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Physicochemical and biological characterization of PNVCL hydrogels and chitosan nanoparticles containing antimicrobial peptides and flavonoids for application in regenerative endodontic procedures

Grant number: 18/07735-0
Support type:Scholarships in Brazil - Post-Doctorate
Effective date (Start): May 01, 2019
Effective date (End): April 30, 2021
Field of knowledge:Health Sciences - Dentistry
Principal Investigator:Cristiane Duque
Grantee:Karina Sampaio Caiaffa
Home Institution: Faculdade de Odontologia (FOA). Universidade Estadual Paulista (UNESP). Campus de Araçatuba. Araçatuba , SP, Brazil

Abstract

This study has two main objectives: 1) to evaluate in vitro the antimicrobial, anti-inflammatory and inductive effects of cellular differentiation and dentin mineralization of the combination of antimicrobial peptides and flavonoids and 2) to characterize physicochemical and biologically hydrogels or chitosan nanoparticles containing these compounds for endodontic regeneration purposes. First of all, it is intended to evaluate the antimicrobial activity of the antimicrobial peptides LL-37 and its analogue KR-12-a5 in combination with the following flavonoids: taxifolin and EGCG by determining the Fractional Inhibitory Concentration (FIC) on microorganisms of endodontic interest. Then, the best combinations will be selected to determine the effect on single and dual-species biofilms by counting the CFU/mL. In addition, these combinations will also be evaluated on undifferentiated dental pulp cells (hDPC) or dental apical papilla (SCAP - part to be developed abroad) to evaluate cell differentiation and mineralization induction capacity, by determining ALP activity, quantification of mineralization nodules and expression of dentin mineralization markers: DMP-1 and DSPP. The anti-inflammatory activity of the peptide-flavonoid combinations will be assessed by quantification of TNF-± levels in hDPC, by ELISA assays. The combinations with the best results in the previous tests will be incorporated into two types of systems: thermosensitive hydrogels of PNVCL or chitosan nanoparticles. The physico-chemical characterization of the systems will be performed by rheological tests, the release of the compounds and by mass spectroscopy. The biological effect of peptide-flavonoid-containing systems will be evaluated by the antibiofilm action in confocal microscopy, cytotoxicity in macrophages, hDPC and SCAP cultures and gene expression of mineralization markers. Based in the results of this study, we intend to propose polimeric carriers containing natural products with interesting therapeutic properties for future application in endodontic tissue engineering.

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