Biology subjects

Krakora, M. Z.

Publications and source records attributed to Krakora, M. Z..

2 recordsLinked to original sources

bioRxiv2020

Silencing of E-cadherin in induced human pluripotent stem cells promotes extraembryonic fates accompanying multilineage differentiation

Summary/AbstractIn embryonic development, symmetry breaking events and the mechanical milieus in which they occur coordinate the specification of separate cell lineages. Here, we use 3D aggregates of human pluripotent stem cells (hPSCs) encapsulated in alginate microbeads to model the early blastocyst prior to zona pellucida hatching. We demonstrate that 3D confinement combined with modulation of cell-cell adhesions is sufficient to drive differentiation and collective migration reminiscent of the pre-implantation embryo. Knockdown of the cell adhesion protein CDH1 in encapsulated hPSC aggregates resulted in protrusion morphologies and emergence of extra-embryonic lineages, whereas unencapsulated CDH1(-) aggregates displayed organized radial delamination and mesendoderm specification bias. Transcriptomic similarities between single-cell RNA-sequencing data of early human embryos and encapsulated CDH1(-) aggregates establishes this in vitro system as a competent surrogate for studying early embryonic fate decisions and highlights the relationship between cell-cell adhesions and the mechanical microenvironment in directing cell fate and behavior. HighlightsO_LIGeneration of embryonic scale 3D morphogenesis using hydrogel encapsulation C_LIO_LIManipulating adhesion triggers emergence of specific morphologies and cell fates C_LIO_LIAcquisition of germ layer cell fates mimics early human embryonic diversity C_LI

developmental biology↗

bioRxiv2020

Axial Elongation of Caudalized Human Pluripotent Stem Cell Organoids Mimics Neural Tube Development

Axial elongation of the neural tube is critical during mammalian embryogenesis to establish the anterior-posterior body axis1, but this process is difficult to interrogate directly because it occurs post-implantation2,3. Here we report an organoid model of neural tube extension by caudalized human pluripotent stem cell (hPSC) aggregates that recapitulates the morphologic and temporal gene expression patterns of neural tube development. Axially extending organoids consisting largely of longitudinally elongated neuroepithelial compartments also contained TBXT(+)SOX2(+) neuromesodermal progenitors, PAX6(+)nestin(+) neural progenitor populations, and MEOX1(+) paraxial mesoderm populations. Wnt agonism stimulated singular axial extensions in a dose-dependent manner, and elongated organoids displayed regionalized rostral-caudal HOX gene expression, with spatially distinct hindbrain (HOXB1) expression from brachial (HOXC6) and thoracic (HOXB9) regions. CRISPR-interference-mediated silencing of the TBXT, a downstream Wnt target, increased neuroepithelial compartmentalization and resulted in multiple extensions per aggregate. Further, knock-down of BMP inhibitors, Noggin and Chordin, induced elongation phenotypes that mimicked murine knockout models. These results indicate the potent morphogenic capacity of caudalized hPSC organoids to undergo axial elongation in a manner that can be used to dissect the cellular organization and patterning decisions that dictate early nervous system development in humans.

bioengineering↗