Role of multicellular organisation in mesoderm differentiation
View/ Open
Date
07/03/2022Author
Wisniewski, Darren
Metadata
Abstract
What cues and signals are important for the differentiation of pluripotent cells in
vitro and in vivo? During development, pluripotent cell fate decisions are spatiotemporally
coordinated with changing tissue architecture. Typically, changes in
tissue architecture are thought to be a downstream consequence of changes in cell
fate, which in turn are caused by change in exposure to secreted extracellular
signalling molecules. However, recent evidence in multiple vertebrate systems
has shown that tissue architecture and tissue mechanics can influence and
causally feedback into cell fate decision-making processes. During gastrulation
in the mouse embryo the pseudostratified epithelium of the proximal epiblast
undergoes an epithelial to mesenchymal transition (EMT), forming the primitive
streak. This is accompanied by a number of changes in tissue architecture
including the loss of apical-basal polarity and cell-cell adhesion, an increase in
cell clustering and migratory behaviour, and changes in cell shape associated with
differentiation into mesodermal and endodermal derivatives. Primitive streak
formation and gastrulation are orchestrated by Nodal, BMP, Wnt and Fgf
signalling pathway activity. However, it is unclear if and how changes in tissue
architecture influence the differentiation responses in the context of this
signalling environment.
Here I set out to ask which aspects of changing tissue architecture can provide
regulatory feedback to influence cell fate decisions during gastrulation in the
mouse, and to explore the underlying mechanisms. Decoupling the contributions
of tissue architecture and the chemical signalling environment in cell fate
regulatory feedback is technically challenging in vivo. Therefore, I developed a
more controllable in vitro ECM-based mESC model in which I could manipulate
tissue architecture. Qualitative and quantitative analysis between in vitro and in
vivo cellular organisation confirmed that in vitro ECM-based models
recapitulated features of primitive streak morphogenesis. Further, I find that this
ECM-based culture model displays increased mesodermal differentiation and
demonstrates a more coherent patterning of cell fates, when compared with
monolayer cultures exposed to the same exogenous signals.
Using quantitative image analysis and further experimental manipulation of in
vitro tissue architecture, I identify a role for local cell density in promoting
mesodermal fates in ECM-based models. Finally, I demonstrate that active Nodal
and Wnt signalling are crucial in mediating enhanced mesodermal differentiation
induced by increased local cell density in ECM-based cultures. These results
highlight a potential positive regulatory feedback role for tissue architecture, in
the form of increased local cell density, during mesodermal differentiation in an
in vitro primitive streak model. We propose that the data suggests the requirement
of a critical population size for robust and coherent mesodermal differentiation
and that this mechanism is reminiscent of a community effect.