FoxO1 mediates odontoblast differentiation of hDPSCs via B cell-derived ANGPTL1 in dental caries: A laboratory investigation

IF 5.4 1区医学 Q1 DENTISTRY, ORAL SURGERY & MEDICINE

International endodontic journal Pub Date : 2025-02-04 DOI:10.1111/iej.14206

Peimeng Zhan, Zhu Huang, Zhuo Xie, Xinfang Zhang, Zongshan Shen, Lingling Chen, Shuheng Huang, Qiting Huang, Zhengmei Lin, Runfu Wang

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引用次数: 0

Abstract

Aim

Clinical and in vitro evidence indicates that chronic inflammatory responses initiated by dental caries can persist in the dental pulp even after treatment, necessitating the formation of reparative dentin to restore tissue homeostasis and health. Human dental pulp stem cells (hDPSCs) serve as crucial precursors in this reparative process. This study explores the role of B cells and their secreted factor, Angiopoietin Like 1 (ANGPTL1), in promoting hDPSCs differentiation into odontoblasts under carious conditions, with a particular focus on the activation of Forkhead box O1 (FoxO1).

Methodology

Single-cell RNA sequencing (scRNA-Seq) data from the GEO database were analysed to explore cellular interactions and molecular mechanisms in dental pulp. Immunofluorescence staining was used to investigate the expression patterns of B cells or hDPSCs in dental pulp and hydroxyapatite/tricalcium phosphate (HA/TCP) scaffolds. The expression levels of ANGPTL1 were quantified using enzyme-linked immunosorbent assay (ELISA). Odontoblast differentiation capacity was assessed by alkaline phosphatase activity, alizarin red S staining, and western blotting analysis. hDPSCs were overexpressed or knocked down FoxO1 with lentiviruses. The regulatory interaction between FoxO1 and the DSPP promoter was evaluated through dual-luciferase reporter assay and chromatin immunoprecipitation assay. Statistical analyses were conducted using Student's t-test or one-way analysis of variance (anova) with a p-value of <.05 considered statistically significant.

Results

scRNA-Seq data indicated a significant increase in B cells and ANGPTL1 expression in carious dental pulp. Functional analyses confirmed that ANGPTL1 secreted by B cells activated FoxO1 expression in hDPSCs, enhancing their differentiation into odontoblast-like cells. Blocking ANGPTL1 signalling with a specific antibody reduced FoxO1 expression, indicating a regulatory link between ANGPTL1 and FoxO1. Overexpression of FoxO1 in hDPSCs promoted their differentiation into odontoblasts and facilitated mineralized matrix formation. Mechanistic studies revealed that FoxO1 directly binds to the DSPP promoter, thereby inducing its expression.

Conclusions

Our study reveals a novel mechanism in which ANGPTL1 secreted by B cells in a carious environment promotes the odontoblast differentiation of hDPSCs by upregulating FoxO1. This finding highlights a potential therapeutic target for enhancing dental pulp repair and regeneration.

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FoxO1 通过 B 细胞衍生的 ANGPTL1 在龋齿中介导 hDPSCs 的牙本质分化：实验室研究

目的：临床和体外证据表明，龋齿引起的慢性炎症反应即使在治疗后也会在牙髓中持续存在，需要形成修复性牙本质来恢复组织的稳态和健康。人牙髓干细胞（hDPSCs）在这一修复过程中起着至关重要的前体作用。本研究探讨了B细胞及其分泌因子血管生成素样1 （ANGPTL1）在促进龋病条件下hdpsc向成牙细胞分化中的作用，特别关注叉头盒O1 （FoxO1）的激活。方法：对GEO数据库中的单细胞RNA测序（scRNA-Seq）数据进行分析，以探索牙髓中的细胞相互作用和分子机制。采用免疫荧光染色法观察B细胞或hDPSCs在牙髓和羟基磷灰石/磷酸三钙（HA/TCP）支架中的表达规律。采用酶联免疫吸附法（ELISA）定量测定ANGPTL1的表达水平。通过碱性磷酸酶活性、茜素红S染色和western blotting分析评估成牙髓细胞分化能力。hdpsc被慢病毒过度表达或敲低fox01。通过双荧光素酶报告基因法和染色质免疫沉淀法评估FoxO1与DSPP启动子之间的调控相互作用。采用Student’st检验或单因素方差分析（anova）进行统计学分析，p值为：结果：scRNA-Seq数据显示龋牙髓中B细胞和ANGPTL1表达显著升高。功能分析证实，B细胞分泌的ANGPTL1激活了hDPSCs中FoxO1的表达，促进其向成牙细胞样细胞的分化。用特异性抗体阻断ANGPTL1信号传导可降低FoxO1的表达，表明ANGPTL1与FoxO1之间存在调控联系。FoxO1在hDPSCs中的过表达促进了它们向成牙髓细胞的分化，促进了矿化基质的形成。机制研究表明fox01直接与DSPP启动子结合，从而诱导其表达。结论：我们的研究揭示了一种新的机制，即在龋病环境下B细胞分泌的ANGPTL1通过上调FoxO1促进hdpsc成牙细胞分化。这一发现突出了促进牙髓修复和再生的潜在治疗靶点。

本文章由计算机程序翻译，如有差异，请以英文原文为准。

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来源期刊

International endodontic journal 医学-牙科与口腔外科

CiteScore

10.20

自引率

28.00%

发文量

195

审稿时长

4-8 weeks

期刊介绍： The International Endodontic Journal is published monthly and strives to publish original articles of the highest quality to disseminate scientific and clinical knowledge; all manuscripts are subjected to peer review. Original scientific articles are published in the areas of biomedical science, applied materials science, bioengineering, epidemiology and social science relevant to endodontic disease and its management, and to the restoration of root-treated teeth. In addition, review articles, reports of clinical cases, book reviews, summaries and abstracts of scientific meetings and news items are accepted. The International Endodontic Journal is essential reading for general dental practitioners, specialist endodontists, research, scientists and dental teachers.