layer and contributes significantly to the functional properties of the EDJ.
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References
1.
Bègue-Kirn C, Krebsbach PH, Bartlett JD, Butler WT -1998-. Dentin sialoprotein, dentin phosphoprotein, enamelysin and ameloblastin: tooth-specific molecules that are distinctively expressed during murine dental differentiations. Eur J Oral Sci106:963–970.
2.
Burgess RC, Maclaren CM -1965-. Proteins in developing bovine enamel. In: Tooth enamel I. Stack MV, Fearnhead RW, editors. Bristol: John Wright & Sons Ltd.,
pp. 74-82.
3.
Cerny R, Slaby I, Hammarström L, Wurtz T -1996-. A novel gene expressed in rat ameloblasts codes for proteins with cell binding domains. J Bone Miner Res11:883–891.
4.
Chomczynski P, Sacchi N -1987-. Single-step method of RNA isolation by acid guanidinium thiocyanate-phenol-chloroform extraction. Anal Biochem162:156–159.
5.
Fukae M, Shimizu M -1974-. Studies on the proteins of developing bovine enamel. Arch Oral Biol19:381–386.
6.
Fukae M, Tanabe T -1998-. Degradation of enamel matrix proteins in porcine secretory enamel. Connect Tissue Res39:123–129.
7.
Fukae M, Tanabe T, Yamada M -1994-. Action of metalloproteinases on porcine dentin mineralization. Calcif Tissue Int55:426–435.
8.
Fukae M, Tanabe T, Uchida T, Lee SK, Ryu OH, Murakami C, et al. -1998-. Enamelysin -matrix metalloproteinase-20-: localization in the developing tooth and effects of pH and calcium on amelogenin hydrolysis. J Dent Res77:1580–1588.
9.
Fukae M, Tanabe T, Yamakoshi Y, Yamada M, Ujiie Y, Oida S -2001-. Immunoblot detection and expression of enamel proteins at the apical portion of the forming root in porcine permanent incisor tooth germs. J Bone Miner Metab19:236–243.
10.
Hu CC, Fukae M, Uchida T, Qian Q, Zhang CH, Ryu OH, et al. -1997a-. Sheathlin: cloning, cDNA/polypeptide sequences, and immunolocalization of porcine enamel sheath proteins. J Dent Res76:648–657.
11.
Hu CC, Fukae M, Uchida T, Qian Q, Zhang CH, Ryu OH, et al. -1997b-. Cloning and characterization of porcine enamelin mRNAs. J Dent Res76:1720–1729.
12.
Hu CC, Ryu OH, Chen JJ, Uchida T, Wakida K, Murakami C, et al. -2000-. Cloning of the murine EMSP1 cDNA and analysis of expression during tooth development. J Dent Res79:70–76.
13.
Krebsbach PH, Lee SK, Matsuki Y, Kozak CA, Yamada K, Yamada Y -1996-. Full-length sequence, localization, and chromosomal mapping of ameloblastin: a novel tooth-specific gene. J Biol Chem271:4431–4435.
14.
McMeekin T, Marshall K -1952-. Specific volumes of proteins and the relationship to their amino acid contents. Science116:142–143.
15.
Miake Y, Shimoda S, Fukae M, Aoba T -1993-. Epitaxial overgrowth of apatite crystals on the thin-ribbon precursor at early stages of porcine enamel mineralization. Calcif Tissue Int53:249–256.
16.
Munksgaard EC, Rhodes M, Mayne R, Butler WT -1978-. Collagen synthesis and secretion by rat incisor odontoblasts in organ culture. Eur J Biochem82:609–617.
17.
Oida S, Nagano T, Yamakoshi Y, Ando H, Yamada M, Fukae M -2002-. Amelogenin gene expression in porcine odontoblasts. J Dent Res81:103–108.
18.
Robinson C, Lowe NR, Weatherell JA -1975-. Amino acid composition, distribution and origin of "tuft" protein in human and bovine dental enamel. Arch Oral Biol20:29–42.
19.
Robinson C, Kirkham J, Weatherell JA, Richards A, Josephsen K, Fejerskov O -1988-. Mineral and protein concentrations in enamel of the developing permanent porcine dentition. Caries Res22:321–326.
20.
Schiess AV, Okigaki T, Rounds DE -1966-. Human odontoblasts in vitro. I. The harvest of homogeneous cell populations. J Dent Res45:1101–1104.
21.
Simmer JP, Fukae M, Tanabe T, Yamakoshi Y, Uchida T, Xhu J, et al. -1998-. Purification, characterization, and cloning of enamel matrix serine proteinase 1. J Dent Res77:377–386.
22.
Suga S -1983-. Comparative histology of the progressive mineralization pattern of developing enamel. In: Mechanisms of tooth enamel formation. Suga S, editor. Tokyo:
Quintessence Publishing Co. Inc., pp. 167-203.
23.
Tanabe T -1984-. Purification and characterization of proteolytic enzymes in porcine immature enamel. Tsurumi Shigaku10:443–452.
24.
Tanabe T, Fukae M, Uchida T, Shimizu M -1992-. The localization and characterization of proteinases for the initial cleavage of porcine amelogenin. Calcif Tissue Int51:213–217.
25.
Tanabe T, Fukae M, Shimizu M -1996-. Possible actions of metalloproteinases found in porcine enamel in an early secretory stage. Adv Dent Res10:170–172.
26.
Uchida T, Tanabe T, Fukae M -1989-. Immunocytochemical localization of amelogenins in the deciduous tooth germs of the human fetus. Arch Histol Cytol52:543–552.
27.
Uchida T, Tanabe T, Fukae M, Shimizu M -1991-. Immunocytochemical and immunochemical detection of a 32 kDa nonamelogenin and related proteins in porcine tooth germs. Arch Histol Cytol54:527–538.
28.
Uchida T, Fukae M, Tanabe T, Yamakoshi Y, Satoda T, Murakami C, et al. -1995-. Immunochemical and immunocytochemical study of a 15 kDa non-amelogenin and related proteins in the porcine immature enamel: proposal of a new group of enamel proteins sheath proteins. Biomed Res16:131–140.
