Department of Biological Sciences, Faculty of Science, Kanagawa University, Tsuchiya 2946, Hiratsuka City, Kanagawa, 259-1293, Japan.
Department of Biological Sciences, Faculty of Science, Kanagawa University, Tsuchiya 2946, Hiratsuka City, Kanagawa, 259-1293, Japan; Research Institute for Integrated Science, Kanagawa University, Japan.
Zoology (Jena). 2021 Jun;146:125905. doi: 10.1016/j.zool.2021.125905. Epub 2021 Feb 12.
In vertebrates, gut coiling proceeds left-right asymmetrically during expansion of the gastrointestinal tract with highly organized muscular structures facilitating peristalsis. In this report, we explored the mechanisms of larval gut coiling morphogenesis relevant to its nascent smooth muscle cells using highly transparent Xenopus early larvae. First, to visualize the dynamics of intestinal smooth muscle cells, whole-mount specimens were immunostained with anti-smooth muscle-specific actin (SM-actin) antibody. We found that the nascent gut of Xenopus early larvae gradually expands the SM-actin-positive region in a stage-dependent manner. Transverse orientation of smooth muscle cells was first established, and next, the cellular longitudinal orientation along the gut axis was followed to make a meshwork of the contractile cells. Finally, anisotropic torsion by the smooth muscle cells was generated in the center of gut coiling, suggesting that twisting force might be involved in the late phase of coiling morphogenesis of the gut. Administration of S-(-)-Blebbistatin to attenuate the actomyosin contraction in vivo resulted in cancellation of coiling of the gut. Development of decapitation embryos, trunk 'torso' explants, and gut-only explants revealed that initial coiling of the gut proceeds without interactions with the other parts of the body including the central nervous system. We newly developed an in vitro model to assess the gut coiling morphogenesis, indicating that coiling pattern of the nascent Xenopus gut is partially gut-autonomous. Using this gut explant culture technique, inhibition of actomyosin contraction was performed by administrating either actin polymerization inhibitor, myosin light chain kinase inhibitor, or calmodulin antagonist. All of these reagents decreased the extent of gut coiling morphogenesis in vitro. Taken together, these results suggest that the contraction force generated by actomyosin-rich intestinal smooth muscle cells during larval stages is essential for the normal coiling morphogenesis of this muscular tubular organ.
在脊椎动物中，胃肠道扩张过程中肠道呈左右不对称卷曲，高度组织化的肌肉结构促进蠕动。在本报告中，我们使用高度透明的非洲爪蟾早期幼虫探索了与新生平滑肌细胞相关的幼虫肠道卷曲形态发生的机制。首先，为了可视化肠平滑肌细胞的动力学，使用抗平滑肌特异性肌动蛋白（SM-actin）抗体对全胚胎标本进行免疫染色。我们发现，非洲爪蟾早期幼虫的新生肠道以依赖于阶段的方式逐渐扩展 SM-actin 阳性区域。平滑肌细胞的横向取向首先建立，然后沿着肠道轴的细胞纵向取向被跟随以形成收缩细胞的网格。最后，在肠道卷曲的中心产生了平滑肌细胞的各向异性扭曲，表明扭转力可能参与肠道卷曲形态发生的后期阶段。在体内施用 S-(-)-Blebbistatin 以减弱肌动球蛋白收缩导致肠道卷曲的取消。断头胚胎、躯干“躯干”外植体和仅肠道外植体的发育表明，肠道的初始卷曲不需要与身体的其他部分（包括中枢神经系统）相互作用而进行。我们新开发了一种体外模型来评估肠道卷曲形态发生，表明新生非洲爪蟾肠道的卷曲模式部分是肠道自主的。使用这种肠道外植体培养技术，通过给予肌动蛋白聚合抑制剂、肌球蛋白轻链激酶抑制剂或钙调蛋白拮抗剂来抑制肌动球蛋白收缩。所有这些试剂都减少了体外肠道卷曲形态发生的程度。总之，这些结果表明，肌球蛋白丰富的肠道平滑肌细胞在幼虫阶段产生的收缩力对于这种肌肉管状器官的正常卷曲形态发生是必不可少的。
