Polycaprolactone/nanohydroxyapatite nanofibrous scaffolds for the dentin-pulp complex regeneration: synthesis, characterization, and evaluation on human dental pulp cells

Author(s)

Igor Paulino Mendes Soares

• Document type: Master's Dissertation
• Press: Araraquara. 2021-07-01.
• Institution: Universidade Estadual Paulista (Unesp). Faculdade de Odontologia. Araraquara
• Defense date: 2021-06-08
• Advisor: Josimeri Hebling

Abstract

Targeting a cytocompatible and bioactive therapy for the application on the exposed pulp tissue, this study developed and characterized polycaprolactone (PCL) nanofibrous scaffolds incorporated with nano-hydroxyapatite (nHA) and evaluated the response of human dental pulp cells (HDPCs) seeded on their surface. PCL-based solutions (10% w/v in chloroform/dimethylformamide) were incorporated or not with nHA (0.5; 1.0; or 2.0 % w/v) and electrospun into nanofibrous scaffolds. The scaffolds were characterized for morphology (SEM), composition (EDS), solubility (mass change), release of calcium/phosphate (spectrophotometry), and pH medium modulation. HDPCs were cultured on the surface of the scaffolds and evaluated for cell viability (Live/Dead and alamarBlue) and adhesion/spreading (F-actin) over time. The expression of COL1A1, ALPL, DSPP, and DMP1 genes (RT-qPCR), total protein synthesis (TP; Lowry), and alkaline phosphatase activity (ALP; thymolphthalein assay) were investigated at 14 and 21 days. The formation of a mineralized matrix (Alizarin Red) was assessed at 21 days. Data were analyzed with one- or two-way ANOVA complemented with Tukey, Games-Howell, or Sidak post-hocs (α=0.05). All formulations generated fibers ranging from 600 to 900 nm in diameter, with random arrangement. The incorporation of nHA into the nanofibers was dose-dependent. Higher nHA concentrations roughened nanofibers surfaces. PCL+0.5%nHA enlarged fiber diameter whereas PCL+2%nHA increased interfibrillar spaces. PCL+1%nHA or PCL+2%nHA promoted greater release of calcium and phosphate, but the medium pH was maintained below 8.0. HDPCs viability was not affected by the addition of nHA, while adhesion/spreading were favored. The expression of DSPP and DMP1 was upregulated in 14 days, and COL1A1, ALPL, and DMP1 in 21 days by the PCL+1%nHA and PCL+2%nHA formulations. The incorporation of nHA promoted higher protein synthesis and increased ALP activity. The formation of a mineralized matrix was concentration-dependent, about 9× higher for the PCL+2%nHA in comparison to the control. In conclusion, nanofibrous PCL scaffolds incorporated with nHA were cytocompatible, stimulated adhesion, spreading, proliferation, and the odontogenic potential of HDPCs. The PCL+2%nHA formulation is a bioactive tissue engineering strategy for vital pulp therapy. (AU)

• FAPESP's process: 19/07400-0 - Polycaprolactone/nano-hydroxyapatite nanofibrous scaffolds for dentin-pulp complex regeneration: synthesis, characterization, and evaluation on human dental pulp cells
• Grantee: Igor Paulino Mendes Soares
• Support Opportunities: Scholarships in Brazil - Master