Effect of calcium hydroxide on LTA-activated apical papila cells differentiation in vitro

• Grant number: 15/09750-8
• Support type: Scholarships in Brazil - Scientific Initiation
• Effective date (Start): July 01, 2015
• Effective date (End): December 31, 2016
• Field of knowledge: Health Sciences - Dentistry - Endodontics
• Principal researcher: Carla Renata Sipert
• Grantee: Juliana Garuba Rahhal
• Home Institution: Faculdade de Odontologia (FO). Universidade de São Paulo (USP). São Paulo , SP, Brazil

Abstract

The treatment of teeth with open apices is one of the greatest challenges of current Endodontics. The study of the effect of intracanal dressings on apical papilla cells has received special attention in this knowledge area over the last years. However, few studies have considered the previous activation of these cells by bacterial byproducts regarding survival and differentiation ability. Therefore, this project aims to investigate the cytotoxicity and osteo/odontogenic differentiation of cultured human apical papilla cells under contact with calcium hydroxide after the previous activation. Primary cultures of apical papilla cells (n = 3) will be established based on an explant technique. Part of the cells will be primed with 1 µg/mL of Enterococcus faecalis lipoteichoic acid (LTA) for seven days. The cells will be distributed at 96-wells or 24-wells plates. The evaluation of the cytotoxicity of increasing concentrations of calcium hydroxide will be assessed by means of MTT method in order to select the highest concentration unable to affect cellular viability for five days. Primed or naïve cells will be kept in contact with the selected concentration of calcium hydroxide for five days when the cells will be subjected to odontogenic differentiation for 14 and 28 days. The cellular differentiation will be assessed by calcium deposition by using alizarin red S staining and the gene expression of dentin matrix protein-1, dentin sialophosphoprotein, and osteocalcin by means of reverse transcription followed by quantitative polymerase chain reaction (RT-qPCR). Data will be analyzed by using one-way analysis of variance setting p < 0.05.