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1.
J Periodontal Res ; 50(6): 846-54, 2015 Dec.
Article in English | MEDLINE | ID: mdl-25808290

ABSTRACT

BACKGROUND AND OBJECTIVE: After tooth formation is complete, it is suggested that continuity exists between the epithelial cell rests of Malassez (ERM), reduced enamel epithelium (REE) and subsequently the junctional epithelium. However, the junctional epithelium was reported to differ from REE and ERM. The developmental relationships between and among them remain controversial. Therefore, in the present study we examined the expression of cytokeratins in the three types of epithelia to investigate the epithelial phenotypes. MATERIAL AND METHODS: The maxillae of Wistar rats, 1, 2, 3 and 7 wk of age, were used, and the expression of CK14, CK17, CK19, CK10/CK13 and AE1/AE3 was detected using immunoperoxidase techniques. RESULTS: There was negative staining for CK10/CK13 in all the epithelia. ERM stained strongly for AE1/AE3, CK14, CK17 and CK19. During the transformation of inner enamel epithelial (IEE) cells into reduced ameloblasts and subsequently into junctional epithelium, strong staining for CK14 was evident in IEE, REE and junctional epithelium, whereas the expression of AE1/AE3 and of CK19 were initially negative in IEE and then strong in REE and junctional epithelium, respectively. In particular, the expression of CK17 was strongly positive in ERM and REE, but was negative in IEE and junctional epithelium. CONCLUSION: ERM are of odontogenic origin and junctional epithelium has an epithelial phenotype different from REE and ERM. This is the first report to demonstrate that CK17 can be used as a marker to distinguish junctional epithelium from ERM.


Subject(s)
Cell Differentiation , Enamel Organ/chemistry , Epithelial Attachment/chemistry , Epithelial Cells/chemistry , Keratins/analysis , Animals , Enamel Organ/physiology , Epithelial Attachment/physiology , Epithelial Cells/physiology , Immunohistochemistry , Rats, Wistar
2.
J Oral Sci ; 56(4): 261-8, 2014 Dec.
Article in English | MEDLINE | ID: mdl-25500923

ABSTRACT

Amelotin (AMTN) is a secreted protein transcribed predominantly during the maturation stage of enamel formation and localized in the junctional epithelium. We investigated differences in the levels of AMTN gene expression between non-inflamed gingiva and inflamed gingiva from patients with chronic periodontitis. Total RNAs were isolated from these tissues and their gene expression profiles were monitored by DNA microarray. The observed induction of AMTN mRNA in inflamed gingiva and cultured human gingival fibroblasts (HGF) was confirmed by real-time PCR. Transient transfection assays were performed using chimeric constructs of mouse AMTN gene promoter fragments linked to a luciferase reporter gene. Immunohistochemical localization of AMTN in inflamed and non-inflamed gingiva was assessed by immunohistochemistry. Among many differentially expressed genes, the level of AMTN mRNA was significantly increased in inflamed gingiva. Treatment of HGF with interleukin-1ß (IL-1ß), IL-6 and tumor necrosis factor-α (TNF-α) induced the expression of AMTN mRNA, and increased the luciferase activities of the AMTN promoter constructs. AMTN protein was detected in inflamed gingival connective tissue and junctional epithelium. These findings demonstrate that proinflammatory cytokines induce AMTN gene expression in human gingival fibroblasts and suggest a role for AMTN in gingival inflammation.


Subject(s)
Cytokines/pharmacology , Dental Enamel Proteins/drug effects , Fibroblasts/drug effects , Gingiva/drug effects , Inflammation Mediators/pharmacology , Transcription, Genetic/drug effects , Adult , Animals , Cells, Cultured , Chimera/genetics , Chronic Periodontitis/genetics , Chronic Periodontitis/pathology , Connective Tissue/chemistry , Dental Enamel Proteins/analysis , Dental Enamel Proteins/genetics , Epithelial Attachment/chemistry , Female , Fibroblasts/chemistry , Gene Expression Profiling , Genes, Reporter/genetics , Gingiva/chemistry , Gingiva/cytology , Gingivitis/genetics , Gingivitis/pathology , Humans , Interleukin-1beta/pharmacology , Luciferases/genetics , Male , Mice , Microarray Analysis , Middle Aged , Promoter Regions, Genetic/genetics , Real-Time Polymerase Chain Reaction , Tumor Necrosis Factor-alpha/pharmacology
3.
J Periodontol ; 85(8): e313-22, 2014 Aug.
Article in English | MEDLINE | ID: mdl-24527855

ABSTRACT

BACKGROUND: Many pathogens are known to modulate epithelial physical barriers, particularly tight-junction (TJ) proteins, to enter host cells and/or tissues. Growth factors have been implicated in the regulation of TJ proteins. The aim of this study is to determine differences in the levels of TJ proteins, growth factors, and their receptors in relation to bacterial invasion in diseased gingival tissues obtained from patients with periodontitis. METHODS: The presence of bacteria and expression of junctional adhesion molecule (JAM)-A, occludin, epidermal growth factor (EGF), keratinocyte growth factor (KGF), insulin-like growth factor-I (IGF-I), EGF receptor, KGF receptor, and IGF-1 receptor (IGF-1R) were evaluated in gingival tissues from healthy (n = 10) and diseased (n = 10) sites in patients with periodontitis by in situ hybridization and immunohistochemistry. RESULTS: The bacterial invasion of gingival tissue was increased in periodontal lesions compared with healthy sites. Although the levels of JAM-A and occludin were not significantly different between the healthy and diseased sites, aberrant cytoplasmic expression of JAM-A and occluding was often observed in the lesions. In addition, more leukocytes expressing JAM-A or occludin were observed within the disease-associated epithelia. Compared with the healthy sites, the differential expression of KGF, IGF-I, and IGF-1R was observed in the periodontal lesions. The levels of TJ proteins showed positive correlations with those of growth factors. CONCLUSION: The aberrant expression of growth factors and TJ proteins may contribute to increased bacterial invasion and disease progression in periodontal lesions.


Subject(s)
Bacteria/pathogenicity , Gingiva/chemistry , Intercellular Signaling Peptides and Proteins/analysis , Periodontitis/metabolism , Receptors, Growth Factor/analysis , Tight Junction Proteins/analysis , Adult , Bacterial Load , Cell Adhesion Molecules/analysis , Cytoplasm/chemistry , Disease Progression , Epidermal Growth Factor/analysis , Epithelial Attachment/chemistry , Epithelial Attachment/microbiology , ErbB Receptors/analysis , Female , Fibroblast Growth Factor 7/analysis , Gingiva/microbiology , Humans , Insulin-Like Growth Factor I/analysis , Leukocytes/chemistry , Male , Middle Aged , Occludin/analysis , Periodontitis/microbiology , Receptor, IGF Type 1/analysis , Receptors, Cell Surface/analysis
4.
Angle Orthod ; 84(3): 534-40, 2014 May.
Article in English | MEDLINE | ID: mdl-23984990

ABSTRACT

OBJECTIVE: To identify the regenerating junctional epithelium (JE) during orthodontic tooth movement in rats. MATERIALS AND METHODS: Closed-coil springs were used to create a 20 g mesial force to the maxillary first molars. On days 1, 3, 7, 10, and 14 after force application, histologic changes in JE were examined by immunohistochemistry using proliferating cell nuclear antigen (PCNA), odontogenic ameloblast-associated protein (ODAM), and cytokeratin 14 (CK14). RESULTS: On day 1, JE was destroyed and lost attachment to the tooth surface. Cell division activity was rarely observed in JE, and ODAM localization was weakly detected in damaged JE. By day 3, regenerating JE had not fully recovered. High cell proliferation activity and CK14 expression started to appear in most basal cells of JE. ODAM expression was reduced and appeared in a small area. By day 7, JE had almost recovered. Cell proliferation activity was still observed in several basal cells of JE, and ODAM expression was detected among JE cells. CK14 was hardly observed in JE except in the basal cells. By days 10 and 14, regenerated JE appeared. ODAM, PCNA, and CK14 expression was similar to that of the control. CONCLUSIONS: Damaged JE might recover rapidly during orthodontic tooth movement because basal cells of the remaining JE, which show higher proliferation activity, are involved in JE regeneration. Reduced ODAM expression during proliferation of JE cells may increase again after JE regeneration is complete. Therefore, ODAM may be associated with the normal function of JE.


Subject(s)
Calcium-Binding Proteins/analysis , Epithelial Attachment/chemistry , Keratin-14/analysis , Proliferating Cell Nuclear Antigen/analysis , Tooth Movement Techniques/methods , Animals , Basement Membrane/chemistry , Basement Membrane/pathology , Cell Division/physiology , Cell Proliferation , Epithelial Attachment/pathology , Epithelial Cells/chemistry , Epithelial Cells/pathology , Extracellular Space/chemistry , Gingiva/pathology , Male , Molar/pathology , Neutrophils/pathology , Orthodontic Wires , Rats , Rats, Sprague-Dawley , Regeneration/physiology , Stress, Mechanical , Time Factors , Tooth Movement Techniques/instrumentation
5.
Can J Physiol Pharmacol ; 86(3): 105-12, 2008 Mar.
Article in English | MEDLINE | ID: mdl-18418437

ABSTRACT

Epithelial cells form a tight barrier against environmental stimuli via tight junctions (TJs) and adherence junctions (AJs). Defects in TJ and AJ proteins may cause changes in epithelial morphology and integrity and potentially lead to faster trafficking of inflammatory cells through the epithelium. Bronchial epithelial fragility has been reported in asthmatic patients, but little is known about the expression of TJ and AJ proteins in asthma. We studied epithelial expression of zonula occludens-1 (ZO-1) and AJ proteins E-cadherin, alpha-catenin, and beta-catenin in bronchial biopsies from nonatopic nonasthmatic (healthy) subjects (n = 14), and stable atopic asthmatic subjects (n = 22) at baseline conditions. Immunostaining for these proteins was semi-quantified for separate cellular compartments. E-cadherin, alpha-catenin and beta-catenin were present in the cellular membrane and less in the cytoplasm. Only beta-catenin was present in the nucleus in agreement with its potential function as transcription factor. ZO-1 was present in the apicolateral membrane of superficial cells. alpha-Catenin expression was significantly lower in subjects with asthma than without and correlated inversely with numbers of eosinophils within the epithelium. ZO-1 and E-cadherin expression were significantly lower in asthmatic than in nonasthmatic subjects. Expression of beta-catenin was not different. Our results suggest that the lower epithelial alpha-catenin, E-cadherin and (or) ZO-1 expression in patients with atopic asthma contributes to a defective airway epithelial barrier and a higher influx of eosinophils in the epithelium.


Subject(s)
Asthma/metabolism , Cadherins/biosynthesis , Epithelial Attachment/chemistry , Membrane Proteins/biosynthesis , Phosphoproteins/biosynthesis , alpha Catenin/biosynthesis , beta Catenin/biosynthesis , Asthma/pathology , Bronchi/chemistry , Bronchi/pathology , Cell Adhesion Molecules/biosynthesis , Eosinophils/chemistry , Eosinophils/pathology , Epithelial Attachment/pathology , Epithelial Cells/chemistry , Epithelial Cells/pathology , Humans , Immunohistochemistry , Inflammation/metabolism , Inflammation/pathology , Intercellular Junctions/chemistry , Intercellular Junctions/pathology , Mucous Membrane/chemistry , Mucous Membrane/pathology , Zonula Occludens-1 Protein
6.
Arch Oral Biol ; 53(7): 652-8, 2008 Jul.
Article in English | MEDLINE | ID: mdl-18316062

ABSTRACT

OBJECTIVE: This study introduced the usefulness of LYVE-1 immunoreactivity for identification of lymphatic vessels in decalcified tissues, and demonstrated the fine distribution and organization of these vessels in mouse gingiva. DESIGN: After confirming the specificity of anti-mouse LYVE-1, frozen sections of mouse decalcified gingiva were immunostained with the antibody. RESULTS: The LYVE-1-positive lymphatic vessels were clearly found in the connective tissue under the gingival epithelium; these vessels appeared to pass through the lamina propria of the gingiva toward the alveolar crest and run along the external surface of the alveolar bone. The lymphatic vessels were sparse and apart from the oral gingival and sulcular epithelia, while they were dense adjacent to the junctional epithelium. CONCLUSIONS: The dense network of the lymphatic vessels adjacent to the junctional epithelium, which is apparently exposed to foreign antigens, may act as an efficient drainage pathway of the excessive interstitial fluid and immune cells, and play an active role in the immune defense of the gingiva. The present study also revealed the absence of lymphatic connection between gingiva and periodontal ligament.


Subject(s)
Epithelial Attachment/anatomy & histology , Gingiva/anatomy & histology , Glycoproteins/analysis , Lymphatic Vessels/anatomy & histology , Animals , Epithelial Attachment/chemistry , Epithelial Attachment/immunology , Epithelium/anatomy & histology , Epithelium/chemistry , Epithelium/immunology , Gingiva/chemistry , Gingiva/immunology , Immunohistochemistry , Lymphatic Vessels/chemistry , Lymphatic Vessels/immunology , Male , Membrane Transport Proteins , Mice , Microscopy, Confocal , Mouth Mucosa/chemistry , Mouth Mucosa/immunology , Periodontal Ligament/anatomy & histology , Periodontal Ligament/chemistry
7.
J Periodontal Res ; 41(4): 322-8, 2006 Aug.
Article in English | MEDLINE | ID: mdl-16827727

ABSTRACT

BACKGROUND AND OBJECTIVE: The gingival epithelium is the physiologically important interface between the bacterially colonized gingival sulcus and periodontal soft and mineralized connective tissues, requiring protection from exposure to bacteria and their products. However, of the three epithelia comprising the gingival epithelium, the junctional epithelium has much wider intercellular spaces than the sulcular epithelium and oral gingival epithelium. Hence, the aim of the present study was to characterize the cell adhesion structure in the junctional epithelium compared with the other two epithelia. MATERIAL AND METHODS: Gingival epithelia excised at therapeutic flap surgery from patients with periodontitis were examined for expression of adhesion molecules by immunofluorescence. RESULTS: In the oral gingival epithelium and sulcular epithelium, but not in the junctional epithelium, desmoglein 1 and 2 in cell-cell contact sites were more abundant in the upper than the suprabasal layers. E-cadherin, the main transmembranous molecule of adherens junctions, was present in spinous layers of the oral gingival epithelium and sulcular epithelium, but was scarce in the junctional epithelium. In contrast, desmoglein 3 and P-cadherin were present in all layers of the junctional epithelium as well as the oral gingival epithelium and sulcular epithelium. Connexin 43 was clearly localized to spinous layers of the oral gingival epithelium, sulcular epithelium and parts of the junctional epithelium. Claudin-1 and occludin were expressed in the cell membranes of a few superficial layers of the oral gingival epithelium. CONCLUSION: These findings indicated that the junctional epithelium contains only a few desmosomes, composed of only desmoglein 3; adherens junctions are probably absent because of defective E-cadherin. Thus, the anchoring junctions connecting junctional epithelium cells are lax, causing widened intercellular spaces. In contrast, the oral gingival epithelium, which has a few tight junctions, functions as a barrier.


Subject(s)
Cell Adhesion Molecules/biosynthesis , Epithelial Attachment/metabolism , Gingiva/metabolism , Intercellular Junctions/chemistry , Keratins/biosynthesis , Membrane Proteins/biosynthesis , Actins/analysis , Actins/biosynthesis , Aged , Catenins/analysis , Catenins/biosynthesis , Cell Adhesion Molecules/analysis , Epithelial Attachment/chemistry , Epithelial Attachment/cytology , Epithelium/chemistry , Epithelium/metabolism , Female , Fluorescent Antibody Technique , Gingiva/chemistry , Gingiva/cytology , Humans , Intercellular Junctions/metabolism , Keratins/analysis , Male , Membrane Proteins/analysis , Middle Aged
8.
J Periodontol ; 76(5): 696-704, 2005 May.
Article in English | MEDLINE | ID: mdl-15898929

ABSTRACT

BACKGROUND: Syndecans are cell surface heparan sulfate proteoglycans (PG) which can bind to and modulate the action of growth factors and extracellular matrix components (ECM). Syndecan- 1 has been shown to play important roles during early tooth development and wound healing and repair. Among diverse cells and tissues that comprise the periodontium, the junctional epithelium (JE) constitutes a region of significant anatomic and clinical importance, but the nature of inductive signals and molecules involved in its formation is still unclear. Therefore, this work examines if syndecan-1 is associated with formation of JE, and the distribution of other syndecan family members in the epithelium. METHODS: In situ hybridization and immunohistochemical techniques were performed using oral tissues from 4-day-old to 10-week-old mice to investigate the expression of syndecan- 1, -2, -3 and -4 mRNAs and their corresponding proteins. RESULTS: Based on in situ hybridization experiments, all syndecan mRNAs were detected in sulcular epithelium (SE), gingival epithelium (GE), and JE with varying intensity and distribution. Syndecan-1 immunostaining was localized on the cell surface while that of syndecan-2 did not show clear membrane localization. Our experiments in the developing tooth demonstrated that syndecan-1 protein followed characteristic patterns of expression during JE formation and that immunoreactivity for syndecan-1 protein decreased with age when JE cells underwent terminal differentiation. CONCLUSION: Results of syndecan-1 mRNA and protein expression patterns suggested that this proteoglycan might be an important molecule during the formation of JE.


Subject(s)
Epithelial Attachment/growth & development , Membrane Glycoproteins/analysis , Proteoglycans/analysis , Ameloblasts , Animals , Epithelial Attachment/chemistry , Epithelial Attachment/cytology , In Situ Hybridization/methods , Male , Mice , Mice, Inbred ICR , Microscopy, Immunoelectron/methods , Molar , Odontogenesis , RNA, Messenger/analysis , Rabbits , Syndecan-1 , Syndecans , Tooth Eruption
9.
J Periodontal Res ; 34(1): 7-15, 1999 Jan.
Article in English | MEDLINE | ID: mdl-10086881

ABSTRACT

This investigation was undertaken to further study cyclosporin A (CsA)-induced gingival overgrowth. Thirty mg/kg/d of vehicle or CsA solutions were given orally to 6-wk-old male Sprague-Dawley rats. After 4, 9, 14 and 19 wk 2 control and 2 experimental rats were anaesthetized, tissues fixed by intracardiac perfusion of fixative solution and jaws processed for Epon inclusion. Histological and ultrastructural studies conducted in a gingival portion (free gingiva) revealed the presence of hyalinization areas and of multinucleated cells (MCs) containing collagen fibrils (connective tissue), of amorphous areas and disorders of keratinization (epithelia). Histomorphometric evaluation indicated that in the CsA rats the mean cross-sectional area of the free gingiva was 2.52-fold increased compared to the controls. The connective tissue comprised 41.43% of this area (instead of 31.49% in controls). Additional histomorphometric evaluation was performed in 3 groups of free gingival portions: control (C group), CsA-non-respondent (CsA-nR) and CsA-respondent (CsA-R). The cross-sectional gingival areas studied were slightly lower than the mean area of all the control sites previously defined (groups C and CsA-nR) or showed the higher degrees of enlargement (CsA-R). In the CsA-R group the mean cross-sectioned area of the vessel profiles was increased and the number of fibroblast profiles decreased. In the CsA-nR group the number of vessel profiles and that of MCs profiles were increased. In the epithelia of the CsA-R group were increased (a) keratinized epithelia: thickness; thickness of the inner and of the outer compartments; surface area of spinous cell profiles; (b) oral gingival epithelium: number of cell layers (inner compartment); (c) oral sulcular epithelium: surface area of granular cell profiles; (d) junctional epithelium: thickness; number of cell layers. These results indicate that (a) the CsA induced modifications are not limited to enlarged gingiva (b) the overgrowth of the GCT is the result of a vasodilatation and of an increase in the volume of the extracellular matrix and (c) the increase of the epithelial thickness is mainly the result of a cell hypertrophy in the keratinized epithelia and of a cell hyperplasia in the junctional epithelium.


Subject(s)
Cyclosporine/adverse effects , Gingival Overgrowth/chemically induced , Immunosuppressive Agents/adverse effects , Anatomy, Cross-Sectional , Animals , Cell Count , Collagen/analysis , Connective Tissue/chemistry , Connective Tissue/drug effects , Connective Tissue/ultrastructure , Epithelial Attachment/chemistry , Epithelial Attachment/drug effects , Epithelial Attachment/ultrastructure , Epithelium/chemistry , Epithelium/drug effects , Epithelium/ultrastructure , Extracellular Matrix/chemistry , Extracellular Matrix/drug effects , Extracellular Matrix/ultrastructure , Fibroblasts/chemistry , Fibroblasts/drug effects , Fibroblasts/ultrastructure , Follow-Up Studies , Gingiva/blood supply , Gingiva/chemistry , Gingiva/drug effects , Gingiva/ultrastructure , Gingival Overgrowth/metabolism , Gingival Overgrowth/pathology , Hyalin/chemistry , Hyperplasia , Keratins/analysis , Male , Microscopy, Electron , Pharmaceutical Vehicles , Rats , Rats, Sprague-Dawley , Vasodilation/drug effects
10.
J Periodontal Res ; 32(4): 388-400, 1997 May.
Article in English | MEDLINE | ID: mdl-9210093

ABSTRACT

To establish a three-dimensional in vitro test system mimicking the physiological situation of the oral cavity, organotypic co-cultures consisting of primary gingival cells on a collagen matrix with fibroblasts were generated. The histomorphological development after 7 and 14 d revealed close similarity with the non-keratinized gingiva epithelium. Furthermore, as epithelial specific markers synthesis and localization of keratins as well as the deposition of basement membrane components were assessed on frozen sections by immunofluorescence and keratin expression by in situ hybridization. Primary keratinocytes in conventional culture strained positive for keratin K14 and the mucosal differentiation-specific keratins K4 and K13, while primary fibroblasts, isolated from the same tissue source, and also some keratinocytes, were positive for vimentin. In organotypic co-cultures the keratinocytes formed a multilayered epithelium within 14 d containing basal cells and flattened cells in the uppermost layers. Comparable to native non-keratinized gingiva keratin 14 gene expression was clearly detectable in the basal cell compartment but showed extending immunolocalization. In addition, particularly at the early stage (7 d), basally located keratinocytes were also vimentin positive. According to morphological differentiation K4 and K13 were detectable in suprabasal position a the RNA and protein level. The major basement membrane constituents collagen type IV and laminin increased with time revealing first an interrupted and later a fully extended staining underneath the basal cells. Maintenance of basal cell function was further demonstrated by cell proliferation (BrdU incorporation) which was initially high (7 d) but declined towards the later stages (14-21 d). The results demonstrate i) that this co-culture system leads to a stratified surface epithelium with morphological and biochemical characteristics of the non-keratinized gingiva epithelium and ii) that a state of physiological tissue balance was reached, thus rendering a suitable model for tissue compatibility studies.


Subject(s)
Gingiva/chemistry , Gingiva/cytology , Keratinocytes/cytology , Basement Membrane/chemistry , Cell Differentiation/physiology , Cells, Cultured , Coculture Techniques , Collagen/biosynthesis , Epithelial Attachment/chemistry , Epithelial Attachment/cytology , Epithelial Attachment/ultrastructure , Epithelial Cells , Epithelium/chemistry , Epithelium/metabolism , Epithelium/ultrastructure , Fibroblasts/chemistry , Fibroblasts/cytology , Fibroblasts/metabolism , Fibroblasts/ultrastructure , Fluorescent Antibody Technique, Indirect , Gene Expression , Gingiva/ultrastructure , Humans , In Situ Hybridization , Keratinocytes/ultrastructure , Keratins/biosynthesis , Keratins/genetics , Laminin/biosynthesis , Mesoderm/physiology , Organ Specificity , Vimentin/biosynthesis
11.
J Periodontol ; 66(11): 933-43, 1995 Nov.
Article in English | MEDLINE | ID: mdl-8558394

ABSTRACT

The oral, oral sulcular, and junctional epithelia of the natural gingiva each possess distinct patterns of differentiation that are demonstrable both ultrastructurally and by their individual patterns of macromolecular synthesis. The supracrestal tissues reformed around oral implants structurally resemble those of natural gingiva but little is known about phenotype changes occurring in the epithelia. To investigate whether peri-implant epithelia acquire similar patterns of differentiation to those of natural gingiva, biopsies from the supracrestal regions of five oral implants were examined by immunofluorescent methods using a panel of monoclonal antibodies with specificities for individual cytokeratins and ICAM-1, macromolecules which act as markers of the three gingival epithelial phenotypes. The observed staining patterns indicated the formation of oral, oral sulcular, and junctional epithelia which were phenotypically indistinguishable from those of natural gingival epithelia. This degree of reprogramming of epithelial gene expression is a surprising observation and the potential mechanisms leading to the development of those new epithelial phenotypes are discussed in the context of what is known about the development of natural gingiva, in terms of the possible effects of inflammation, and in relation to the known connective tissue influences on epithelial differentiation.


Subject(s)
Dental Implants , Gingiva/anatomy & histology , Cell Differentiation , Epithelial Attachment/chemistry , Epithelial Attachment/cytology , Epithelial Attachment/metabolism , Epithelial Cells , Epithelium/chemistry , Epithelium/metabolism , Fluorescent Antibody Technique , Gingiva/chemistry , Gingiva/metabolism , Humans , Immunophenotyping , Intercellular Adhesion Molecule-1/analysis , Intercellular Adhesion Molecule-1/biosynthesis , Keratins/analysis , Keratins/biosynthesis , Laminin/analysis , Laminin/biosynthesis , Osseointegration
12.
Differentiation ; 59(2): 113-26, 1995 Sep.
Article in English | MEDLINE | ID: mdl-8522068

ABSTRACT

We have determined the patterns of synthesis of cytokeratins and other epithelial marker proteins in the "rests of Malassez" of the periodontium of rabbits and humans, by immunofluorescence microscopy of cryosections prepared from fixed and decalcified rabbit teeth with attached ligament or from manually isolated human periodontal ligaments. Proteins of the major cell structures characterizing epithelial differentiation are present in Malassez cells: a complex set of cytokeratins as well as desmosomal, hemidesmosomal and basal lamina proteins. In addition, we have shown these cytoskeletal and extracellular matrix structures by electron microscopy. The cytokeratin complement of Malassez cells was found to be highly complex, as 8 of the total of 20 known epithelial cytokeratins were detected (nos. 5, 7, 8, 14, 15, 17, 18, 19). This pattern, together with the presence of the desmosomal cadherins Dsg2 and Dsc2 and the cytoplasmic desmosome plaque-associated protein plakophilin 1, indicates that the cells of the rests of Malassez are derived from the basal cell layer of a stratified squamous epithelium rather than from simple epithelial or neuroendocrine epithelial cells. Our observations show that Malassez cells retain the major characteristics of epithelial cells throughout their differentiation from the root sheath epithelium into the rests of Malassez, even though the surface location and the polar tissue architecture that typify epithelial are lost during this process. From this study we further conclude that the specific cytoskeletal complement of the Malassez cells represents an intrinsic gene expression program that neither depends on nor causes the formation of a stratified epithelium. We also compare the specific cytoskeletal features of Malassez cells with those of other persisting epithelial residues and discuss the potential value of these findings in relation to the histogenesis and diagnostic classification of dental and periodontal cysts and tumors.


Subject(s)
Cytoskeletal Proteins/chemistry , Epithelial Attachment/chemistry , Periodontal Ligament/chemistry , Adolescent , Adult , Animals , Cytoskeletal Proteins/biosynthesis , Epithelial Attachment/cytology , Epithelial Cells , Epithelium/chemistry , Extracellular Matrix Proteins/chemistry , Extracellular Matrix Proteins/immunology , Humans , Intermediate Filament Proteins/chemistry , Intermediate Filament Proteins/immunology , Keratins/chemistry , Microscopy, Electron , Microscopy, Fluorescence , Rabbits , Tooth/chemistry , Tooth/ultrastructure
13.
J Periodontal Res ; 30(5): 349-54, 1995 Sep.
Article in English | MEDLINE | ID: mdl-7494177

ABSTRACT

The continuous growth of the rat incisor is associated with renovation of the junctional epithelium and resorption of the periodontal ligament. The circumdental papilla separates the connective tissue suffering resorption from the rest of the gingiva. Laminin and collagen IV were detected by the immunoperoxidase technique on the basement membrane of all regions of the gingival epithelium of the rat incisor, except the internal basal lamina and the internal surface of the circumdental papilla. The internal basal lamina is formed by a granular electron-dense material, without the organization of a typical basal lamina. Areas of the internal surface of the circumdental papilla, negative for laminin and collagen IV, lack the basal lamina. These data suggest that these molecules are not components of the dento-epithelial junction of the distal surface of the rat incisor. In addition, the basal lamina is absent or fragmented on the internal surface of the circumdental papilla, adjacent to the areas of the connective tissue undergoing resorption.


Subject(s)
Collagen/analysis , Epithelial Attachment/chemistry , Epithelial Attachment/ultrastructure , Gingiva/chemistry , Gingiva/ultrastructure , Incisor , Laminin/analysis , Animals , Basement Membrane/chemistry , Basement Membrane/ultrastructure , Desmosomes/chemistry , Desmosomes/ultrastructure , Immunoenzyme Techniques , Incisor/chemistry , Incisor/ultrastructure , Male , Microscopy, Electron , Rats , Rats, Wistar
14.
J Periodontal Res ; 30(2): 97-107, 1995 Mar.
Article in English | MEDLINE | ID: mdl-7539838

ABSTRACT

The unique features of junctional epithelium involve lack of keratinization, limited differentiation and a relatively permeable structure. In order to study the relationship between differentiation and permeability of stratified epithelium a model system was developed. Porcine periodontal ligament epithelial cells were cultured on the polycarbonate nucleopore membrane of the Transwell two-compartment culture system. Within 5 days of culture the cells formed a confluent multilayered structure. Subsequently, maturation of the structure and differentiation of surface cells took place. Transmission electron microscopy showed that the cells were arranged into basal and suprabasal layers with sparse desmosomal attachments and wide intercellular spaces resembling the organization of junctional epithelium. The basal cells attached to a subepithelial basal lamina through numerous hemidesmosomes. The cytokeratin profile of the cultured epithelium (K5, 6, 14, 16, 19) resembled that of the cells of junctional epithelium attached to the tooth surface. The older cultures expressed differentiation markers, K4, K13 and involucrin, thereby resembling sulcular epithelium. The epithelial permeability, measured by diffusion of phenol red, radioactive dextran or methionine tracers, and as transepithelial electrical resistance, decreased with the increased cell number and maturation of the cultures. The new model provides an organotypic culture system which allows to control differentiation of a multilayered periodontal epithelium. It thus may serve as a valuable new tool for studies on the permeability and behaviour of periodontal epithelium under the influence of exogenous and endogenous factors.


Subject(s)
Epithelial Attachment/cytology , Periodontal Ligament/cytology , Analysis of Variance , Animals , Cell Differentiation , Cell Membrane Permeability , Cells, Cultured , Culture Techniques/methods , Desmosomes , Epithelial Attachment/chemistry , Epithelial Attachment/ultrastructure , Keratins/analysis , Microscopy, Electron , Periodontal Ligament/ultrastructure , Swine
15.
Cell Tissue Res ; 278(3): 563-71, 1994 Dec.
Article in English | MEDLINE | ID: mdl-7850866

ABSTRACT

A characteristic layer containing numerous fibrils is associated with the basement membrane of the inner enamel epithelium during the early stages of odontogenesis. However, its nature is not well understood. In this study, the layer was examined with high-resolution electron microscopy and immuno-histochemical staining. Tooth germs of monkeys (Macaca fuscata) were studied and each fibril in the layer was found to be a tubular structure, 8-9 nm in width, resembling a "basotubule", the tubular structure previously observed in various basement membranes. The space between the fibrils was filled with a network formed by irregular anastomosing strands with an average thickness of 4 nm; these strands resembled the "cords" forming the network in the lamina densa of basement membranes. After immunoperoxidase staining, fine threads immunoreactive for laminin staining were seen winding along the strands of the network, and 1.5-nm wide filaments, immunoreactive for type IV collagen, took the form of a network arrangement. The 5-nm-wide ribbon-like structures associated with the strands were identified as heparan sulfate proteoglycan by immunostaining. These results are similar to those obtained for the cord network of the lamina densa. The "fibrillar layer" therefore represents a highly specialized lamina fibroreticularis of the basement membrane of the inner enamel epithelium, and rich in basotubules.


Subject(s)
Epithelial Attachment/ultrastructure , Tooth Germ/ultrastructure , Animals , Basement Membrane/chemistry , Basement Membrane/ultrastructure , Collagen/analysis , Cytoskeleton/chemistry , Cytoskeleton/ultrastructure , Dental Enamel/chemistry , Dental Enamel/ultrastructure , Epithelial Attachment/chemistry , Heparitin Sulfate/analysis , Immunoenzyme Techniques , Macaca , Mesoderm/chemistry , Mesoderm/ultrastructure , Microscopy, Electron , Odontogenesis , Tooth Germ/chemistry
16.
J Periodontal Res ; 29(3): 185-95, 1994 May.
Article in English | MEDLINE | ID: mdl-7515960

ABSTRACT

This study uses cytokeratins (CK) as markers to investigate the phenotype of the junctional epithelium (JE) in partially erupted human teeth. The gingival samples, which were clinically healthy, were carefully dissected from the teeth. Cryostat sections were cut for histological staining, immunofluorescence microscopy and gel electrophoresis. Cytokeratins were extracted after microdissection. The basal and suprabasal epithelial cell markers, cytokeratins 4, 5, 13, 14 and 19 were detected with specific monoclonal antibodies. They showed that the junctional epithelium in erupting teeth has a complex topography. The cytokeratin immunohistochemical profile distinguished between the primary junctional epithelium (CK 5, 14 and 19 in basal and suprabasal cells and CK 13 faintly stained throughout the suprabasal layers) and the adjacent epithelium that had the same cytokeratin profile as the sulcular epithelium (CK 5, 14 and 19 in basal cells and CK 4 and 13 intensively stained in the suprabasal cells). Extraction, two-dimensional electrophoresis and western blotting showed that this transitional JE during eruption also contained CK 6, 16 and perhaps CK 4. Thus, the JE in erupting teeth shows patterns of CK distribution that are very similar to that of developing oral epithelia.


Subject(s)
Epithelial Attachment/chemistry , Keratins/chemistry , Tooth Eruption , Adolescent , Adult , Cell Differentiation , Child , Electrophoresis, Gel, Two-Dimensional , Epithelial Attachment/anatomy & histology , Humans , Immunoblotting , Immunohistochemistry , Keratins/analysis , Microscopy, Fluorescence
17.
Brain Res Mol Brain Res ; 21(3-4): 235-46, 1994 Feb.
Article in English | MEDLINE | ID: mdl-8170348

ABSTRACT

The immunofluorescent distribution of ZO-1, a tight junction-associated protein, was studied in murine circumventricular organs. These regions generally express a less restrictive blood-brain barrier than is found in other areas of the CNS. In the remaining brain parenchyma, where a characteristic blood-brain barrier exists, ZO-1 was localized in discrete, continuous lines along blood vessels, presumably in association with endothelial cell tight junctions. The ependymal cells in the ventricular walls displayed a more punctate pattern of ZO-1 distribution, indicative of discontinuous tight junctions. In two of the circumventricular organs examined, the median eminence and the subfornical organ, many capillaries lacked detectable ZO-1 immunoreactivity while the apical aspects of the specialized ependymal cells (tanycytes) revealed an unbroken ZO-1 distribution. Scant labelling of ZO-1 in blood vessels was found in the area postrema, and only weak and discontinuous ZO-1 labelling was present in the ventricular wall. Capillaries of the organum vasculosum laminae terminalis expressed ZO-1 immunoreactivity which was comparable to the pattern observed in CNS regions with typical blood-brain barrier. The subcommissural organ, known to contain a blood-brain barrier, also displayed continuous ZO-1 staining in blood vessels. Unbroken ZO-1 distribution was observed in the specialized ependymal cells adjacent to both the organum vasculosum laminae terminalis and subcommissural organ. These immunocytochemical data demonstrate a distribution of ZO-1 in CNS parenchyma outside the circumventricular organs that is consistent with an organization of tight junctions which prevent free paracellular exchange of substances between blood and neuropil but which allow for continuity between CSF and the neuronal environment. The ZO-1 staining pattern in blood vessels and ventricular walls of the circumventricular organs is heterogeneous despite the prevalent absence of a functional blood-brain barrier.


Subject(s)
Blood-Brain Barrier/physiology , Brain Chemistry/physiology , Epithelial Attachment/chemistry , Membrane Proteins/analysis , Phosphoproteins/analysis , Animals , Male , Median Eminence/chemistry , Mice , Mice, Inbred BALB C , Subcommissural Organ/chemistry , Subfornical Organ/chemistry , Zonula Occludens-1 Protein
18.
Anat Rec ; 238(1): 1-14, 1994 Jan.
Article in English | MEDLINE | ID: mdl-8116883

ABSTRACT

The region of epithelial apposition with a tooth surface is the site of an unusual stratified integument, the junctional epithelium, which combines tight attachment to the tooth, cell turnover, tissue permeability, and epithelial versatility into the first line of defense against periodontal destruction by oral pathogens. To better understand the structure and function of the junctional epithelium we have reviewed its developmental and cell biology, and undertaken a multidisciplinary analysis of its composition in the pig, an omnivore whose dietary and dental development and occlusion patterns are similar to the human condition, and which, because of its size, is more readily amenable to experimental manipulation. The porcine junctional epithelium was also compared with this well-described epithelium in the rat. Morphological analyses by light microscopy and scanning and transmission electron microscopy showed the porcine junctional epithelium and epithelial attachment were similar to that in the rat except that apically, extracellular matrix lamellae associated with the internal basal lamina were more complex, and more coronally there was extensive layering of a dental cuticle-like material. Biochemical analysis of the porcine junctional epithelium by dissociative extraction and SDS-PAGE revealed the presence of some proteins not present in gingival epithelium. Together, these studies show that the porcine junctional epithelium has predictable morphological and biochemical features which establish the pig as an advantageous model to study the basic and clinical biology of this unique epithelium.


Subject(s)
Epithelial Attachment/ultrastructure , Molar/ultrastructure , Animals , Epithelial Attachment/chemistry , Humans , Male , Microscopy, Electron , Microscopy, Electron, Scanning , Models, Biological , Molar/chemistry , Proteins/isolation & purification , Rats , Rats, Wistar , Species Specificity , Swine
19.
J Dent Res ; 72(2): 502-7, 1993 Feb.
Article in English | MEDLINE | ID: mdl-8423247

ABSTRACT

Localization of cathepsin D was studied in the junctional epithelium (JE) of healthy rat gingivae by immuno-light and -electron microscopy, by means of both the avidin-biotin-peroxidase complex method and a colloidal gold IgG method. At the light-microscopic level, cathepsin D was demonstrated in the JE and oral sulcular epithelium (OSE). Cathepsin D immunoreactivity was remarkable in the coronal portion of the JE and decreased toward its apical portion. However, cathepsin D immunoreactivity in the basal cell layer of the JE was negligible or negative. In the OSE, the granular layer was positive for cathepsin D. In the adjacent connective tissue, many macrophage-like cells (not clear at this level) close to the basal cell layer showed strong immunoreactivity. At the electron microscopic level, cathepsin D was found in the primary lysosomes and trans-cisternae of Golgi apparatus in the JE cells. These lysosomes were often fused together or were fused with cathepsin D-negative intracytoplasmic vacuoles to form secondary lysosomes, which indicated that intracellular digestion may have been in progress. However, neutrophils contained few gold particles based on cathepsin D. It is likely that the amounts of cathepsin D contained in the JE cells and macrophages are larger than those of cathepsin D contained in the neutrophils. These findings provided morphological evidence that JE cells have the same endocytotic capacity as macrophages and neutrophils, and that JE cells participate in the intracellular digestion that is carried out by lysosomal enzymes such as cathepsin D. It is suggested, in addition, that maximum intracellular digestion occurs in the coronal portion of the JE.


Subject(s)
Cathepsin D/analysis , Epithelial Attachment/metabolism , Animals , Cathepsin D/metabolism , Endocytosis , Epithelial Attachment/chemistry , Epithelial Attachment/ultrastructure , Female , Immunohistochemistry , Male , Microscopy, Immunoelectron , Phagosomes/ultrastructure , Rats , Rats, Wistar , Vacuoles/ultrastructure
20.
J Periodontal Res ; 28(1): 49-59, 1993 Jan.
Article in English | MEDLINE | ID: mdl-7678864

ABSTRACT

Fourteen specimens of periodontal pockets and the associated marginal gingiva were collected and either frozen for examination using antibodies against various defined cytokeratin specificities or processed for 2-dimensional gel electrophoresis. The epithelium forming the pocket lining typically extended into the connective tissue of the pocket wall in the form of a network of finger-like strips. Immunocytological staining indicated that keratins (K) 5, 6, 14 and 19 were expressed by almost all cells of the pocket lining and K13 and K16 by the suprabasal cells. The coronal region of the pocket lining showed some cells staining for K4. Staining for K8 and K18 was seen in the apical region of the pocket lining and in the finger-like extensions of epithelium into the connective tissue. Compared with normal gingiva, the sulcular and the oral gingival epithelia showed a marked increase in staining for K19. Surprisingly, the pattern of keratin expression of the epithelium of the pocket lining was found to be essentially similar to that of normal junctional epithelium and the anatomical position of the boundaries between each epithelial phenotype were not significantly altered. These patterns of keratin expression were confirmed by the 2D electrophoretic analyses of microdissected regions of epithelium. The potential significance of inflammation to the epithelial changes associated with pocket formation is discussed.


Subject(s)
Gingivitis/metabolism , Keratins/metabolism , Periodontal Pocket/metabolism , Adult , Antibodies, Monoclonal , Electrophoresis, Gel, Two-Dimensional , Epithelial Attachment/chemistry , Epithelial Attachment/metabolism , Epithelium/chemistry , Epithelium/metabolism , Humans , Immunoenzyme Techniques , Keratins/analysis , Middle Aged
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