Acquired pellicle engineering with a new cystatin derived from Maqui Berry (Aristotelia chilensis - MaquiCPI-3): defining concentrations for the prevention of enamel and dentin erosion in vitro

• Grant number: 21/11152-2
• Support Opportunities: Scholarships in Brazil - Scientific Initiation
• Effective date (Start): March 01, 2022
• Effective date (End): February 28, 2023
• Field of knowledge: Health Sciences - Dentistry - Social and Preventive Dentistry
• Principal Investigator: Marília Afonso Rabelo Buzalaf
• Grantee: Cecília Guimarães Giannico Borges
• Host Institution: Faculdade de Odontologia de Bauru (FOB). Universidade de São Paulo (USP). Bauru , SP, Brazil

Abstract

The incorporation of proteins (eg CaneCPI-5) into the acquired pellicle (AP) has favored its ability to protect against erosion. In this context, a new cysteine protease inhibitor protein (cystatin) derived from Maqui Berry (MaquiCPI-3) deserves to be studied, due to some characteristics similar to CaneCPI-5 and its high yield when heterologously expressed in bacteria. Initial in vitro results with MaquiCPI-3 might open a new preventive possibility against tooth erosion, aiming for its incorporation in dental products to increase the acid-resistant capacity of AP (AP Engineering). Thus, the aim of this study will be to analyze the protective effect and define concentrations of MaquiCPI-3 by modifying AP protection against initial erosion of enamel and dentin in vitro. 238 samples will be prepared, 119 bovine enamel and 119 bovine dentin (4 X 4 mm). Afterwards, the samples will be randomized and divided into 7 groups, according to the treatments (n/group=17): 1) Deionized water (negative control); 2) 0.1 mg/ml CaneCPI-5 (positive control); 3) Commercial solution with SnCl2/NaF/AmF (800 ppm Sn+2,500 ppm F-, pH 4.5, Erosion Protection® - GABA, (positive control)); 4) 0.1 mg/ml MaquiCPI-3; 5) 0.5 mg/ml MaquiCPI-3; 6) 1.0 mg/ml MaquiCPI-3 and 7) 2.0 mg/ml MaquiCPI-3. Initially, the samples will be treated individually (250 µL) with the respective solutions for 2h, 37ºC, under constant agitation. Then, the AP will be individually formed (250 µL) for 2h, 30ºC, under constant agitation (the stimulated saliva will be collected from 3 volunteers). Subsequently, the erosive challenge will be done individually (1 mL) with 0.65% citric acid (pH = 3.4) for 1 min, at 37ºC, under constant agitation. These procedures will be performed once a day, for 3 consecutive days. Surface microhardness (SHM) analyses will be performed and changes in SMH (baseline SHM - post-erosion SMH) will be calculated. Data will be analyzed using the GraphPad Prism software. First, the data will be checked for normality (Kolmogorov-Smirnov test) and homogeneity (Bartlett test), to select the appropriate statistical test. The significance level will be 5%.(AU)