Influence of the NF-8B pathway on the effects of chalcone T4 in vitro osteoclastogenesis

• Grant number: 19/15546-5
• Support Opportunities: Scholarships in Brazil - Scientific Initiation
• Effective date (Start): February 01, 2020
• Effective date (End): July 31, 2022
• Field of knowledge: Health Sciences - Dentistry - Periodontology
• Principal Investigator: Morgana Rodrigues Guimarães Stabili
• Grantee: Giovani Cirelli
• Host Institution: Faculdade de Odontologia (FOAr). Universidade Estadual Paulista (UNESP). Campus de Araraquara. Araraquara , SP, Brazil

Abstract

Chalcone derivatives are polyphenolic structures found in medicinal plants, with numerous biological activities. These compounds have a simple molecular architecture, allowing structural modifications that can alter their physicochemical properties and significantly increase the potency and duration of their biological effects. In fact, several synthetic chalcone compounds have demonstrated effectiveness in the clinical treatment of inflammatory diseases and bone pathologies, such as osteoporosis and rheumatoid arthritis. Inhibition of the production of pro-inflammatory cytokines, regulation of intracellular signaling pathways, and modulation of the activity of transcription factors appears to be at least part of the mechanisms used by chalcones to exert their anti-inflammatory activities. Considering the biological effects of these compounds, our research group evaluated the ability of a new chalcone derivative (Chalcone T4) to suppress the expression of relevant mediators in inflammation. Chalcone T4 was able to reduce the gene expression of inflammatory markers (IL-6, Tnf-±, Mmp-13) by more than 90% in macrophages, even when used in low concentrations. These results, although preliminary, suggest that Chalcone T4 can act by inhibiting osteoclastogenesis and may be efficient in the treatment of osteolytic diseases. It is even possible, considering the critical role of NF-8B on the expression of pro-inflammatory genes, as well as on the activity and differentiation of osteoclasts, that the possible inhibitory effect of chalcone on osteoclastogenesis involves the modulation of this transcription factor. In order to investigate such hypotheses, this study has the following specific objectives: Specific objective 1: To investigate the effects of chalcone T4 on the expression of bone resorption markers, differentiation and activity of osteoclast precursor cells (RAW 264.7) stimulated with RANKL. The expression of certain gene markers of osteoclastogenesis will be observed compared by RT-qPCR and the number of differentiated osteoclasts, identified by fluorescence microscopy visualization actin ring containing three or more nuclei. The osteoclastic activity will be determined by the area of resorption in plates containing inorganic material. Specific objective 2: To evaluate the influence of the NF-8B signaling pathway on the effects of chalcone on osteoclast differentiation and activity. RAW 264.7 will be treated with different non cytotoxic concentrations of chalcone and/or the inhibitor of NF-8B (BAY 11-7082), and then stimulated with RANKL and used in the experiments described above (gene expression by RT-qPCR, count of osteoclasts identified by fluorescence microscopy and osteoclast activity determined by measurement of the area of resorption).