Effect of a combination of proteins/peptide on the acquired pellicle engineering, including a new cystatin derived from Maqui Berry (MaquiCPI-3), Stn15pSpS, and Hemoglobin, associated with EGCG, for protection against dental erosion in vitro

• Grant number: 24/22199-8
• Support Opportunities: Scholarships in Brazil - Scientific Initiation
• Start date: March 01, 2025
• End date: February 28, 2026
• Field of knowledge: Health Sciences - Dentistry - Social and Preventive Dentistry
• Principal Investigator: Talita Mendes Oliveira Ventura
• Grantee: Felipe José Costa Corrêia
• Host Institution: Faculdade de Odontologia de Bauru (FOB). Universidade de São Paulo (USP). Bauru , SP, Brazil
• Associated research grant: 19/26070-1 - Modulation of acquired enamel pellicle to control dental mineral loss: unravelling mechanisms to make therapies possible, AP.TEM

Abstract

The acquired enamel pellicle (AEP) is an organic layer formed in vivo by the selective adsorption of salivary proteins on the dental surface, also containing glycoproteins and lipids. This bacteria-free integument acts as a protective interface that plays a crucial role in the interfacial events of the oral cavity. Moreover, the AEP is considered one of the main protective factors against erosive tooth wear (ETW). Among the proteins present in the AEP with the potential to bind to enamel and confer acid resistance, cystatin, statherin, and hemoglobin have recently been identified as notable examples. Modifying the protein profile of the AEP through the incorporation of specific proteins has shown promise in controlling ETW. However, studies exploring the interaction of AEP with other bioactive components, such as epigallocatechin gallate (EGCG), are still scarce, opening new perspectives for investigation. Based on this, the present research project, part of "acquired pellicle engineering," aims to evaluate the protective effect of a new cystatin derived from Maqui Berry (MaquiCPI-3), a statherin-derived peptide (Stn15pSpS), or hemoglobin, isolated or combined, associated or not with EGCG, on the protective impact against initial dental erosion in vitro. Unstimulated saliva will be collected from 10 participants with good general and oral health. For the evaluation of initial dental erosion in vitro, 120 bovine enamel samples (4 x 4 mm) will be prepared. Subsequently, the samples will be randomized and divided into 8 groups according to the previous treatments: 1. Deionized water (negative control); 2. Commercial solution with SnCl2/NaF/AmF (800 ppm Sn + 2,500 ppm F-, pH 4.5, Erosion Protection® - GABA, (positive control)); 3. MaquiCPI-3 0.05 mg/mL; 4. Stn15pSpS 1.88 x 10-5 M; 5. Hemoglobin 1.0 mg/mL; 6. EGCG 1.25 mM; 7. Combination of proteins/peptide; 8. Combination of proteins/peptide associated with EGCG. The AEP will then be formed individually (250 µL of saliva) for 120 minutes at 37 °C under constant agitation. Subsequently, the erosive challenge will be performed individually with 1 mL of 0.65% citric acid (pH = 3.4) for 1 minute at 25 °C under constant agitation. These procedures will be performed once a day for 3 consecutive days. The citric acid will be collected and analyzed by the Arsenazo III colorimetric method. Surface microhardness (SMH) and surface reflection intensity (SRI) analyses will be performed, and changes in SMH (SMH baseline - SMH final) and SRI (%SRI = (SRI final / SRI initial) × 100) will be calculated, respectively. The data will be analyzed using GraphPad Prism software. The data will be checked for normality (Kolmogorov-Smirnov test) and homogeneity (Bartlett's test) for selection of the appropriate statistical test. The significance level will be 5%.