Langerhans cells express matrix metalloproteinases 9 and 2 and tissue inhibitors of metalloproteinases 1 and 2 in healthy human gingival tissue and in periodontitis.

BACKGROUND: As antigen-presenting cells, Langerhans cells may play an important role in the initiation and maintenance of periodontal disease. This study is the first report that extends our knowledge of the expression of matrix metalloproteinases and their endogenous tissue inhibitors by Langerhans cells in healthy and diseased gingival tissues. METHODS: Single and double immunolabeling procedures were carried out using monoclonal antibodies against CD1a, matrix metalloproteinases 2 and 9, and tissue inhibitors of matrix metalloproteinases 1 and 2, and analyzed by conventional and confocal microscopes. RESULTS: Langerhans cells expressed matrix metalloproteinases 2 and 9, and tissue inhibitors of matrix metalloproteinases 1 and 2 in healthy and diseased gingival tissues. The tissue inhibitors of matrix metalloproteinase-positive Langerhans cells were mainly observed in the upper epithelial layers. Matrix metalloproteinase 9-positive Langerhans cells were observed especially during periodontitis and in the basal epithelial layer or crossing the basement membrane. CONCLUSION: During periodontal disease, changes in the expression of matrix metalloproteinases and their tissue inhibitors by gingival Langerhans cells could be implicated in the migration of the cells towards the connective tissue.

Is collagen breakdown during periodontitis linked to inflammatory cells and expression of matrix metalloproteinases and tissue inhibitors of metalloproteinases in human gingival tissue?

BACKGROUND: Evidence of the role of matrix metalloproteinases (MMPs) produced by resident and inflammatory cells in periodontal destruction is now well established. The purpose of this study was to quantify, in healthy and diseased upper gingival connective tissue, the area fraction (AA%) occupied by collagen fibers, the cell number belonging to inflammatory cell subsets, and the amounts of MMPs and TIMPs (tissue inhibitors of MMPs) in order to investigate the possible correlations, if any, between such molecules, collagen loss, and inflammatory cell subsets. METHODS: Gingival tissue specimens from 6 healthy controls (C) and 6 patients with severe periodontitis (P) were divided into 2 groups. The first group of specimens was frozen and used for the staining of collagen fibers by sirius red F3Ba and for immunohistochemistry with antibodies against CD8, CD4, CD22, CD68, and TIA-1 molecules. The second group was used for organ culture, zymography, Western blotting, and dot blotting. Morphometric and automated image analysis was performed for the evaluation of the area fraction occupied by collagen fibers, the number of inflammatory cell subsets and for enzymatic activities developed by MMPs, and the amounts of TIMPs expressed during periodontal disease. RESULTS: In group P, the area fraction of collagen fibers (33 +/- 10%) was significantly decreased (P < 0.0002) when compared to group C (60 +/- 7%), and was correlated with the number of all inflammatory cells and amounts of MMPs and TIMPs. In group P, there were significant increases of CD8+, CD22+, CD68+, and TIA-1+ cells, as well as increases in the amounts of MMP-1, MMP-2, MMP-3, MMP-9, and the active form of MMP-9. The active form of MMP-9 and the amount of TIMP-1 were positively correlated with the number of CD22+, CD68+, and TIA-1+ cells. CONCLUSIONS: The present study showed an imbalance between MMPs and TIMPs associated with the pathologic breakdown of the extracellular matrix during periodontitis. The active form of MMP-9 could be a marker for the clinical severity of periodontal disease.

Quantitative morphological analysis of Langerhans cells in healthy and diseased human gingiva.

Langerhans cells (LC) are implicated in the initiation and maintenance of inflammatory periodontal diseases. The purpose of this immunohistological study using morphometric and automated image analysis was to determine the morphological features of CD1a+ LC in healthy and inflammatory gingiva according to their localisation in the upper epithelium or the basal layer. The study was on gingival samples from 11 healthy controls (C), eight patients with gingivitis (G) and 12 patients with severe chronic adult periodontitis (P). The results show that in the basal layer of all experimental groups, the perimeter, surface and equivalent diameter of CD1a+ LC were significantly decreased (P<0.005) when compared with those in the upper epithelium of the same group. Furthermore, CD1a+ LC had become more rounded, reflected by a significant increase in form factor (P<0.005), when located close to the epithelial basal membrane. In the upper epithelium of group P, the perimeter, surface and equivalent diameter of CD1a+ LC were significantly decreased (P<0. 05) and the form factor significantly increased (P<0.05) when compared with the upper epithelium of group C. This work provides evidence for important morphological variations in CD1a+ LC according to their location within the epithelium and the severity of the periodontal disease. The observed morphological changes may reflect a cellular adaptation during the epithelial transmigration and could eventually be involved in immune stimulation during periodontitis.

Collagen fibers and inflammatory cells in healthy and diseased human gingival tissues: a comparative and quantitative study by immunohistochemistry and automated image analysis.

BACKGROUND: Periodontal disease is histologically characterized by the degradation of extracellular matrix components associated with a gingival infiltration of inflammatory cell populations. The purpose of this in situ study was to quantify inflammatory cell subsets and the area fraction (AA%) occupied by collagen fibers in healthy and diseased upper gingival connective tissue in order to investigate the association, if any, between collagen loss and inflammatory cell infiltrate. METHODS: Paraffin gingival tissue sections from 10 healthy controls (C), 9 patients with gingivitis (G), and 10 patients with severe chronic periodontitis (P) were immunohistochemically stained by antibodies against CD45, CD3, CD8, CD20, CD68, TIA-1, and GrB molecules, and the collagen fibers were stained using sirius red F3Ba. The quantitative evaluations of inflammatory cell numbers and the AA% occupied by collagen fibers were performed by morphometric and automated image analysis. RESULTS: In group P, CD45+, CD20+, CD68+, TIA-1+, and GrB+ cell numbers were significantly increased (P<0.05) when compared to both C and G groups. The present study revealed significant differences (P <0.01) between means of AA% observed in group C (63%), group G (46%), and group P (26%), and AA% of group G and group P was inversely correlated with the numbers of TIA-1+ cells (P<0.01) and GrB+ cells (P<0.01 and P<0.05, respectively). CONCLUSIONS: This study showed great differences in the number of the distinct inflammatory cell subsets according to the severity of the periodontal disease and suggested that activated cytotoxic cells could play a pivotal role in the loss of collagen fibers observed during these pathological states. During periodontitis, collagen loss was significantly correlated with all inflammatory cell subset numbers. Finally, the quantitative evaluation of the area fraction occupied by gingival collagen fibers may reflect the clinical severity of the periodontal disease.

Immunohistological and morphometric analysis of intra-epithelial lymphocytes and Langerhans cells in healthy and diseased human gingival tissues.

Periodontal diseases are histologically characterized by an infiltration of several inflammatory cell populations into the gingival epithelium and connective tissue, associated with degradation of extracellular matrix components. The purpose of this in situ study was to evaluate the inflammatory state of gingival tissues by the number of intra-epithelial lymphocyte (IEL) subsets and the area fraction (AA%) occupied by collagen fibres in the upper gingival connective tissue, and also to evaluate the number of CD1a+ Langerhans cells (LC) in order to show correlation(s), if any, between these histological findings. The gingival samples were from 10 clinically healthy controls (group C), 8 patients with gingivitis (group G) and 9 with chronic adult periodontitis (group P). A quantitative evaluation of the number of cell populations (CD1a+, CD45RB+, CD3+, CD8+, CD20+, TIA-1+ and GrB+ cells) and the area fraction (AA%) occupied by collagen fibres in the upper gingival connective tissue was made by morphometric and automated image analysis. The results showed that, compared with group C, all IEL subset numbers were significantly increased (p<0.05) in G and P groups, CD20+ excepted. In addition, there was a significant increase in the cytotoxic TIA-1+ IEL number (p<0.05) in group P when compared with group G. The study also showed a significant decrease in the number of CD1a+ LC in groups G and P (p<0.02 and p<0.001, respectively) when compared with group C. No significant difference was found in CD1a+ LC number between groups G and P. The determination of coefficients of correlation (r) with data obtained for each patient showed that in group G, CD1a+ LC number was significantly correlated with CD45RB+ (p<0.05) and CD3+ (p<0.01) IEL numbers whereas during periodontitis, CD1a+ LC number was significantly and inversely correlated with CD20+ (p<0.01), cytotoxic TIA-1+ (p<0.01) and with activated cytotoxic GrB+ (p<0.01) IEL numbers. Moreover, in group P a significant (p<0.05) positive correlation was shown between CD1a+ LC number and the AA% occupied by collagen fibres. This work demonstrates a decrease in CD1a+ LC number according to the severity of the periodontal disease estimated by the number of IEL and by the area fraction occupied by collagen fibres in human gingiva. The decrease of such cells could represent a way to avoid immune overstimulation.

Immunohistologic and morphometric analysis of cytotoxic T lymphocytes in gingivitis.

BACKGROUND: Gingivitis is an inflammatory phenomenon localized in gingival tissues and histologically characterized by an infiltration of several inflammatory cell populations. The purpose of this study was to characterize, localize, and quantify in situ inflammatory and cytotoxic T lymphocytes using immunolabeled gingival tissue sections in order to specify their implication during human gingivitis, since it is well known that such cells play an important role in the defense against bacterial elements. METHODS: Paraffin gingival tissue sections from 7 patients with gingivitis (G) and from 7 clinically and histologically healthy controls (C) were immunohistochemically stained by specific antibodies (anti-CD45, anti-CD3, anti-CD8, anti-CD20, anti-TIA-1, anti-GrB, and anti-CD68), allowing the quantification of inflammatory cells in upper gingival epithelium (Ep), in the basal epithelium layer (BEp), and in upper connective tissue (CT). Collagen fibers were stained by sirius red F3Ba in order to evaluate, by morphometric and automated image analysis, the surface occupied by collagen bundles and to histologically confirm the absence of pathology of the clinically selected healthy controls. RESULTS: In the gingivitis group, CD45+, CD3+, CD8+, TIA-1+, and GrB+ lymphocyte numbers were significantly increased in Ep (P<0.05); and CD45+, CD3+, and TIA-I+ lymphocyte numbers were significantly increased in BEp (P <0.05) compared respectively to Ep and BEp of group C. In Ep of group G, mean CD8+/CD3+ cell ratio was significantly increased (P<0.05) compared to BEp and CT, and 25% of TIA-1+ cytotoxic cells were activated GrB+ cells. CONCLUSIONS: The present study suggests that intraepithelial cytotoxic T lymphocytes play an important role during gingivitis and CD8 expression and that activation of TIA-1+ cytotoxic cells could be induced in Ep in response to epithelial environment. Thus, gingival epithelial tissue, which is generally only considered as a physical barrier, in fact contains numerous immune cell populations preventing the infiltration of pathogenic elements into the connective tissue. Particular clinical attention must be taken for the preservation of the epithelial tissue integrity.