Random migration of induced pluripotent stem cell-derived human gastrulation-stage mesendoderm.

Yuta Yamamoto,Akiko Kondow,Ayumu Kano,Ryo Kobayashi,Shuji Fujii,Shota Miyazaki,Kiyoshi Ohnuma,Minh Nguyen Tuyet Le,Kenshiro Maruyama,Masaharu Seno

doi:10.1371/journal.pone.0201960

Yuta Yamamoto, Akiko Kondow + Show 8 more

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https://doi.org/10.1371/journal.pone.0201960

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Abstract

Gastrulation is the initial systematic deformation of the embryo to form germ layers, which is characterized by the placement of appropriate cells in their destined locations. Thus, gastrulation, which occurs at the beginning of the second month of pregnancy, is a critical stage in human body formation. Although histological analyses indicate that human gastrulation is similar to that of other amniotes (birds and mammals), much of human gastrulation dynamics remain unresolved due to ethical and technical limitations. We used human induced pluripotent stem cells (hiPSCs) to study the migration of mesendodermal cells through the primitive streak to form discoidal germ layers during gastrulation. Immunostaining results showed that hiPSCs differentiated into mesendodermal cells and that epithelial–mesenchymal transition occurred through the activation of the Activin/Nodal and Wnt/beta-catenin pathways. Single-cell time-lapse imaging of cells adhered to cover glass showed that mesendodermal differentiation resulted in the dissociation of cells and an increase in their migration speed, thus confirming the occurrence of epithelial–mesenchymal transition. These results suggest that mesendodermal cells derived from hiPSCs may be used as a model system for studying migration during human gastrulation in vitro. Using random walk analysis, we found that random migration occurred for both undifferentiated hiPSCs and differentiated mesendodermal cells. Two-dimensional random walk simulation showed that homogeneous dissociation of particles may form a discoidal layer, suggesting that random migration might be suitable to effectively disperse cells homogeneously from the primitive streak to form discoidal germ layers during human gastrulation.

Highlights

Gastrulation, which is critical to human body formation, is the initial dynamic deformation of the embryo and occurs at the beginning of the second month of pregnancy, which is 2–3 weeks after fertilization (Fig 1a) [1,2,3]
To confirm that human induced pluripotent stem cells (hiPSCs) differentiated into mesendodermal cells during gastrulation, we first assessed the expression of mesendodermal and epithelial to mesenchymal transition (EMT) markers
We assessed the expression of four marker proteins, namely, OCT3/4 and E-cadherin, BRACHYURY and SNAIL [25, 26]

Summary

Introduction

Gastrulation, which is critical to human body formation, is the initial dynamic deformation of the embryo and occurs at the beginning of the second month of pregnancy, which is 2–3 weeks after fertilization (Fig 1a) [1,2,3]. The epiblast, which is comprised of epithelial cells, differentiates into the ectoderm, which forms the neural system, and the mesendoderm, comprised of mesenchymal cells that form the muscles, heart, bone, and the digestive tract. Mesendodermal cells migrate to their appropriate locations through the primitive streak to form the discoidal mesoderm and endoderm layers. The three germ layers, ectoderm, mesoderm, and endoderm, derived from the epiblast, in turn, form the entire body (Fig 1a). Histology as well as genetic regulation of early human embryogenesis have been studied [5, 6], cellular dynamics of early human embryogenesis is not well understood largely due to ethical and technical limitations

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