Keratin filaments mediate the expansion of extra‐embryonic membranes in the post‐gastrulation mouse embryo

Abstract

Mesoderm arises at gastrulation and contributes to both the mouse embryo proper and its extra‐embryonic membranes. Two‐photon live imaging of embryos bearing a keratin reporter allowed recording filament nucleation and elongation in the extra‐embryonic region. Upon separation of amniotic and exocoelomic cavities, keratin 8 formed apical cables co‐aligned across multiple cells in the amnion, allantois, and blood islands. An influence of substrate rigidity and composition on cell behavior and keratin content was observed in mesoderm explants. Embryos lacking all keratin filaments displayed a deflated extra‐embryonic cavity, a narrow thick amnion, and a short allantois. Single‐cell RNA sequencing of sorted mesoderm cells and micro‐dissected amnion, chorion, and allantois, provided an atlas of transcriptomes with germ layer and regional information. It defined the cytoskeleton and adhesion expression profile of mesoderm‐derived keratin 8‐enriched cells lining the exocoelomic cavity. Those findings indicate a novel role for keratin filaments in the expansion of extra‐embryonic structures and suggest mechanisms of mesoderm adaptation to the environment.

Regionally specific levels and patterns of keratin 8 expression in the mouse embryo visceral endoderm emerge upon anterior-posterior axis determination

“…At E7.5 ( Figure 5A ), K8 marked the extra-embryonic ectoderm and mesoderm, as previously described ( Saykali et al, 2019 ; Nahaboo et al, 2022 ), and was enriched at cell-cell junctions in the endoderm ( Figures 5B–E,B′–E′ ; Supplementary Video S9 ), which at this stage is composed of VE only in the extra-embryonic region, and of a mixture of VE and definitive endoderm cells that intercalate between VE cells in the embryonic region ( Kwon et al, 2008 ). Single cell RNA sequencing showed that definitive endoderm cells have a lower mean level of Krt8 transcripts than VE at E7.25; between E7.5 and E8 Krt8 mRNA content tends to equalize in the high range in all endoderm cells ( https://marionilab.cruk.cam.ac.uk/MouseGastrulation2018/ ( Pijuan-Sala et al, 2019 )).…”

“…Over time, K8 positive dots became connected by filamentous elements, primarily at the membrane of outer cells at the morula stage, then in the cytoplasm at the blastocyst stage, progressively creating a dense web. The generation of a K8-containing mesh was also observed in extra-embryonic mesoderm cells surrounding the extra-embryonic cavity in late gastrulation embryos ( Nahaboo et al, 2022 ).…”

“…3D reconstructions of stills from confocal imaging of an E5.75 K8-eYFP embryo (yellow) imaged every 15 min for a total of 120 min. (B) 3D reconstructions of stills from confocal imaging of an E6.5 K8-eYFP (yellow, also depicted in ( Nahaboo et al, 2022 ) imaged every 60 min for a total of 300 min. (C) 3D reconstructions of stills from confocal imaging of an E7.5 K8-eYFP (yellow, also depicted in ( Nahaboo et al, 2022 ) imaged every 60 min for a total of 180 min.…”

Modeling human extraembryonic mesoderm cells using naive pluripotent stem cells

“…The extraembryonic mesoderm (EXM) is an important tissue with essential roles in development. EXM is implicated in primitive erythropoiesis and extracellular matrix formation; becomes an integral part of the amnion, yolk sac, allantois, and chorion; and forms the primitive umbilical cord (Enders and King, 1988;Luckett, 1978;Nahaboo et al, 2022;Ross and Boroviak, 2020;Sadler, 2012;Saykali et al, 2019;Shepard, 1989;Spencer Chapman et al, 2021). Intriguingly, the mechanisms of EXM specification differ dramatically between species of mammals.…”

Biofunctionalization of hydrogel-based scaffolds for vascular tissue regeneration

“…The neoformation of blood vessels is mandatory for tissue regeneration, since it is the main step for the success of the new tissue functions, allowing rapid adaptation and providing microvessels that connect with the host's existing vessels, achieving prompt homeostasis (HUA et al, 2021;Szklanny et al, 2021). The development of different techniques to convert embryonic cells into vascularized tissues enable the formation of specialized tissues and even mini-organs, which are capable of creating a vascular network when they are auto transplanted, but also when they are grafted onto extraembryonic membrane systems (Nahaboo et al, 2022). For example, the Chorioallantoic Membrane System (CAM), which is one of the most widely used assays for the study of vasculogenesis, allows easy handling and manipulation.…”