Function of laminins and laminin-binding integrins in gingival epithelial cell adhesion.

BACKGROUND: In human gingiva, epithelial cells attach to their adjacent tissues by means of specialized molecular adhesion complexes and a basement membrane. Little is known about the synthesis of adhesion proteins by gingival keratinocytes; we, therefore, studied how cultured immortalized gingival epithelial cells produce laminins and express laminin-binding integrins. We presumed that different laminins and integrins would be involved in the adhesion of gingival epithelial cells. METHODS: We cultured gingival keratinocytes and studied their production of laminins and expression of integrins using immunofluorescence microscopy, immunoprecipitation, and immunoblotting methods and by quantitative cell adhesion experiments. We also studied how gingival tissue expresses these adhesion proteins in vivo by using immunofluorescence microscopy. RESULTS: In immunofluorescence microscopy, the cells were seen to organize chains of laminin-5 (alpha3beta03gamma2) to extracellular patches, whereas the alpha5 chain of laminin-10 (alpha5betalgamma1) could only be seen intracellularly. Of the laminin-binding integrin subunits, integrin a6 subunit was organized to dotted arrays, typical of prehemidesmosomal adhesions, whereas integrin alpha3 subunit was located at cell-cell junctions, in prehemidesmosomal structures, and at some locations also in small focal-contact like patches. Integrin beta1 subunit was found at cell-cell junctions and in focal contacts. Immunoprecipitation experiments showed that the cells synthesize and secrete chains of laminin-5 and laminin-10. In quantitative cell adhesion experiments, the cells adhered efficiently to these laminins by using cooperatively integrin alpha3beta1 and alpha6beta1 integrin complexes. None of the other known laminin-binding integrin subunits appeared to be significantly involved in cell adhesion to these laminin isoforms. CONCLUSIONS: Our results provide new information on gingival epithelial cell adhesion and extracellular matrix production and may thus aid in the understanding of periodontal physiology.

An organotypic in vitro model that mimics the dento-epithelial junction.

BACKGROUND: The dento-epithelial junction forms the primary periodontal defense structure against oral microbes. The cells of the junctional epithelium (JE) attach both to a basement membrane (BM) facing the connective tissue and to a hard dental tissue by structurally similar but molecularly distinct mechanisms. Here we describe a new organotypic cell culture model for the dento-epithelial junction comprising not only epithelial and mesenchymally derived components, but also a tooth surface equivalent. METHODS: Rat palatal keratinocytes were seeded on fibroblast-collagen gels. A tooth slice was placed on top of the epithelial cells and the multilayer cultures were grown at the air-liquid interface. Formation of the epithelial structures, BM components, and the epithelial attachment to the tooth surface were studied by immunofluorescence and light and electron microscopy. The findings were compared to the structure of the dento-epithelial junction in vivo. RESULTS: A well-differentiated stratified epithelium was formed. Under the tooth slice the epithelium remained thin and non-differentiated. Attachment of the epithelial cells to the tooth surface was mediated by hemidesmosomes (HDs) as in vivo. Laminin-5 (Ln-5) was present in the extracellular matrix (ECM) between the tooth and the epithelium as well as in the BM structure between the epithelium and the fibroblast-collagen matrix. Instead, Ln-10/11 was present only at the mesenchymal tissue side as is known to be the case in vivo. CONCLUSIONS: The organotypic model presented expresses the characteristic structural and molecular features of the dento-epithelial junction and may be applied for studying physiological and pathological processes in the epithelial attachment.