Upregulated nicotinic ACh receptor signaling contributes to intestinal stem cell function through activation of Hippo and Notch signaling pathways
Upregulated nicotinic ACh receptor signaling contributes to intestinal stem cell function through activation of Hippo and Notch signaling pathways
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烟碱 ACh 受体信号上调通过激活 Hippo 和 Notch 信号通路促进肠道干细胞功能
DOI：
10.1016/j.intimp.2020.106984
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发表时间：
2020-11-01
影响因子：
5.6
通讯作者：
Osawa, Masatake
中科院分区：
文献类型：
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作者：
Takahashi, Toshio;Shiraishi, Akira;Osawa, Masatake
背景：近年来的研究表明，多种哺乳动物非神经元细胞可原位合成乙酰胆碱（ACh），并通过烟碱和毒蕈碱ACh受体（nAChRs和mAChRs）调控胆碱能信号传导。了解通过激活nAChR信号控制肠干细胞（ISC）功能的机制对于开发炎症性肠病（IBD）等疾病的治疗干预措施至关重要。先前，通过使用隐绒毛类器官培养物进行RNA测序（RNA- seq）分析，我们发现Hippo信号通路，一个干细胞调节网络，在尼古丁治疗后在ISCs中上调。在这里，我们通过激活Hippo信号通路来探索nAChR信号的作用。方法：采用定量反转录聚合酶链反应（qRT-PCR）对RNA-Seq数据进行验证。产生了β 4敲入小鼠，并使用敲入小鼠及其肠道类器官进行了实验。结果：RNA-Seq和qRT-PCR分析显示，尼古丁处理后YAP1/TAZ和Notch1/Dll1的表达上调。然而，一种nAChR拮抗剂，甲胺，强烈抑制这些基因的表达。值得注意的是，我们发现在β 4敲入小鼠小肠中，YAP1和Notch1的表达显著降低，而TAZ和Dll1的表达不明显，这表明Hippo和Notch信号通路可能是nAChR信号传导的靶点。此外，在隐窝底部与ISCs相互作用的Paneth细胞中检测到荧光信号，表明Paneth细胞通过激活Hippo和Notch信号通路，通过nAChR信号通路与ISCs相互作用。结论：我们的研究结果表明，nAChR信号的上调有助于维持ISC活性，并通过激活Hippo和Notch信号通路来平衡分化。
Backgrounds: Recent studies have shown that various mammalian non-neuronal cells synthesize acetylcholine (ACh) in situ and operate cholinergic signaling via nicotinic and muscarinic ACh receptors (nAChRs and mAChRs). Understanding the mechanisms that control intestinal stem cell (ISC) function through activation of nAChR signaling is critical for developing therapeutic interventions for diseases such as inflammatory bowel disease (IBD). Previously, by conducting RNA sequencing (RNA-Seq) analysis using crypt-villus organoid cultures, we found that the Hippo signaling pathway, a stem cell regulating network, is upregulated in ISCs after treatment with nicotine. Here, we explored the roles of nAChR signaling through activation of the Hippo signaling pathway.Methods: RNA-Seq data were validated by quantitative reverse transcription polymerase chain reaction (qRT-PCR) analysis. beta 4-knock-in mice were generated, and experiments using the knock-in mice and their intestinal organoids were carried out.Results: RNA-Seq and qRT-PCR analyses demonstrated that the expression of YAP1/TAZ and Notch1/Dll1 was upregulated after treatment with nicotine. However, a nAChR antagonist, mecamylamine, strongly inhibited the expression of these genes. Notably, we found that in beta 4-knock-in mouse small intestines, expression of YAP1 and Notch1 was significantly reduced, but not that of TAZ and Dll1, suggesting that Hippo and Notch signaling pathways are putative targets for nAChR signaling. Furthermore, fluorescent signals were detected in Paneth cells that interact with ISCs at the crypt bottom, indicating an interaction between Paneth cells and ISCs via nAChR signaling through the activation of Hippo and Notch signaling pathways.Conclusion: Our results indicate that upregulated nAChR signaling contributes to the maintenance of ISC activity and balances differentiation through activation of Hippo and Notch signaling pathways.