Bioactive potential of calcium-containing nanofiber scaffolds enriched with fibronectin on human pulp cells

• Grant number: 19/11192-4
• Support type: Scholarships in Brazil - Master
• Effective date (Start): September 01, 2019
• Effective date (End): February 28, 2021
• Field of knowledge: Health Sciences - Dentistry - Dental Materials
• Principal researcher: Josimeri Hebling Costa
• Grantee: Caroline Anselmi de Oliveira
• Home Institution: Faculdade de Odontologia (FOAr). Universidade Estadual Paulista (UNESP). Campus de Araraquara. Araraquara , SP, Brazil

Abstract

The pulp tissue of human teeth is a rich source of undifferentiated mesenchymal stem cells and, therefore, has a great reparative potential. Despite that potential, the direct pulp capping materials currently used to promote tissue repair, mostly based in calcium hydroxide, cause superficial necrosis of the pulp, intense cell death and inflammation. This mechanism of action does not respect the biocompatibility and bioactivity principles and, therefore, the development of new materials to apply in direct contact with the pulp is still necessary. Tissue engineering has made possible the development of new treatments based on three main pillars, a cell source, scaffolds and signalizing molecules. Transposing that triad to the pulp biology, undifferentiated mesenchymal stem cells residing in the tissue would be attracted by chemotaxis to a nanofibrillar scaffold and then induced to differentiate into odontoblast-like cells. Therefore, the aim of this study is to synthetize and characterize polycaprolactone (PCL) nanofiber scaffolds incorporated with low concentrations of a mineral phase, as well as, to evaluate the scaffolds bioactivity on human dental pulp cells (HDPCs) when impregnated with fibronectin. In the phase 1, different concentrations of calcium hydroxide (0; 0.1%; 0.2% or 0.4%) will be incorporated into PCL scaffolds produced by the electrospinning technique. The scaffolds will be characterized with regards to the morphology (SEM) and chemical composition (EDS), and analyzed as to the hydrolytic degradation, pH change of the medium and calcium release. Additionally, HDPCs will be placed in contact with the scaffolds for the analysis of the cell viability (alamarBlue), proliferation (Live/Dead assay), and adhesion and spread (F-actine). Based on the cellular response, the best calcium hydroxide concentration will be selected to the next phase. Therefore, in the phase 2, scaffolds incorporated with the best calcium hydroxide concentration will be impregnated with fibronectin (20 µg/mL) and will be evaluated with regards to HDPCs migration (Trans-well), viability (alamarBlue), proliferation (Live/Dead assay), alkaline phosphatase activity, mineralized nodules formation (Alizarin red), and gene expression of odontogenic differentiation markers (RT-PCR). Sample size will vary according to the protocol. The dataset of each response variable will be analyzed for the adherence to the normal distribution and homoscedasticity among groups. Whether the dataset satisfied these prerequisites or not will be used to select the statistical tests. The level of significance for statistical decisions will be set as 5%. (AU)