Labeling and Characterization of Human GLP-1-Secreting L-cells in Primary Ileal Organoid Culture
Labeling and Characterization of Human GLP-1-Secreting L-cells in Primary Ileal Organoid Culture
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DOI：
10.1016/j.celrep.2020.107833
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发表时间：
2020-06-30
期刊：
影响因子：
8.8
通讯作者：
Gribble, Fiona M.
中科院分区：
文献类型：
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作者：
Goldspink, Deborah A.;Lu, Van B.;Gribble, Fiona M.
胰高血糖素样肽-1（GLP-1）来自肠道L细胞，刺激胰岛素分泌，减少食物摄入后的食欲，是治疗2型糖尿病和肥胖症药物的基础。靶向L-和其他肠内分泌细胞的药物正在开发中，目的是模拟胃旁路手术的内分泌效应，但如果没有人类L-细胞模型，它们很难开发。通过CRISPR-Cas9工程化的人类回肠类器官在胰高血糖素原基因座中表达荧光蛋白Venus，从而能够在培养物中维持活的、可识别的人类L细胞。通过RNA测序（RNA-seq）分析的荧光激活细胞分选（FACS）纯化的类器官来源的L细胞表达激素、受体和离子通道，这在很大程度上是其鼠对应物的典型特征。L-细胞是电活性的，并且响应于G-蛋白偶联受体配体而表现出膜去极化和钙升高。类器官响应于葡萄糖和其他刺激而分泌激素。在类器官培养物中标记和维持人L细胞的能力为探索L细胞功能和开发靶向人肠内分泌系统的药物开辟了途径。
Glucagon-like peptide-1 (GLP-1) from intestinal L-cells stimulates insulin secretion and reduces appetite after food ingestion, and it is the basis for drugs against type-2 diabetes and obesity. Drugs targeting L- and other enteroendocrine cells are under development, with the aim to mimic endocrine effects of gastric bypass surgery, but they are difficult to develop without human L-cell models. Human ileal organoids, engineered by CRISPR-Cas9, express the fluorescent protein Venus in the proglucagon locus, enabling maintenance of live, identifiable human L-cells in culture. Fluorescence-activated cell sorting (FACS)-purified organoid-derived L-cells, analyzed by RNA sequencing (RNA-seq), express hormones, receptors, and ion channels, largely typical of their murine counterparts. L-cells are electrically active and exhibit membrane depolarization and calcium elevations in response to G-protein-coupled receptor ligands. Organoids secrete hormones in response to glucose and other stimuli. The ability to label and maintain human L-cells in organoid culture opens avenues to explore L-cell function and develop drugs targeting the human enteroendocrine system.