Purification of pancreatic endocrine subsets reveals increased iron metabolism in beta cells.,Molecular Metabolism

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Purification of pancreatic endocrine subsets reveals increased iron metabolism in beta cells.
Molecular Metabolism ( IF 7.0 ) Pub Date : 2020-08-05 , DOI: 10.1016/j.molmet.2020.101060
C Berthault ₁ , W Staels ₂ , R Scharfmann ₁

Affiliation

Objectives

The main endocrine cell types in pancreatic islets are alpha, beta, and delta cells. Although these cell types have distinct roles in the regulation of glucose homeostasis, inadequate purification methods preclude the study of cell type-specific effects. We developed a reliable approach that enables simultaneous sorting of live alpha, beta, and delta cells from mouse islets for downstream analyses.

Methods

We developed an antibody panel against cell surface antigens to enable isolation of highly purified endocrine subsets from mouse islets based on the specific differential expression of CD71 on beta cells and CD24 on delta cells. We rigorously demonstrated the reliability and validity of our approach using bulk and single cell qPCR, immunocytochemistry, reporter mice, and transcriptomics.

Results

Pancreatic alpha, beta, and delta cells can be separated based on beta cell-specific CD71 surface expression and high expression of CD24 on delta cells. We applied our new sorting strategy to demonstrate that CD71, which is the transferrin receptor mediating the uptake of transferrin-bound iron, is upregulated in beta cells during early postnatal weeks. We found that beta cells express higher levels of several other genes implicated in iron metabolism and iron deprivation significantly impaired beta cell function. In human beta cells, CD71 is similarly required for iron uptake and CD71 surface expression is regulated in a glucose-dependent manner.

Conclusions

This study provides a novel and efficient purification method for murine alpha, beta, and delta cells, identifies for the first time CD71 as a postnatal beta cell-specific marker, and demonstrates a central role of iron metabolism in beta cell function.

中文翻译：

胰腺内分泌亚群的纯化揭示了 β 细胞中铁代谢的增加。

目标

胰岛中的主要内分泌细胞类型是 α、β 和 δ 细胞。尽管这些细胞类型在葡萄糖稳态的调节中具有不同的作用，但不充分的纯化方法妨碍了对细胞类型特异性影响的研究。我们开发了一种可靠的方法，可以同时分选来自小鼠胰岛的活 alpha、beta 和 delta 细胞，用于下游分析。

方法

我们开发了一种针对细胞表面抗原的抗体组合，能够根据 β 细胞上 CD71 和 delta 细胞上 CD24 的特异性差异表达，从小鼠胰岛中分离出高度纯化的内分泌亚群。我们使用批量和单细胞 qPCR、免疫细胞化学、报告小鼠和转录组学严格证明了我们方法的可靠性和有效性。

结果

胰腺α、β 和δ 细胞可以根据β 细胞特异性CD71 表面表达和δ 细胞上CD24 的高表达进行分离。我们应用我们的新分类策略来证明 CD71（介导转铁蛋白结合铁摄取的转铁蛋白受体）在出生后早期在 β 细胞中上调。我们发现 β 细胞表达更高水平的其他几种与铁代谢和铁缺乏有关的基因，这会显着损害 β 细胞功能。在人类 β 细胞中，铁摄取同样需要 CD71，并且 CD71 表面表达以葡萄糖依赖性方式进行调节。