DOI
https://doi.org/10.25772/C34C-ZT87
Defense Date
2016
Document Type
Thesis
Degree Name
Master of Science
Department
Biomedical Engineering
First Advisor
Dr.Daniel Conway
Abstract
Numerous three-dimensional model systems have emerged for emulating the biochemical and physiological states of native tissue. Yet little is known about the effects of mechanical forces on cell behavior in the context of an organized tissue structure in three-dimensional cell-culture. Epithelial cells cultured in a three-dimensional environment comprised of extracellular matrix proteins form spheroids of polarized cells. Cellular responses to mechanical cues, generated from dynamic interactions with the extracellular matrix and neighboring cells, are known to influence cellular behavior to a great extent. Previous studies have shown that tumorigenic progression has been frequently linked to the down regulation of E-cadherin, a cell-cell adhesion protein. This work proposes that E-cadherin plays a pivotal role in maintaining epithelial tissue integrity and homeostasis. Novel FRET-based biosensors were used to measure force across E-cadherin. First, I observed that 3D acini had significantly higher force than 2D monolayers. Next, I determined that low-force mutant phenotypes of E-cadherin resulted in impaired lumen formation. In order to examine the effects of E-cadherin force on the disruption of homeostasis, TGF-b was used to induce epithelial to mesenchymal transition (EMT). TGF-b resulted in a decrease in E-cadherin force, even at early time points prior to transcriptional changes. Forskolin, a known regulator of acini lumen size, was shown to increase E-cadherin force. Furthermore, forskolin was able to prevent TGF-b disruptions in acini homeostasis. Finally, I examined how changes in substrate stiffness, known to affect acini lumen structure, altered E-cadherin forces. Stiffer substrates (mediated by collagen doping of Matrigel) delivered higher E-cadherin forces while simultaneously including acinar luminal filling. It is possible that signaling through non-junction forces, due to changes in ECM proteins, may mediate loss of the lumen. Thus, the major conclusion of these studies is that higher E-cadherin force is required for the formation and maintenance of a single central lumen in epithelial acini. Lower junctional forces induced acinar luminal filling, possibly through disruption in the polarity and subsequent cellular reorganization. This work, thus, establishes the role of E-cadherin as a key regulator of tissue homeostasis.
Rights
© The Author
Is Part Of
VCU University Archives
Is Part Of
VCU Theses and Dissertations
Date of Submission
5-12-2016
Included in
Biological Engineering Commons, Biotechnology Commons, Cancer Biology Commons, Cell Biology Commons, Molecular, Cellular, and Tissue Engineering Commons, Molecular Genetics Commons, Other Biomedical Engineering and Bioengineering Commons