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Biological characterization of polydioxanone mesh, functionalized or not with i-PRF, relative to conjunctive tissue healing: in vitro, ex vivo and in vivo study

Grant number: 23/17056-0
Support Opportunities:Regular Research Grants
Duration: May 01, 2024 - April 30, 2026
Field of knowledge:Health Sciences - Dentistry - Periodontology
Principal Investigator:Daniela Bazan Palioto Bulle
Grantee:Daniela Bazan Palioto Bulle
Host Institution: Faculdade de Odontologia de Ribeirão Preto (FORP). Universidade de São Paulo (USP). Ribeirão Preto , SP, Brazil
Associated researchers:Paulo Tambasco de Oliveira

Abstract

The development and improvement of existing biomaterials as mucosal substitutes is required, as it allows alternatives to the use of autogenous grafts. Currently, volume-stable collagen matrices present promising results when compared to subepithelial connective tissue grafts. However, this type of biomaterial is of allogeneic origin (human skin) or xenogeneic origin (porcine derivative). A mucous substitute with a synthetic composition would be an advantageous alternative, as it would combine the benefits of the properties of existing matrices with the advantage of less immunoreactivity and ethical issues as it is not a derivative of animal origin. Polydioxanone (PDO) meshes and filaments appear as promising materials in dermatology and plastic surgery, as collagen stimulators or tissue support. The biological characterization of this material (polydioxanone polymer) will allow us to understand its biological characteristics to subsequently enable subsequent studies investigating its application as a synthetic mucous substitute. Furthermore, 3D scaffolds seem conducive to the incorporation of bioactives in order to optimize cellular behavior by directing the mechanisms involved in clot formation, tissue integration and, consequently, the regeneration process. The objective of this study is, through in vitro, ex vivo and in vivo analyses, to characterize the cellular behavior and formation of the physiological clot of polydioxanone and also to understand whether biofunctionalization with i-PRF could add or modulate the connective tissue relationship to the material. Fibroblast cells (Fb) from the C57BL/6 mouse lineage (SCRC-1008, ATCC, Manassas, VA, USA) will be cultured on polydioxanone (PDO+Fb) directly or polydioxanone functionalized with iPRF (PDO+iPRF+Fb), totaling 5 days of culture, composing the in vitro analyses. In ex vivo analyses, C57BL/6 fibroblasts will be cultured on PDO plus a 16-hour physiological clot produced in the animal (PDO+PhC+Fb) and PDO functionalized with iPRF plus a physiological clot (PDO+PhC+iPRF+Fb). Morphology will be analyzed (scanning electron microscopy - 24h post-plating), cell viability, migration and differentiation (Live/Dead assay, 24h and 5 days post-plating and MTT assay in 5 days). The ELISA assay will be performed to quantify the glycoproteins fibronectin and tenascin (analysis 5 days after cell plating). In the in vivo test, 16 hours after the procedure (implantation of the matrix on the mouse back - PDO group and PDO+iPRF group), the surface morphology produced (SEM) and characterization of the physiological clot formed by flow cytometry will be evaluated. After 7 and 21 days of implantation of PDO and PDO+iPRF on the back of mice, the inflammatory reaction will be characterized (histomorphological analysis of sections stained with HE), and collagen deposition (histomorphological analysis of sections stained with Picrosirius Red). Angiogenesis will be assessed by indirect immunofluorescence, checking the average percentage rate of newly formed vessels, using a specific anti-VEGF-A monoclonal antibody, to assess Vascular Endothelial Growth Factor A (VEGF-A). The data will be evaluated using the homoscedasticity test, parametric or non-parametric tests, establishing the level of significance (p<0.05). The results will be expressed descriptively and in graphs with mean or median and standard deviation (mean ± standard error) or interquartile deviation (median ± interquartile). (AU)

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