Distinct Cellular Origins for Serotonin-Expressing and Enterochromaffin-like Cells in the Gastric Corpus

The alimentary tract contains a diffuse endocrine system comprising enteroendocrine cells that secrete peptides or biogenic amines to regulate digestion, insulin secretion, food intake, and energy homeostasis. Lineage analysis in the stomach revealed that a significant fraction of endocrine cells in the gastric corpus did not arise from neurogenin3-expressing cells, unlike enteroendocrine cells elsewhere in the digestive tract. We aimed to isolate enriched serotonin-secreting and enterochromaffin-like (ECL) cells from the stomach and to clarify their cellular origin. We used Neurod1 and Neurog3 lineage analysis, and examined differentiation of serotonin-producing and ECL cells in stomach tissues of Neurod1-cre;ROSAtdTom, Tph1-CFP, c-Kitwsh/wsh, and Neurog3Cre;ROSAtdTom mice, by immunohistochemistry. We used fluorescence-activated cell sorting to isolate each cell type for gene expression analysis. We performed RNA-seq analysis of ECL cells. Neither serotonin-secreting nor ECL cells of the corpus arose from cells expressing Neurod1. Serotonin-secreting cells expressed a number of mast cell genes, but not genes associated with endocrine differentiation; they did not develop in c-Kitwsh/wsh mice and were labeled with transplanted bone marrow cells. RNA-seq analysis of ECL cells revealed high expression levels of many genes common to endocrine cells including transcription factors, hormones, ion channels, and solute transporters but not markers of bone marrow cells. We used fluorescence-activated cell sorting to isolate Hdc+ cells from stomach corpus and performed RNA-seq for gene expression analysis to determine the origin of those cells.消化道拥有一套弥散内分泌系统（diffuse endocrine system），该系统由肠内分泌细胞（enteroendocrine cells）构成，此类细胞可分泌肽类或生物胺，参与调控消化过程、胰岛素分泌、食物摄入及能量稳态。针对胃部的谱系分析（lineage analysis）结果显示，与消化道其余部位的肠内分泌细胞不同，胃体（gastric corpus）内有相当比例的内分泌细胞并非起源于表达神经元素3（Neurogenin3）的细胞。本研究旨在从胃部分离富集5-羟色胺分泌细胞与肠嗜铬样（ECL）细胞，并阐明其细胞起源。我们采用神经分化因子1（Neurod1）及神经元素3（Neurog3）谱系分析策略，通过免疫组织化学（immunohistochemistry）方法，检测了Neurod1-cre;ROSAtdTom、Tph1-CFP、c-Kitwsh/wsh及Neurog3Cre;ROSAtdTom小鼠胃部组织中5-羟色胺产生细胞与ECL细胞的分化状态。我们借助荧光激活细胞分选术（fluorescence-activated cell sorting）分离各类细胞以开展基因表达分析，并对ECL细胞进行了RNA测序（RNA-seq）分析。研究发现，胃体中的5-羟色胺分泌细胞与ECL细胞均未起源于表达神经分化因子1的细胞。5-羟色胺分泌细胞可表达多种肥大细胞基因（mast cell genes），但不表达与内分泌分化（endocrine differentiation）相关的基因；此类细胞无法在c-Kitwsh/wsh小鼠体内发育，且可被移植的骨髓细胞（bone marrow cells）标记。对ECL细胞的RNA测序分析结果表明，其高表达多种内分泌细胞共有的基因，包括转录因子（transcription factors）、激素、离子通道（ion channels）及溶质转运蛋白（solute transporters），但不表达骨髓细胞标志物。我们还通过荧光激活细胞分选术从胃体中分离出Hdc+细胞（Hdc+ cells），并进行RNA测序以开展基因表达分析，以此确定此类细胞的起源。