Evaluation of biocompatibility of nanofibers scaffolds containing LL-37 and mineralization potential of DPSC cells sensitized by LPS in contact with this scaffolds.

Abstract

Tissue engineering in regenerative endodontics is some steps from the application of intracanal scaffolds in humans with the purpose of forming a new pulp tissue. However the risk of contamination and infection of the root canal system represents a high challenge for its clinical implementation. The purpose of this study is to evaluate in vitro the biocompatibility of scaffolds; and the growth and differentiation of pulp stem cells in contact with these materials, with consequent formation of mineralized tissue, in addition to the neutralization of bacterial lipopolysaccharide (LPS). For this, in this project will be used nanofibers scaffolds of polydioxanone (PDS) fabricated by electrospinning technique, with the addition of LL-37 peptide (peptide which is released by neutrophils. Its is able to neutralize tissue loaded by LPS and induce favorable conditions to the regeneration of the pulp-dentin complex). These scaffolds containing LL-37, will be placed on DPSC cell cultures sensitized by LPS. The biocompatibility of these scaffolds will be examined through cytotoxicity test by WST assay, and through the genotoxicity test by the comet assay. The behavior of cells in contact with these materials will be evaluated through expression of DSPP, DMP1, RUNX2, and ALP genes by real-time PCR (qRT-PCR) and through production of the proteins DSPP and DMP-1, associated with mineralized tissue, by ELISA immunoassay. In addition, functional tests of mineralized tissue (Alkaline Phosphatase Activity and Alizarin Red), and the quantification of residual LPS after contact with scaffolds containing LL-37, will be performed. The results will be analyzed by ANOVA and Tukey's multiple comparison (post hoc), adopting a significance level of 5%. (AU)