Aims/hypothesis

Pancreatic beta cell mass is dynamically regulated in response to increased physiological and pathological demands. Understanding the mechanisms that control physiological beta cell proliferation could provide valuable insights into novel therapeutic approaches to diabetes. Here, we aimed to analyse the intracellular and extracellular signalling pathways involved in regulating the physiological proliferation of beta cells using single-cell RNA-seq (scRNA-seq) and in vitro functional assays.

Methods

Islets isolated from nulliparous mice, mice at different time points of gestation and mice at day 4 after delivery were analysed using scRNA-seq. Bioinformatics analyses of scRNA-seq data were performed to determine the heterogeneous transcriptomic characteristics of beta cells and to identify the proliferating subpopulation. CellChat was used to analyse cell–cell communication and identify the ligand–receptor pairs between beta cell subclusters as well as between non-beta cells and proliferating beta cells. In vitro functional assays were conducted in mouse and rat beta cell lines and isolated mouse primary islets to validate the role of Kmt5a– mono-methylation of histone H4 at lysine 20 (H4K20me) signalling and endothelial-derived heparin-binding EGF-like growth factor (HBEGF) in beta cell proliferation.

Results

Of 43,724 endocrine and non-endocrine cells within islets analysed by scRNA-seq, 15,569 beta cells were clustered into eight distinct populations, each exhibiting unique heterogeneity. A proliferating beta cell subcluster was identified that highly expressed the histone methyltransferase Kmt5a. Activation of Kmt5a–H4K20me signalling upregulated the expression of Cdk1 and promoted beta cell proliferation. The crosstalk between endothelial cells and the proliferating beta cell subcluster, mediated by the HBEGF–EGF receptor (EGFR) ligand–receptor interaction, increased as beta cell mass expanded. HBEGF increased the expression levels of genes involved in the cell cycle and promoted beta cell proliferation by regulating the Kmt5a–H4K20me signalling pathway.

Conclusions/interpretation

Our study demonstrates that, under physiological conditions, endothelial-derived HBEGF regulates beta cell proliferation through the Kmt5a–H4K20me signalling pathway, which may serve as a potential target to promote beta cell expansion and treat diabetes.

Data availability

The scRNA-seq and RNA-seq datasets are available from the Gene Expression Omnibus (GEO) using the accession numbers GSE278860 and GSE278861, respectively.

Graphical Abstract

中文翻译：

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单细胞 RNA 测序确定内皮来源的 HBEGF 通过 EGFR-Kmt5a-H4K20me 通路促进小鼠胰腺 β 细胞增殖

 目标/假设

胰腺 β 细胞质量受到动态调节，以响应增加的生理和病理需求。了解控制生理性 β 细胞增殖的机制可以为糖尿病的新治疗方法提供有价值的见解。在这里，我们旨在使用单细胞 RNA-seq （scRNA-seq） 和体外功能测定分析参与调节 β 细胞生理增殖的细胞内和细胞外信号通路。

 方法

使用 scRNA-seq 分析从未产小鼠、不同妊娠时间点的小鼠和分娩后第 4 天的小鼠中分离的胰岛。对 scRNA-seq 数据进行生物信息学分析，以确定 β 细胞的异质性转录组学特征并鉴定增殖亚群。CellChat 用于分析细胞间通讯并识别 β 细胞亚簇之间以及非 β 细胞和增殖 β 细胞之间的配体-受体对。在小鼠和大鼠 β 细胞系以及分离的小鼠原代胰岛中进行体外功能测定，以验证 Kmt5a – 组蛋白 H4 在赖氨酸 20 （H4K20me） 信号传导和内皮衍生的肝素结合 EGF 样生长因子 （HBEGF） 在 β 细胞增殖中的作用。

 结果

在通过 scRNA-seq 分析的胰岛内的 43,724 个内分泌和非内分泌细胞中，15,569 个 β 细胞聚成八个不同的群体，每个细胞群都表现出独特的异质性。鉴定出一个高度表达组蛋白甲基转移酶 Kmt5a 的增殖 β 细胞亚簇。Kmt5a-H4K20me 信号的激活上调了 Cdk1 的表达并促进了 β 细胞增殖。由 HBEGF-EGF 受体 （EGFR） 配体-受体相互作用介导的内皮细胞和增殖的 β 细胞亚簇之间的串扰随着 β 细胞质量的扩大而增加。HBEGF 通过调节 Kmt5a-H4K20me 信号通路提高细胞周期相关基因的表达水平并促进 β 细胞增殖。

结论/解释

我们的研究表明，在生理条件下，内皮来源的 HBEGF 通过 Kmt5a-H4K20me 信号通路调节 β 细胞增殖，这可能是促进 β 细胞扩增和治疗糖尿病的潜在靶点。

 数据可用性

scRNA-seq 和 RNA-seq 数据集可从基因表达综合 （GEO） 中分别使用GSE278860和 GSE278861 的登录号获得。

 图形摘要