Brizuela Claudia, Chaparro Alejandra, Valencia María Ignacia, Bendek María José, Duncan Henry F, Segura-Egea Juan J, Alhucema Camila, Ramírez Valeria
Universidad de los Andes, Santiago, Chile.
Division of Restorative Dentistry & Periodontology, Dublin Dental University Hospital, Trinity College Dublin, Dublin, Ireland.
Int Endod J. 2025 Aug 15. doi: 10.1111/iej.70017.
To identify differentially expressed proteins (DEPs) in dentinal fluid across a range of pulpal inflammatory stages-mild, moderate, severe pulpitis-using mass spectrometry.
This cross-sectional study analysed dentinal fluid from 60 patients categorised into healthy pulp (control), mild, moderate and severe pulpitis groups, based on Wolters' pulp diagnostic criteria. Pulp conditions were assessed through cold sensibility tests and radiographs, with inclusion limited to patients aged 12-40, who were systemically healthy and exhibited no advanced periodontal or apical pathology. Dentinal fluid was collected under aseptic conditions, stored at -80°C and pooled into 12 representative samples for proteomic analysis. Proteins were extracted using lysis buffers and processed via LC-MS/MS with label-free quantification (LFQ) to identify DEPs. Enrichment analysis and protein-protein interaction (PPI) networks were conducted using Gene Ontology (GO), KEGG databases and STRING, with hub proteins identified using cytoscape. Statistical analysis employed Bayesian t-tests and linear models to evaluate protein expression, with a significance threshold of p < 0.05.
The severe pulpitis group exhibited the highest prevalence of systemic diseases (40%) compared with other groups (6.6%). LC-MS/MS identified 577 proteins, with 62 consistently quantified across the groups. The number of DEPs increased with inflammation severity, with 13 DEPs in severe pulpitis compared with controls. Principal component analysis (PCA) revealed partial separation between control and severe inflammation groups, with significant overlap between mild and moderate stages. Functional enrichment identified key biological pathways, including immune response, energy metabolism and structural integrity. Proteins such as cofilin-1, haemoglobin subunit alpha and peroxiredoxin-1 were upregulated in severe inflammation, while hornerin and myosin light chain 6 were downregulated. These findings highlight proteomic changes associated with pulpitis progression and identify potential prognostic biomarkers and therapeutic targets.
This study identified distinct proteomic differences across pulpitis stages, with unique proteins in severe cases. These findings highlight novel biomarkers for advancing precise, cost-effective, point-of-care diagnostics and therapies, including multiplex platforms or ELISA assays. Proteomic analysis shows promise for understanding disease mechanisms and enabling personalised treatment strategies in endodontics.
运用质谱分析法鉴定在一系列牙髓炎症阶段（轻度、中度、重度牙髓炎）牙本质液中差异表达的蛋白质（DEPs）。
这项横断面研究依据沃尔特牙髓诊断标准，分析了60例患者的牙本质液，这些患者被分为健康牙髓（对照组）、轻度、中度和重度牙髓炎组。通过冷敏感测试和X光片评估牙髓状况，纳入标准仅限于年龄在12至40岁、全身健康且无晚期牙周或根尖病变的患者。在无菌条件下收集牙本质液，储存在-80°C，合并为12个代表性样本进行蛋白质组分析。使用裂解缓冲液提取蛋白质，并通过液相色谱-串联质谱（LC-MS/MS）和无标记定量（LFQ）进行处理，以鉴定DEPs。使用基因本体论（GO）、KEGG数据库和STRING进行富集分析和蛋白质-蛋白质相互作用（PPI）网络分析，使用Cytoscape识别枢纽蛋白。采用贝叶斯t检验和线性模型进行统计分析，以评估蛋白质表达，显著性阈值为p < 0.05。
与其他组（6.6%）相比，重度牙髓炎组全身疾病的患病率最高（40%）。LC-MS/MS鉴定出577种蛋白质，并在各组中持续定量出62种。DEPs的数量随炎症严重程度增加，重度牙髓炎组与对照组相比有13种DEPs。主成分分析（PCA）显示对照组和重度炎症组之间部分分离，轻度和中度阶段之间有显著重叠。功能富集确定了关键生物途径，包括免疫反应、能量代谢和结构完整性。诸如丝切蛋白-1、血红蛋白亚基α和过氧化物酶-1等蛋白质在重度炎症中上调，而角蛋白和肌球蛋白轻链6则下调。这些发现突出了与牙髓炎进展相关的蛋白质组变化，并确定了潜在的预后生物标志物和治疗靶点。
本研究确定了牙髓炎各阶段不同的蛋白质组差异，重度病例中有独特的蛋白质。这些发现突出了新的生物标志物，有助于推进精确、经济高效的即时诊断和治疗，包括多重平台或酶联免疫吸附测定（ELISA）检测。蛋白质组分析有望用于理解疾病机制并实现牙髓病学的个性化治疗策略。
