File:EB development and TJ formation.png

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Figure 2. Embryoid bodies (EB) development and tight junction (TJ) formation.

(A) Schematic diagram of EB development. The development of EBs starts with the aggregation of ESCs into suspended spheroids of inner cell mass (ICM) and parallels the development of the mouse blastocyst ICM. By day 3 of culture, the outer cells differentiate into a circumferential epithelium known as the primitive endoderm (PrEn). The apical and basal domains of the PrEn face the exterior environment and interior ICM core of the EB respectively. The EBs at this point are called simple EBs. Progressively from day 4 onwards, the PrEn basally secretes key ECM components Laminins and Collagen IV to form a basement membrane (BM). The PrEn also further differentiates in the visceral endoderm (VEn). In conjunction with this, the interior ICM differentiates into the epiblast. By day 5 or 6, the epiblast in contact with the BM in turn differentiates into the primitive ectoderm (PrEc) or epiblast epithelium. The rest of the epiblast not in contact with the BM will undergo apoptosis. The apoptotic bodies are removed by autophagy-initiated phagocytosis, leaving a progressively enlarging lumen or Proamniotic-like cavity (PAC) surrounded by the apical domain of the PrEc and the basal BM. EBs at this stage are called cystic EBs and equivalent to the egg cylinder stage of the mouse peri-implantation embryo just before gastrulation. (B) Immunofluorescence staining of ZO-1 and ZO-2 in Day-5 or -6 EB cryosections. WT (panels a and e), ZO-1-/- (panels b and f), ZO-2-/- (panels c and g) and ZO-1-/- ZO-2-/- (panels d and h) EBs were stained with antibodies to ZO-1 (panels a-d, green color) or ZO-2 (panels e-h, green color). Nuclei are labeled with DAPI (blue color). Magnification of image in insets. ExEn is indicated here as ‘en’. (C) Transmission electron micrographs. TJ complex at the apico-lateral membrane of Day-9 EB ExEn was visualized by TEM as electron-dense material (indicated by arrows in magnification of inset) in WT (panels a and e), ZO-1-/- (panels b and f) and ZO-2-/- (panels c and g) EBs. This was absent in ZO-1-/- ZO-2-/- EBs (panels d and h, arrowheads in magnification of inset). (D) Immunofluorescence staining of Cldn-6 in Day-9 EB cryosections. WT, ZO-1-/-, ZO-2-/- and ZO-1-/- ZO-2-/- EBs were stained with antibodies to Cldn-6 (panels a–d, green color). Nuclei are labeled with DAPI (blue color). Magnification of image in insets. ExEn is indicated here as ‘en’.
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Source https://journals.plos.org/plosone/article?id=10.1371/journal.pone.0099532 Phua DCY, Xu J, Ali SM, Boey A, Gounko NV, Hunziker W (2014) ZO-1 and ZO-2 Are Required for Extra-Embryonic Endoderm Integrity, Primitive Ectoderm Survival and Normal Cavitation in Embryoid Bodies Derived from Mouse Embryonic Stem Cells. PLoS ONE 9(6): e99532. https://doi.org/10.1371/journal.pone.0099532
Author Phua DCY, Xu J, Ali SM, Boey A, Gounko NV, Hunziker W
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Copyright: © 2014 Phua et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

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current17:46, 2 September 2024Thumbnail for version as of 17:46, 2 September 20241,350 × 2,712 (1.93 MB)Rasbak (talk | contribs){{Information |description=Figure 2. EB development and TJ formation.<br> (A) Schematic diagram of EB development. The development of EBs starts with the aggregation of ESCs into suspended spheroids of inner cell mass (ICM) and parallels the development of the mouse blastocyst ICM. By day 3 of culture, the outer cells differentiate into a circumferential epithelium known as the primitive endoderm (PrEn). The apical and basal domains of the PrEn face the exterior environment and interior ICM c...

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