Futaki Sugiko, Nakano Itsuko, Kawasaki Miwa, Sanzen Noriko, Sekiguchi Kiyotoshi
Laboratory of Matrixome Research and Application, Institute for Protein Research, Osaka University, 3-2 Yamadaoka, Suita, Osaka, 565-0871, Japan.
Regen Ther. 2019 May 10;12:55-65. doi: 10.1016/j.reth.2019.04.010. eCollection 2019 Dec 15.
The basement membrane (BM) is a sheet-like extracellular matrix (ECM) lining the basal side of epithelial and endothelial cells. The molecular composition of the BM diversifies as embryonic development proceeds, providing optimized microenvironments for individual cell types. In post-implantation stage embryos, the embryonic BMs are essential for differentiation of the epiblast, a layer of multipotent embryonic stem cells, and subsequent embryogenesis. To better understand the role of BMs and cell-BM interactions in early embryogenesis, it is imperative to accumulate information on the molecular entities of the embryonic BMs.
We analyzed the expressions and localizations of 20 major BM proteins (11 laminin subunits, 6 type IV collagen subunits, nidogen-1 and -2, and perlecan) and other ECM-related proteins such as fibronectin and integrins in post-implantation stage embryos by immunohistochemistry.
We found that a set of BM proteins, laminin α5, β1, and γ1 (comprising laminin-511), type IV collagen α1 and α2 (yielding type IV collagen α1α2 [IV]), nidogen-1 and -2, and perlecan, were consistently present in the epiblast/ectoderm BMs throughout the early post-implantation stages. In contrast, laminin α1 was detected in the epiblast BM at E5.5 but decreased in later stages, suggesting that laminin-511 is a major laminin isoform in the early embryonic BM. In addition, fibronectin, a mesenchymal ECM protein, was enriched in the endoderm BM, indicating that the BM compositions differ between the ectoderm and the endoderm. Consistent with these observations, integrin α5, a high-affinity receptor for fibronectin, was localized in the endoderm, while integrin α6, a receptor for laminin-511, was localized in the ectoderm.
The embryonic BMs underlying the epiblast/ectoderm contain a common toolkit comprising laminin-511, type IV collagen (α1α2 [IV]), nidogen-1 and -2, and perlecan, providing a physiological basis for the utility of laminin-511 as a culture substrate for pluripotent stem cells. The distinctive association of laminin-511 and fibronectin with endodermal and ectodermal cells, together with the differential expression of integrin α5 and α6 in these cells, suggests that the ectodermal and endodermal cells rely on their integrin-dependent interactions with laminin-511 and fibronectin, respectively, to ensure their fate specification in embryonic development.
基底膜（BM）是一种片状细胞外基质（ECM），衬于上皮细胞和内皮细胞的基底侧。随着胚胎发育的进行，基底膜的分子组成多样化，为不同类型的细胞提供优化的微环境。在植入后阶段的胚胎中，胚胎基底膜对于上胚层（一层多能胚胎干细胞）的分化以及随后的胚胎发生至关重要。为了更好地理解基底膜和细胞 - 基底膜相互作用在早期胚胎发生中的作用，积累有关胚胎基底膜分子实体的信息势在必行。
我们通过免疫组织化学分析了植入后阶段胚胎中20种主要基底膜蛋白（11种层粘连蛋白亚基、6种IV型胶原亚基、巢蛋白 -1和 -2以及基底膜聚糖）以及其他细胞外基质相关蛋白（如纤连蛋白和整合素）的表达和定位。
我们发现一组基底膜蛋白，即层粘连蛋白α5、β1和γ1（组成层粘连蛋白 -511）、IV型胶原α1和α2（产生IV型胶原α1α2 [IV]）、巢蛋白 -1和 -2以及基底膜聚糖，在植入后早期阶段的整个上胚层/外胚层基底膜中持续存在。相比之下，层粘连蛋白α1在E5.5时在上胚层基底膜中被检测到，但在后期减少，这表明层粘连蛋白 -511是早期胚胎基底膜中的主要层粘连蛋白异构体。此外，纤连蛋白（一种间充质细胞外基质蛋白）在内胚层基底膜中富集，表明外胚层和内胚层的基底膜组成不同。与这些观察结果一致，纤连蛋白的高亲和力受体整合素α5定位于内胚层，而层粘连蛋白 -511的受体整合素α6定位于外胚层。
上胚层/外胚层下方的胚胎基底膜包含一个由层粘连蛋白 -511、IV型胶原（α1α2 [IV]）、巢蛋白 -1和 -2以及基底膜聚糖组成的共同工具包，为层粘连蛋白 -511作为多能干细胞培养底物的实用性提供了生理基础。层粘连蛋白 -511和纤连蛋白与内胚层和外胚层细胞的独特关联，以及这些细胞中整合素α5和α6的差异表达，表明外胚层和内胚层细胞分别依赖它们与层粘连蛋白 -511和纤连蛋白的整合素依赖性相互作用，以确保它们在胚胎发育中的命运特化。
