■Wnt信号是干细胞稳态的关键调节因子,在肠道隐窝和其他后生动物组织中进行了广泛的研究。然而,几乎没有任何关于体内成人肠神经系统(ENS)中Wnt信号传导成分存在的数据。
■因此,我们采用了RNAscopeHiPlex测定法,一种新的和更敏感的原位杂交技术。通过放大目标特定信号,这种技术可以检测低丰度,紧密调节RNA含量,如Wnt信号组件的情况。此外,我们使用数据挖掘方法将我们的数据与先前发表的生理单细胞RNA和基于RiboTag的肠胶质增生的RNA测序分析进行了比较.
■我们的描述性分析表明,Wnt信号通路的多维调控网络的几个组成部分存在于鼠ENS中。Wnt配体Wnnless以及规范(Wnt3a和Wnt2b)和非规范Wnt配体(Wnt5a,Wnt7a,Wnt8b和Wnt11)在粘膜下和肌间神经丛内可检测到。Further,相应的Frizzled受体(Fzd1,Fzd3,Fzd6和Fzd7)和调节信号传导介质,如R-Sondin/DKK配体存在于小肠和大肠的ENS中。Further,数据挖掘方法揭示,一些Wnt相关分子由肠神经胶质细胞簇表达,并在肠神经胶质增生的炎症表现过程中受到动态调节。
■我们的结果表明,规范和非规范的Wnt信号对成熟的ENS及其在健康和炎症中的细胞稳态具有更广泛的影响,比之前预期的要多。
Wnt-signaling is a key regulator of stem cell homeostasis, extensively studied in the intestinal crypt and other metazoan tissues. Yet, there is hardly any data available on the presence of Wnt-signaling components in the adult enteric nervous system (ENS) in vivo.
Therefore, we employed RNAscope HiPlex-assay, a novel and more sensitive in situ hybridization technology. By amplifying target specific signals, this technique enables the detection of low abundance, tightly regulated RNA content as is the case for Wnt-signaling components. Additionally, we compared our data to previously published physiological single cell RNA and RiboTag-based RNA sequencing analyses of enteric gliosis using data-mining approaches.
Our descriptive analysis shows that several components of the multidi-mensional regulatory network of the Wnt-signaling pathway are present in the murine ENS. The transport and secretion protein for Wnt-ligands Wntless as well as canonical (Wnt3a and Wnt2b) and non-canonical Wnt-ligands (Wnt5a, Wnt7a, Wnt8b and Wnt11) are detectable within submucosal and myenteric plexus. Further, corresponding Frizzled receptors (Fzd1, Fzd3, Fzd6, and Fzd7) and regulatory signaling mediators like R-Spondin/DKK ligands are present in the ENS of the small and large intestine. Further, data mining approaches revealed, that several Wnt-related molecules are expressed by enteric glial cell clusters and are dynamically regulated during the inflammatory manifestation of enteric gliosis.
Our results suggest, that canonical and non-canonical Wnt-signaling has a much broader impact on the mature ENS and its cellular homeostasis in health and inflammation, than previously anticipated.