Ex vivo and in vitro study of the modulation of innate and adaptive immunity by dysplastic keratinocytes in oral leukoplakia and proliferative verrucous leukoplakia

Author(s)

Darcy Fernandes

• Document type: Doctoral Thesis
• Press: Araraquara. 2020-05-01.
• Institution: Universidade Estadual Paulista (Unesp). Faculdade de Odontologia. Araraquara
• Defense date: 2020-03-13
• Advisor: Andreia Bufalino

Abstract

Oral squamous cell carcinoma (SCC) represents more than 95% of all malignant neoplasms that affect the oral cavity and these tumors are often preceded by clinical changes that have an evident potential for malignant transformation, which are called potentially malignant oral disorders (OPMDs). Among these, oral leukoplakia (OL) has a malignant transformation rate that varies from 0.2% to 17.5%; however, another OPMD known as proliferative verrucous leukoplakia (PVL) has a persistent and progressive behavior for malignancy, with a malignant transformation rate greater than 70%. Unlike OL, a risk factor such as tobacco, alcohol and areca nut does not seem to be associated with the development of PVL. Additionally, PVL often presents an inadequate response to all treatment modalities, suffers rapid dissemination through oral sites and often shows recurrence. Recent studies suggest that the inflammatory infiltrate associated with leukoplastic lesions in a patient with PVL may be related to the etiology and / or aggressive clinical behavior of this OPMD. Thus, a comparative analysis was performed between OL and PVL samples that consisted of: (1) evaluating the percentage and identify subtypes of auxiliary T lymphocytes and activation status of cytotoxic T lymphocytes, (2) evaluating the density and activation status of dendritic cells, and (3) determine the effect of soluble products of dysplastic cells on the modulation of innate and / or adaptive immunity and whether such modulation confers an advantage for progression of the tumor. To achieve these goals, immunohistochemical expression of markers for Langerhans dendritic cells (DC), immature and mature submucous DC, plasmacytoid DC, T and B lymphocytes were used. In addition, cell culture assays were performed to access the potential modeling of soluble factors produced by PVL-derived cell cultures on peripheral blood mononuclear cells (PBMCs). These trials included the assessment of apoptosis and immunophenotyping rates of PBMCs after contact with conditioned culture media from different cell cultures, including normal keratinocytes (HOK), dysplastic (DOK and DOK-PVL) and tumor (SCC-25). The results of the immunohistochemical analysis showed a global reduction in T and B cells accompanied by an inhibition of Th17 cell activation and the consequent induction of an immune response mediated by Treg cells in PVL, seems to favor the more aggressive behavior of this OPMD in relation to OL. In addition, Langerhans DC, matures and plasmacytoids are reduced in the epithelial and subepithelial compartments in the OL and PVL in relation to the control. In contrast, immature DCs are increased in the epithelial and subepithelial compartments of these two OPMD. Together, these findings favor an inadequate presentation of pre-tumor antigens, facilitating immunological tolerance. The experiments with cell culture also revealed that the soluble products from dysplastic keratinocytes may to change the PBMCs phenotype favoring the malignant transformation of OPMD. Thus, the results of this study indicate that soluble product from PVL-derived keratinocytes are capable of differently modulating the immune response of this DPMO in relation to OL, and this modulation is accompanied by the gain of advantages that favor the tumorigenesis process. (AU)

• FAPESP's process: 17/01798-7 - In vitro study of the modulation of innate and adaptive immunity by dysplastic keratinocytes
• Grantee: Darcy Fernandes
• Support Opportunities: Scholarships in Brazil - Doctorate