Inflammatory Cells Can Alter the Levels of H3K9ac and γH2AX in Dysplastic Cells and Favor Tumor Phenotype.

Oral potentially malignant disorders (OPMD) are clinical presentations that carry an increased risk of cancer development. Currently, epithelial dysplasia grade is based on architectural and cytological epithelial changes and is used to predict the malignant transformation of these lesions. However, predicting which OPMD will progress to a malignant tumor is very challenging. Inflammatory infiltrates can favor cancer development, and recent studies suggest that this association with OPMD lesions may be related to the etiology and/or aggressive clinical behavior of these lesions. Epigenetic changes such as histone modifications may mediate chronic inflammation and also favor tumor cells in immune resistance and evasion. This study aimed to evaluate the relationship between histone acetylation (H3K9ac) and DNA damage in the context of dysplastic lesions with prominent chronic inflammation. Immunofluorescence of "low-risk" and "high-risk" OPMD lesions (n = 24) and inflammatory fibrous hyperplasia (n = 10) as the control group was performed to assess histone acetylation levels and DNA damage through the phosphorylation of H2AX (γH2AX). Cell co-culture assays with PBMCs and oral keratinocyte cell lines (NOK-SI, DOK, and SCC-25) were performed to assess proliferation, adhesion, migration, and epithelial-mesenchymal transition (EMT). Oral dysplastic lesions showed a hypoacetylation of H3K9 and low levels of γH2AX compared to control. The contact of dysplastic oral keratinocytes with PBMCs favored EMT and the loss of cell-cell adhesion. On the other hand, p27 levels increased and cyclin E decreased in DOK, indicating cell cycle arrest. We conclude that the presence of chronic inflammation associated to dysplastic lesions is capable of promoting epigenetic alterations, which in turn can favor the process of malignant transformation.

Macrophages CD163+ and Factor XIIIa+ Provide a First-Line Defence against Proliferative Verrucous Leukoplakia Antigens.

This study aimed to evaluate the density of the dendritic cells (DCs) and macrophages in oral leukoplakia (OL) and proliferative verrucous leukoplakia (PVL) by immunohistochemical analysis. We analysed paraffined tissue samples of PVL (n = 27), OL (n = 20), and inflammatory fibrous hyperplasia (n = 20) as the control group using the immunomarkers for DCs (CD1a, CD207, CD83, CD208 and CD123) and macrophages (CD68, CD163, FXIIIa and CD209). A quantitative analysis of positive cells in the epithelial and subepithelial areas was determined. Our results showed a reduction in CD208+ cells in the subepithelial area of the OL and PVL compared to the control. Additionally, we found a higher density of FXIIIa+ and CD163+ cells in the subepithelial area in PVL compared to the OL and control. Four-way MANOVA revealed a relationship between increased CD123+ cell density in the subepithelial area of "high-risk" samples regardless of disease. Macrophages provide the first line of defence against PVL antigens, suggesting a distinct pattern of innate immune system activation in PVL compared to OL, which may contribute to the complexity and the high rate of malignant transformation in the PVL.

High density of CD8 T cell and immune imbalance of T lymphocytes subsets are associated with proliferative verrucous leukoplakia.

Oral leukoplakia (OL) and proliferative verrucous leukoplakia (PVL) are oral potentially malignant disorders (OPMDs) that microscopically show no or varying degrees of dysplasia. Even sharing clinical and microscopic aspects, PVL shows a more aggressive clinical behaviour, with a malignant transformation rate greater than 40%. Inflammatory infiltrate associated with dysplastic lesions may favour malignant transformation of OPMDs. This study aimed to evaluate the density of T cells and cytokines in dysplastic lesions from OL and PVL patients. Additionally, we evaluated whether soluble products produced in vitro by dysplastic keratinocytes are capable of modulating apoptosis rates and Th phenotype (Th1, Th2, Th17 and Treg) of peripheral blood mononuclear cells. The density of CD3, CD4 and CD8 T cells was assessed by immunohistochemistry. Cytokines and chemokines profile from frozen tissue samples were analysed using the LUMINEX system. Apoptosis rates and Th phenotype modulation were evaluated by flow cytometry. Our results showed an increase in the number of CD8 T cell in the subepithelial region from PVL dysplastic lesions in relation to OL samples. PVL showed increased levels of IL-5 and a decrease in IL-1ß and IFN-γ levels compared to OL. Soluble products of PVL and oral carcinoma cell cultures were able to reduce apoptosis rate and promote an imbalance of Th1/Th2 and Th17/Treg. The high-subepithelial density of CD8 T cells and immune imbalance of T lymphocytes subsets probably play an important role in the pathogenesis of PVL and may explain its more aggressive behaviour in relation to OL.

The role of CD68+ and CD163+ macrophages in immunopathogenesis of oral lichen planus and oral lichenoid lesions.

Macrophages are phagocytic cells with essential participation in immunological events of the oral cavity. However, the role of these cells in oral lichen planus (OLP) and oral lichenoid lesions (OLL) remains unclear. The present study aimed to evaluate the density of macrophages in OLP and OLL, and to compare it with that of oral inflammatory fibrous hyperplasia (OIFH) (control group). 14 cases of OLP, 14 cases of OLL and 14 cases of OIFH were selected for immunohistochemical analysis of CD68+ (M1) and CD163+ (M2) macrophage expression. CD68+ and CD163+ macrophages densities were measured in the intraepithelial and subepithelial areas. The statistical tests used were multivariate analysis of variance, as well as a correlation and linear regression. OLP has more CD68+ macrophages when comparing with OLL (p = 0.001) and OIFH (p = 0.045). There is a very strong relationship between the macrophages types (p < 0.0001) in OLP and OLL. The linear regression showed that to OLL development (p < 0.0001/R2' = 0.9584), the presence of different types of macrophages are more essential than to OLP (p < 0.0001/R2' = 0.8983). However, in the OLP these dependencies are also largely. CD68+ macrophages may be associated with immunopathogenesis of OLP, indicating a pro-inflammatory activity and regulatory role in the type of T-cell response. Besides, CD68+ macrophages can cooperate in the diagnosis of OLP. These results are essential to future studies that seek a therapeutic target for OLP and OLL.

Immunohistochemical evaluation of Langerhans cells in oral lichen planus and oral lichenoid lesions.

OBJECTIVE: the aim of this study was to evaluate the density of Langerhans cells in oral lichen planus (OLP) and oral lichenoid lesions (OLL). DESIGN: 14 cases of OLP, 15 cases of OLL and 14 cases of oral inflammatory fibrous hyperplasia (OIFH), were selected for immunohistochemical analysis of CD1a, CD207 and S100 expression. The OIFH group was subdivided according to the presence (OIFHL n = 14) or absence (OIFHNL n = 14) of lichenoid inflammatory infiltrate. Positive cells were counted in intraepithelial and subepithelial areas. Results were analyzed by multivariate comparative analysis, correlation analysis, linear regression models and Student's T-test. RESULTS: A significantly higher amount of CD207+ cells in OLL vs OLP was observed (p = 0.015). The prevailing reticular pattern observed was CD207high for OLP (p = 0.0329). A statistically significant difference in the expression of CD1a and CD207 was observed for intraepithelial vs subepithelial areas (p = 0.024 and p=0.015, for CD1a and CD207, respectively). Significant correlations were also observed between the expression of CD1a + and CD207+ cells in the pathogenesis of OLP and OLL. CONCLUSION: High levels of CD207+cells in OLP compared with OLL may help explain the differences in the immunopathogenesis of both diseases. Additionally, CD1a + and CD207+ cells appear to be more essential to immunopathogenesis of OLL than to the pathogenesis of OLP.