Mechanopluripotency – Adherens Junction Mechanotransduction Supports mESC Pluripotency Gene Expression via β-catenin Nuclear Translocation in Response to Fluid Shear Stress

Harper, Lane Ward

Mechanopluripotency – Adherens Junction Mechanotransduction Supports mESC Pluripotency Gene Expression via β-catenin Nuclear Translocation in Response to Fluid Shear Stress

dc.contributor.advisor	Rancourt, Derrick Emile
dc.contributor.author	Harper, Lane Ward
dc.contributor.committeemember	Moorhead, Gregory B.
dc.contributor.committeemember	Sen, Arindom
dc.date	2024-02
dc.date.accessioned	2023-11-08T22:57:44Z
dc.date.available	2023-11-08T22:57:44Z
dc.date.issued	2023-11-07
dc.description.abstract	Pluripotent stem cells are increasingly being investigated as a base platform from which effector cells can be differentiated and utilized in various contexts as whole-cell therapeutics. Current culture methodologies require 2D plate culture and this represents a bottleneck in the form of increased costs associated with consumable usage and difficulties with standardization in manipulation. These difficulties cascade into ballooning therapy costs and troubles with cGMP compliance required for all biopharmaceutical manufacturing. To this end, there has been interest in culturing pluripotent stem cells in bioreactors, shown in previous therapeutic biopharmaceutical platforms to achieve cost-effective scale and compliance. In our bioreactor work, we have observed an increase in core pluripotency gene expression in mouse embryonic stem cells (mESCs) cultured as aggregates at specific impeller speeds in stirred tank bioreactors. We have termed this phenomenon “mechanopluripotency”. This project aimed to investigate the mechanism(s) responsible for our observation of induced pluripotency in the bioreactor. Using mutant cell lines containing a β-catenin reporter system, we show in both the bioreactor and a defined model system that in response to shear stress of ~6 dyne/cm2 β-catenin translocates to the nucleus where it is responsible for co-activation of the Wnt signaling pathway, known to affect pluripotency gene expression. Additionally, we show evidence that this response is mediated via altered molecular dynamics at the adherens junction, a key hub for cell-cell connections. By knocking out the cytoskeleton protein vinculin we show that β-catenin signaling is impaired in both the bioreactor and defined model system and a decrease in core pluripotency gene expression in the absence of exogenous maintenance factors. These results indicate that mechanopluripotency is mediated primarily through β-catenin and by impairing the adherens junction’s ability to adapt to mechanical force the response is attenuated.
dc.identifier.citation	Harper, L. W. (2023). Mechanopluripotency – adherens junction mechanotransduction supports mESC pluripotency gene expression via β-catenin nuclear translocation in response to fluid shear stress (Master's thesis, University of Calgary, Calgary, Canada). Retrieved from https://prism.ucalgary.ca.
dc.identifier.uri	https://hdl.handle.net/1880/117546
dc.identifier.uri	https://doi.org/10.11575/PRISM/42389
dc.language.iso	en
dc.publisher.faculty	Graduate Studies
dc.publisher.institution	University of Calgary
dc.rights	University of Calgary graduate students retain copyright ownership and moral rights for their thesis. You may use this material in any way that is permitted by the Copyright Act or through licensing that has been assigned to the document. For uses that are not allowable under copyright legislation or licensing, you are required to seek permission.
dc.subject	Stem Cells
dc.subject	Biomedical Engineering
dc.subject	Bioreactor
dc.subject	Biochemistry
dc.subject	Flow Chamber
dc.subject.classification	Biochemistry
dc.subject.classification	Engineering--Biomedical
dc.title	Mechanopluripotency – Adherens Junction Mechanotransduction Supports mESC Pluripotency Gene Expression via β-catenin Nuclear Translocation in Response to Fluid Shear Stress
dc.type	master thesis
thesis.degree.discipline	Engineering – Biomedical
thesis.degree.grantor	University of Calgary
thesis.degree.name	Master of Science (MSc)
ucalgary.thesis.accesssetbystudent	I do not require a thesis withhold – my thesis will have open access and can be viewed and downloaded publicly as soon as possible.