Ultrastructure of granular materials in rat incisor enamel organ at early maturation stage

Exposure to chronic high levels of fluoride results in the formation of fluorosed enamel. Although enamel may be more susceptible to fluorotic effects at certain stages of development, fluoride at sufficiently high levels may affect enamel at all stages of formation. Careful study of the changes in enamel caused by chronic fluoride ingestion is needed to understand more fully the mechanisms involved in the formation of fluorotic enamel. This paper discusses the various studies we have completed to define the changes, in developing enamel of the rat incisor, caused by long-term ingestion of fluoride in drinking water. Fluoride has been found to inhibit secretion of enamel proteins. Changes in the maturation stage of enamel formation include the retention of amelogenin proteins during early maturation. The various mechanisms which have been investigated in the formation of fluorosed enamel include a direct effect of fluoride on the enamel organ, and specific interactions of fluoride with the extracellular enamel matrix. Although the same amount of protease appears to be secreted in fluorosed and control enamel, a delay in the digestion of amelogenin protein occurs. This suggests that fluoride may directly or indirectly inhibit the protease present in fluorosed enamel to slow the proteolysis of amelogenins.

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Gene Expression and Localization of Amelogenin in the Rat Incisor

Advances in Dental Research ◽

10.1177/08959374960100021401 ◽

1996 ◽

Vol 10 (2) ◽

pp. 201-207 ◽

Cited By ~ 22

Author(s):

T. Inage ◽

H. Shimokawa ◽

K. Wakao ◽

S. Sasaki

Keyword(s):

Gene Expression ◽

Amino Acids ◽

Maturation Stage ◽

Enamel Matrix ◽

Rat Incisor ◽

Early Maturation ◽

Incisal Edge ◽

Enamel Epithelium ◽

Developing Rat

Gene expression and localization of amelogenin were studied in the developing rat incisor by the methods of in situ hybridization and immunohistochemistry. ISH revealed the first expression of amelogenin mRNA in the inner enamel epithelium of the cervical loop. The signals were clearly observed in pre-ameloblasts in the region bordering on predentin formation and became more intense toward the cells on the initial enamel matrix secretion. The maximal signals were found in the cytoplasm of secretory ameloblasts. From the terminal secretion zone, the signals then became gradually weaker toward the incisal edge but were still evident in the cytoplasm of shortening, transitional ameloblasts and those at the early maturation stage. No signals were found in the cells of the stratum intermedium and stellate reticulum throughout amelogenesis. Immunohistochemistry by means of an antibody against amelogenin C-telopeptide consisting of 12 amino acids revealed immunoreaction in the secretory ameloblasts reacting to the ISH. When a polyclonal antibody against amelogenin was used, immunoreaction was found in the distal ends of ruffle-ended ameloblasts (RA) in the maturation zone. Those results indicated that amelogenin is synthesized by ameloblastic cells from the inner enamel epithelium to the early maturation stage and is then resorbed by the RA.

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Cytochemical and biochemical characterization of glycoproteins in forming and maturing enamel of the rat incisor.

Journal of Histochemistry & Cytochemistry ◽

10.1177/37.11.2809173 ◽

1989 ◽

Vol 37 (11) ◽

pp. 1619-1633 ◽

Cited By ~ 33

Author(s):

A Nanci ◽

J P Ahluwalia ◽

S Zalzal ◽

C E Smith

Keyword(s):

Secretory Granules ◽

Helix Pomatia ◽

Extracellular Proteins ◽

Maturation Stage ◽

Variable Degree ◽

Rat Incisor ◽

Double Labeling ◽

Early Maturation ◽

Enamel Proteins ◽

Lowicryl K4m

Biochemical and histochemical studies have shown the presence of various carbohydrates in enamel. Using lectin-gold cytochemistry, we have examined the distribution of glycoconjugates containing N-acetyl-D-galactosamine (GalNAc) and/or N-acetyl-glucosamine (GlcNAc)/N-acetyl-neuraminic acid (NeuNAc) residues in rat incisor ameloblasts and in forming and maturing enamel embedded in Lowicryl K4M, LR Gold, and LR White resins. The enamel proteins that contain these carbohydrate moieties were further characterized by lectin blotting. All three resins allowed, albeit to a variable degree, detection of the binding sites for Helix pomatia agglutinin (HPA) and wheat germ agglutinin (WGA) GalNAc, and GlcNAc/NeuNAc, respectively. In general, Lowicryl K4M permitted more intense reactions with both lectins. Lectin binding was observed over the rough endoplasmic reticulum (weak labeling with WGA), the Golgi apparatus, lysosomes, secretory granules, and the enamel matrix. These compartments were shown by double labeling with WGA and anti-amelogenin antibody, and by previous immunocytochemical studies, to contain enamel proteins. Furthermore, WGA binding was more concentrated at the growth sites of enamel. Lectin blotting showed that several proteins in the amelogenin group were glycosylated and contained the sugars GalNAc and GlcNAc/NeuNAc. Fewer proteins were stained by HPA than by WGA, and the staining pattern suggested that the extracellular proteins recognized by these two lectins are processed differently. The HPA-reactive proteins were lost by or during the early maturation stage, whereas many of the WGA-reactive proteins persisted into the mid maturation stage. The heterogeneous staining of certain protein bands observed with WGA suggests that they contain more than one component. Two distinct glycoproteins containing GlcNAc/NeuNAc also appeared during the maturation stage. These results are consistent with the notion that ameloblasts produce an extracellular matrix composed mainly of glycosylated amelogenins which are differently processed throughout amelogenesis.

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The Influence of Vanadate on Calcium Uptake in Maturing Enamel of the Rat Incisor

Journal of Dental Research ◽

10.1177/00220345870660120101 ◽

1987 ◽

Vol 66 (12) ◽

pp. 1702-1707 ◽

Cited By ~ 12

Author(s):

Y. Takano ◽

S. Matsuo ◽

S. Wakisaka ◽

H. Ichikawa ◽

S. Nishikawa ◽

...

Keyword(s):

Enamel Organ ◽

Moderate Intensity ◽

Maturation Stage ◽

Vascular Perfusion ◽

Rat Incisor ◽

Experimental Conditions ◽

Efficient Diffusion ◽

Ca Uptake

The influence of vanadate, a potent inhibitor of Ca2+-ATPase and Na+-K+-ATPase, on 45Ca uptake in maturing enamel of the rat incisor was investigated by a vascular perfusion method combined with 45Ca autoradiography. The morphological integrity of the maturation-stage enamel organ was well-retained during vascular perfusion under all the experimental conditions. Distinct patterns of 45Ca labeling, comparable with those found in previous in vivo 45Ca autoradiographic studies, appeared in the maturing enamel after vascular perfusion with a standard perfusate. One mmol/L vanadate added to the standard perfusate caused a drastic decrease in 45Ca uptake in the maturing enamel, corresponding to the ruffle-ended ameloblasts, leaving narrow peaks of moderate intensity corresponding to the bands of the overlying smooth-ended ameloblasts. The in vitro labeling of exposed enamel surfaces with 45Ca revealed blackening of autoradiographic emulsion in wide bands separated by unlabeled or slightly labeled narrow ones resembling the distribution of smooth-ended ameloblasts in both control and vanadate-treated incisors. Our observations indicate that the ruffle-ended ameloblasts of the rat incisor serve as an efficient diffusion barrier to calcium ions and regulate transcellular calcium transport to the maturing enamel, at least in part, by a vanadate-sensitive mechanism.

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Proteinases in Developing Dental Enamel

Critical Reviews in Oral Biology & Medicine ◽

10.1177/10454411990100040101 ◽

1999 ◽

Vol 10 (4) ◽

pp. 425-441 ◽

Cited By ~ 204

Author(s):

J.D. Bartlett ◽

J.P. Simmer

Keyword(s):

Proteinase Inhibitors ◽

Dental Enamel ◽

Serine Proteinase ◽

Organic Matrix ◽

Enamel Organ ◽

Maturation Stage ◽

Enamel Matrix ◽

Enamel Matrix Proteins ◽

Early Maturation

For almost three decades, proteinases have been known to reside within developing dental enamel. However, identification and characterization of these proteinases have been slow and difficult, because they are present in very small quantities and they are difficult to purify directly from the mineralizing enamel. Enamel matrix proteins such as amelogenin, ameloblastin, and enamelin are cleaved by proteinases soon after they are secreted, and their cleavage products accumulate in the deeper, more mature enamel layers, while the full-length proteins are observed only at the surface. These results suggest that proteinases are necessary for "activating" enamel proteins so the parent proteins and their cleavage products may perform different functions. A novel matrix metalloproteinase named enamelysin (MMP-20) was recently cloned from tooth tissues and was later shown to localize primarily within the most recently formed enamel. Furthermore, recombinant porcine enamelysin was demonstrated to cleave recombinant porcine amelogenin at virtually all of the sites that have previously been described in vivo. Therefore, enamelysin is at least one enzyme that may be important during early enamel development. As enamel development progresses to the later stages, a profound decrease in the enamel protein content is observed. Proteinases have traditionally been assumed to degrade the organic matrix prior to its removal from the enamel. Recently, a novel serine proteinase named enamel matrix serine proteinase-1 (EMSP1) was cloned from enamel organ epithelia. EMSP1 localizes primarily to the early maturation stage enamel and may, therefore, be involved in the degradation of proteins prior to their removal from the maturing enamel. Other, as yet unidentified, proteinases and proteinase inhibitors are almost certainly present within the forming enamel and await discovery.

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Role of Innate Inflammation in the Regulation of Tissue Remodeling during Tooth Eruption

Dentistry Journal ◽

10.3390/dj9010007 ◽

2021 ◽

Vol 9 (1) ◽

pp. 7

Author(s):

Yusuke Makino ◽

Kaoru Fujikawa ◽

Miwako Matsuki-Fukushima ◽

Satoshi Inoue ◽

Masanori Nakamura

Keyword(s):

Bacterial Infections ◽

Tooth Movement ◽

Tooth Eruption ◽

Intercellular Adhesion Molecule ◽

Enamel Organ ◽

Pcr Analysis ◽

Maturation Stage ◽

Oral Epithelium ◽

Rt Pcr ◽

Inflammatory Reactions

Tooth eruption is characterized by a coordinated complex cascade of cellular and molecular events that promote tooth movement through the eruptive pathway. During tooth eruption, the stratum intermedium structurally changes to the papillary layer with tooth organ development. We previously reported intercellular adhesion molecule-1 (ICAM-1) expression on the papillary layer, which is the origin of the ICAM-1-positive junctional epithelium. ICAM-1 expression is induced by proinflammatory cytokines, including interleukin-1 and tumor necrosis factor. Inflammatory reactions induce tissue degradation. Therefore, this study aimed to examine whether inflammatory reactions are involved in tooth eruption. Reverse transcription-polymerase chain reaction (RT-PCR) analysis revealed sequential expression of hypoxia-induced factor-1α, interleukin-1β, and chemotactic factors, including keratinocyte-derived chemokine (KC) and macrophage inflammatory protein-2 (MIP-2), during tooth eruption. Consistent with the RT-PCR results, immunohistochemical analysis revealed KC and MIP-2 expression in the papillary layer cells of the enamel organ from the ameloblast maturation stage. Moreover, there was massive macrophage and neutrophil infiltration in the connective tissue between the tooth organ and oral epithelium during tooth eruption. These findings suggest that inflammatory reactions might be involved in the degradation of tissue overlying the tooth organ. Further, these reactions might be induced by hypoxia in the tissue overlying the tooth organ, which results from decreased capillaries in the tissue. Our findings indicate that bacterial infections are not associated with the eruption process. Therefore, tooth eruption might be regulated by innate inflammatory mechanisms.

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Relative Levels of mRNA Encoding Enamel Proteins in Enamel Organ Epithelia and Odontoblasts

Journal of Dental Research ◽

10.1177/154405910308201209 ◽

2003 ◽

Vol 82 (12) ◽

pp. 982-986 ◽

Cited By ~ 59

Author(s):

T. Nagano ◽

S. Oida ◽

H. Ando ◽

K. Gomi ◽

T. Arai ◽

...

Keyword(s):

Tooth Enamel ◽

Structural Proteins ◽

Enamel Organ ◽

Maturation Stage ◽

Mrna Levels ◽

Rt Pcr ◽

Enamel Protein ◽

Enamel Proteins ◽

Enamel Layer

Amelogenin, enamelin, sheathlin (ameloblastin/ amelin), enamelysin (MMP-20), and KLK4 (EMSP-1) are the major structural proteins and proteinases in developing tooth enamel. Recently, odontoblasts were reported to express amelogenin, the most abundant enamel protein. In this study, we hypothesized that odontoblasts express all enamel proteins and proteases, and we measured their relative mRNA levels in enamel organ epithelia and odontoblasts associated with porcine secretory- and maturation-stage enamel by RT-PCR, using a LightCycler instrument. The results showed that amelogenin mRNA in secretory-stage EOE is 320-fold higher than in odontoblasts beneath secretory-stage enamel, and over 20,000-fold higher than in odontoblasts under maturation-stage enamel. Similar results were obtained for enamelin and sheathlin. Enamelysin mRNA levels were equivalent in these two tissues, while KLK4 mRNA was higher in odontoblasts than in secretory-stage EOE. These results support the conclusion that odontoblasts are involved in the formation of the enamel layer adjacent to enamel-dentin junction.

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Penetration of various molecular-weight proteins into the enamel organ and enamel of the rat incisor

Archives of Oral Biology ◽

10.1016/0003-9969(86)90042-7 ◽

1986 ◽

Vol 31 (5) ◽

pp. 287-296 ◽

Cited By ~ 39

Author(s):

M.D. McKee ◽

B. Martineau-Doize ◽

H. Warshawsky

Keyword(s):

Molecular Weight ◽

Enamel Organ ◽

Rat Incisor

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Cytochemical localization of calcium and Ca2+,Mg2(+)-adenosine triphosphatase in colchicine-altered rat incisor ameloblasts.

Journal of Histochemistry & Cytochemistry ◽

10.1177/38.10.2144864 ◽

1990 ◽

Vol 38 (10) ◽

pp. 1469-1478 ◽

Cited By ~ 9

Author(s):

D R Eisenmann ◽

A H Salama ◽

A M Zaki ◽

S H Ashrafi

Keyword(s):

Electron Microscopy ◽

Transmission Electron Microscopy ◽

Adenosine Triphosphatase ◽

Morphological Changes ◽

Rat Incisor ◽

Cytochemical Localization ◽

Early Maturation ◽

Transmission Electron ◽

Maturation Stages ◽

Secretory Transport

Colchicine is known to affect secretory, transport, and degradative functions of ameloblasts. The effects of colchicine on membrane-associated calcium and Ca2+,Mg2(+)-ATPase in secretory and maturation ameloblasts were investigated cytochemically. The pyroantimonate (PPA) method was used for localizing calcium and a modified Wachstein-Meisel medium was used to localize Ca2+,Mg2(+)-ATPase. Sections representing secretory and early maturation stages were examined by transmission electron microscopy. Morphological changes induced by colchicine included dislocated organelles and other well-established reactions to such anti-microtubule drugs. Calcium pyroantimonate (Ca-PA) deposits in most ameloblast types were markedly reduced, with the greater reduction occurring in those cells more severely altered morphologically. However, the cell membranes of both control and experimental smooth-ended maturation ameloblasts were essentially devoid of Ca-PA. The normal distribution and intensity of Ca2+,Mg2(+)-ATPase was not affected by colchicine. Because the observed reduction of membrane-associated calcium is apparently not mediated by Ca2+,Mg2(+)-ATPase in this case, other aspects of the calcium regulating system of ameloblasts are apparently targeted by colchicine.

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