During early development, the enteric nervous system forms from the migration of enteric neural crest cells (ENCCs) from the foregut to the hindgut, where they undergo proliferation and differentiation facilitated by interactions with enteric mesenchymal cells (EMCs). This study investigates the impact on ENCC migration of EMC-ENCC communication mediated by GFRA1b expressed in EMCs. GFRA1-expressing cells in day 11–12 (E11–12) mouse embryos differentiated into smooth muscle cells from E12 onwards. Observations at E12–13.5 revealed high levels of GFRA1 expression on the anti-mesenteric side of the hindgut, correlating with enhanced ENCC migration. This indicates that GFRA1 in EMCs plays a role in ENCC migration during development. Examining GFRA1 isoforms, we found high levels of GFRA1b, which lacks amino acids 140–144, in EMCs. To assess the impact of GFRA1 isoforms on EMC-ENCC communication, we conducted neurosphere drop assays. This revealed that GFRA1b-expressing cells promoted GDNF-dependent extension and increased neurite density in ENCC neurospheres. Co-culture of ENCC mimetic cells expressing RET and GFRA1a with EMC mimetic cells expressing GFRA1a, GFRA1b, or vector alone showed that only GFRA1b-expressing co-cultured cells sustained RET phosphorylation in ENCC-mimetic cells for over 120 min upon GDNF stimulation. Our study provides evidence that GFRA1b-mediated cell-to-cell communication plays a critical role in ENCC motility in enteric nervous system development. These findings contribute to understanding the cellular interactions and signaling mechanisms that underlie enteric nervous system formation and highlight potential therapeutic targets for gastrointestinal motility disorders.

中文翻译：

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间充质细胞通过 RET-GFRA1b 反式信号调节肠神经嵴细胞迁移

在早期发育过程中，肠神经系统由肠神经嵴细胞（ENCC）从前肠迁移到后肠形成，在后肠中它们通过与肠间充质细胞（EMC）的相互作用促进增殖和分化。本研究调查了 EMC 中表达的 GFRA1b 介导的 EMC-ENCC 通讯对 ENCC 迁移的影响。第 11-12 天 (E11-12) 小鼠胚胎中的 GFRA1 表达细胞从 E12 开始分化为平滑肌细胞。 E12-13.5 的观察结果显示，后肠反肠系膜侧 GFRA1 表达水平较高，与 ENCC 迁移增强相关。这表明EMC中的GFRA1在开发过程中ENCC迁移中发挥作用。通过检查 GFRA1 亚型，我们发现在 EMC 中存在高水平的 GFRA1b（缺乏氨基酸 140-144）。为了评估 GFRA1 同种型对 EMC-ENCC 通讯的影响，我们进行了神经球滴落测定。这表明表达 GFRA1b 的细胞促进了 ENCC 神经球中 GDNF 依赖性延伸并增加了神经突密度。将表达 RET 和 GFRA1a 的 ENCC 模拟细胞与表达 GFRA1a、GFRA1b 或单独载体的 EMC 模拟细胞共培养表明，在 GDNF 刺激后，只有表达 GFRA1b 的共培养细胞在 ENCC 模拟细胞中维持 RET 磷酸化超过 120 分钟。我们的研究提供了证据，表明 GFRA1b 介导的细胞间通讯在肠神经系统发育中的 ENCC 运动中发挥着关键作用。这些发现有助于了解肠神经系统形成的细胞相互作用和信号传导机制，并强调胃肠动力障碍的潜在治疗靶点。