Dynamics, structure and assembly of the basement membrane in developing salivary glands revealed by an exogenous EGFP-tagged nidogen probe
Dynamics, structure and assembly of the basement membrane in developing salivary glands revealed by an exogenous EGFP-tagged nidogen probe
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外源性 EGFP 标记巢探针揭示唾液腺发育中基底膜的动力学、结构和组装
DOI：
10.1093/jmicro/dfac040
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发表时间：
2022
期刊：
影响因子：
1.8
通讯作者：
Sekiguchi Kiyotoshi
中科院分区：
文献类型：
--
作者：
Kadoya Yuichi;Futaki Sugiko;Shimono Chisei;Kimura Taketoshi;Sekiguchi Kiyotoshi
大多数上皮组织在胚胎发育期间迅速变得复杂，同时被基底膜（BM）包围。因此，BM形状被认为随着上皮细胞的生长而发生显著变化，但其潜在机制尚不清楚。巢蛋白-1在BM中普遍存在，并结合各种其他BM组分，包括层粘连蛋白和IV型胶原。为了阐明BM在上皮形态发生过程中的行为，我们尝试用与增强型绿色荧光蛋白（hNid 1-EGFP）融合的重组人巢蛋白-1活标记发育中的BM。小鼠胚胎的下颌下腺在玻璃底培养皿中培养，并在含有hNid 1-EGFP的培养基中孵育。随后的共聚焦显微镜清楚地显示了上皮终芽周围的BM。在从Z系列共聚焦切片的三维重建上，观察到上皮BM作为围绕整个上皮基底表面连续扩张的薄片。因为外植体在hNid 1-EGFP存在下继续生长良好，所以进行延时共聚焦显微镜以跟踪BM的动力学。我们发现，上皮BM是一种适应性结构，其根据发育上皮的快速形状变化而变形。此外，hNid 1-EGFP被发现被纳入不同的上皮BM相比，纤连蛋白或IV型胶原蛋白，这表明个别BM组件组装以不同的方式形成BM。
Most epithelial tissues rapidly become complex during embryonic development while being surrounded by the basement membrane (BM). Thus, the BM shape is thought to change dramatically as the epithelium grows, but the underlying mechanism is not yet clear. Nidogen-1 is ubiquitous in the BM and binds to various other BM components, including laminin and type IV collagen. To elucidate the behavior of the BM during epithelial morphogenesis, we attempted to live-label the developing BM with recombinant human nidogen-1 fused to an enhanced green fluorescent protein (hNid1-EGFP). Submandibular glands of mouse embryos were cultured in glass-bottomed dishes and incubated in media containing hNid1-EGFP. Subsequent confocal microscopy clearly visualized the BMs surrounding the epithelial end buds. On three-dimensional reconstruction from Z-series confocal sections, the epithelial BM was observed as a thin sheet that expanded continuously around the entire epithelial basal surface. Because the explants continued to grow well in the presence of hNid1-EGFP, time-lapse confocal microscopy was performed to follow the dynamics of the BM. We found that the epithelial BM is an adaptive structure that deforms in accordance with the rapid shape changes of the developing epithelium. Furthermore, hNid1-EGFP was found to be incorporated differently into the epithelial BM compared with that reported for fibronectin or type IV collagen, suggesting that individual BM components assemble in different ways to form the BM.