Araujo Tamara Teodoro, Debortolli Ana Luiza Bogaz, Carvalho Thamyris Souza, Rodrigues Chelsea Maria Vilas Boas Feitosa, Dionizio Aline, de Souza Beatriz Martines, Vertuan Mariele, Ventura Talita Mendes, Grizzo Larissa Tercilia, Marchetto Reinaldo, Henrique Silva Flavio, Chiaratti Marcos, Santos Angélica Camargo, Alves Lindomar Oliveira, Ferro Milene, Buzalaf Marília Afonso Rabelo
Department of Biological Sciences, Bauru School of Dentistry, University of São Paulo, Bauru 17012-901, Brazil.
Department of Biochemistry and Technological Chemistry, Institute of Chemistry, São Paulo State University, Araraquara 14801-902, Brazil.
Arch Oral Biol. 2025 Mar;171:106159. doi: 10.1016/j.archoralbio.2024.106159. Epub 2024 Dec 5.
This proof-of-concept sequence of in vivo/in vitro studies aimed to unveil the role of acquired enamel pellicle (AEP) engineering with statherin-derived peptide (StN15) on the AEP protein profile, enamel biofilm microbiome in vivo and on enamel demineralization in vitro.
In vivo studies, 10 volunteers, in 2 independent experiments (2 days each), rinsed (10 mL,1 min) with: deionized water (negative control) or 1.88 × 10 M StN15. The AEP, formed along 2 h and the biofilm, along 3 h, were collected. AEP was analyzed by quantitative shotgun-label-free proteomics. The enamel biofilm microbiome was evaluated using 16S-rRNA Next Generation Sequencing (NGS). An in vitro model with microcosm biofilm was employed. Bovine enamel samples (n = 72) were treated with 1) Phosphate-Buffer-Solution (PBS), 2) 0.12 %Chlorhexidine, 3) 500ppmNaF; 4) 1.88 × 10MStN15; 5) 3.76 × 10MStN15 and 6) 7.52 × 10MStN15. Biofilm was supplemented with human saliva and McBain saliva and cultivated for 5 days. Resazurin, colony forming units (CFU) and Transversal Microradiography Analysis-(TMR) were performed.
Proteomic results showed several proteins with acid-resistant, calcium-binding, and antimicrobial properties in the StN15 group. The microbiome corroborated these findings, reducing bacteria that are closely related to dental caries in the StN15 group, compared to the PBS. The microcosm biofilm showed that the lowest concentration of StN15 was the most efficient in reducing bacterial activity, CFU and enamel demineralization compared to PBS.
StN15 can effectively alter the AEP proteome to inhibit initial bacterial colonization, thereby mitigating enamel demineralization. Future research should explore clinical applications and elucidate the mechanisms underlying the protective effects of StN15.
本体内/体外研究的概念验证序列旨在揭示用含组蛋白衍生肽（StN15）的获得性釉质 pellicle（AEP）工程对 AEP 蛋白质谱、体内釉质生物膜微生物群以及体外釉质脱矿的作用。
体内研究中，10 名志愿者在 2 个独立实验（各 2 天）中用以下溶液冲洗（10 mL，1 分钟）：去离子水（阴性对照）或 1.88×10⁻⁶ M StN15。收集 2 小时形成的 AEP 和 3 小时形成的生物膜。通过定量鸟枪法无标记蛋白质组学分析 AEP。使用 16S - rRNA 下一代测序（NGS）评估釉质生物膜微生物群。采用带有微观生物膜的体外模型。牛牙釉质样本（n = 72）用以下溶液处理：1）磷酸盐缓冲溶液（PBS），2）0.12%洗必泰，3）500 ppm 氟化钠；4）1.88×10⁻⁶ M StN15；5）3.76×10⁻⁶ M StN15 和 6）7.52×10⁻⁶ M StN15。生物膜补充人唾液和 McBain 唾液并培养 5 天。进行刃天青、菌落形成单位（CFU）和横向微放射照相分析 -（TMR）。
蛋白质组学结果显示 StN15 组中有几种具有耐酸、钙结合和抗菌特性的蛋白质。微生物群证实了这些发现，与 PBS 相比，StN15 组中与龋齿密切相关的细菌减少。微观生物膜显示，与 PBS 相比，最低浓度的 StN15 在降低细菌活性、CFU 和釉质脱矿方面最有效。
StN15 可有效改变 AEP 蛋白质组以抑制初始细菌定植，从而减轻釉质脱矿。未来的研究应探索临床应用并阐明 StN15 保护作用的潜在机制。
