Notch and Wnt signaling pathway in the modulation of the immune response and tissue destruction associated with periodontal diseases

• Grant number: 11/23136-0
• Support Opportunities: Scholarships in Brazil - Post-Doctorate
• Effective date (Start): May 01, 2012
• Effective date (End): February 02, 2014
• Field of knowledge: Health Sciences - Dentistry - Periodontology
• Principal Investigator: Carlos Rossa Junior
• Grantee: Sabrina Garcia de Aquino
• Host Institution: Faculdade de Odontologia (FOAr). Universidade Estadual Paulista (UNESP). Campus de Araraquara. Araraquara , SP, Brazil

Abstract

Periodontal diseases represent an interesting model to the study of chronic inflammatory conditions, particularly the host-microbial interactions. Tisse destruction associated with progression of periodontal diseases is mostly due to cytokines and other biological mediators produced by the host itself, with fundamental participation of the innate and adaptive immune responses. The expression of these mediators is a strictly-controlled process that may be initiated by of multiple external stimuli acting on various receptors. However, these external signals converge towards a relatively limited number of intracellular signaling pathways. A better comprehension of the role played by diffreent signaling pathways may provide novel and alternative strategies for the modulation of the host response by altering the whole profile of cytokines and mediators produced, as opposed to the strategy of inhibiting or blocking a specific candidate-cytokine, which is limited because of the frequently redundant and compensatory role of inflammatory mediators. The role of Wnt and Notch signaling on the pathogenesis of periodontal diseases is largely unknown. In spite of being initially related with embryonic development and fundamental cellular processes such as control of proliferation and cell death, these pathways have important roles on the regulation of innate and adaptive immunity and bone tissue homeostasis, and have important crosstalk activation with p38 MAPK and NF-kB; which are signaling pathways classically involved with the expression of inflammatory mediators. Our main hypothesis is that the modulation of Wnt and Notch signaling has an important effect on the destruction of non-mineralized and mineralized tissues during periodontal disease progression. As secondary hypotheses, mechanistically these pathways exert this influence on the immune response by crosstalk with p38 MAPK and NF-kB in cells of the innate and adaptive immune response. To test these hypotheses, we propose the following specific aims: 1) To determine the effect of modulating Wnt and Notch signaling on the homeostasis of bone tissue during experimental periodontal diseaseWe will use an established model of LPS-induced experimental periodontal disease in mice, which will be carried out in the presence of Notch and Wnt agonists and inhibitors. The outcomes will be: quantitation of bone resorption by µCT, density of osteoclasts and T cells by immunohistochemistry. Production of the main cytokine regulators of bone homeostasis, RANKL and OPG, will be determined by ELISA, whereas activation of signaling pathways will be studied by Western blot.2) To describe the role of Wnt and Notch signaling on the modulation of innate and adaptive immune response in periodontal diseasesIn vitro, we will use primary T cells and macrophages from mice stimulated with bacterial LPS in the presence of agonists and inhibitors of Wnt and Notch signaling. Polarization of cellular response in macrophages (M1, M2) and T cells (Th1, Th2. Th17, Treg) will be determined by flow cytometry. Using ELISA we will assess activation of AP-1, NF-kB and NFATc1. Multi-ligand ELISA assays and Western blot will be used to describe the profile of Th1,Th2, Th17 cytokines and the activation intracelllular signaling pathways on the gingival tissues harvested from the mice used in the in vivo experiment.