Immunocytochemical and immunochemical study of enamelins, using antibodies against porcine 89-kDa enamelin and its N-terminal synthetic peptide, in porcine tooth germs.
Enamelins comprise an important family of the enamel matrix proteins. Porcine tooth germs were investigated immunochemically and immunocytochemically using two antibodies: a polyclonal antibody raised against the porcine 89-kDa enamelin (89 E) and an affinity purified anti-peptide antibody against the porcine enamelin amino-terminus (EN). Immunochemical analysis of layers of immature enamel from the matrix formation stage detected immunopositive protein bands ranging from 10 kDa to 155 kDa in the outer layer enamel sample irrespective of the antibodies used. In contrast, the middle and inner enamel layer mainly contained lower molecular weight enamelins. In immunocytochemical analyses of the differentiation stage, 89 E stained enamel matrix islands around mineralized collagen fibrils of dentin, while EN stained both enamel matrix islands and stippled material. At the matrix formation stage, both antibodies intensely stained enamel prisms located in the outer layer. In the inner layer, 89 E moderately stained enamel matrix homogeneously, while EN primarily stained the prism sheath. The intense immunoreaction over the surface layer of enamel matrix at the matrix formation stage, following staining with 89 E and EN, disappeared by the end of the transition stage and the early maturation stage, respectively. The Golgi apparatus and secretory granules in the ameloblasts from the late differentiation stage to the transition stage were immunostained by both antibodies. These results suggest that expression of enamelin continues from late differentiation to the transition stage and the cleavage of N-terminal region of enamelin occurs soon after secretion. Some enamelin degradation products, which apparently have no affinity for hydroxyapatite crystals, concentrate in the prism sheaths during enamel maturation.
Dohi
N
N
Department of Endodontology and Periodontology, Hiroshima University School of Dentistry, Kasumi 1-2-3, Minami-ku, Hiroshima 734-8553, Japan.
Murakami
C
C
Tanabe
T
T
Yamakoshi
Y
Y
Fukae
M
M
Yamamoto
Y
Y
Wakida
K
K
Shimizu
M
M
Simmer
J P
JP
Kurihara
H
H
Uchida
T
T
D000595:Amino Acid Sequence; D000818:Animals; D000906:Antibodies / Q000276:immunology; D003746:Dental Enamel Proteins / Q000276:immunology / Q000378:metabolism*; D007150:Immunohistochemistry; D008854:Microscopy, Electron; D008969:Molecular Sequence Data; D010455:Peptides / Q000276:immunology; D013552:Swine; D014083:Tooth Germ / Q000378:metabolism* / Q000648:ultrastructure
D000906:Antibodies; D003746:Dental Enamel Proteins; D010455:Peptides; C038456:tuftelin