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Differentiation of Primary Human Pro-Fibrotic Mesenchymal Cells Influenced by Extracellular Matrix Environment Determined by MicroRNA Expression
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- Author / Creator
- Müller, Alison L
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Fibrosis, a multi-faceted process that exacerbates numerous cardiovascular pathologies leading
to heart failure, is the result of excessive extracellular matrix protein deposition. Although fibrosis is
classically thought to be the result of myofibroblasts activated from interstitial fibroblasts endogenously
present within the heart, recent research has indicated another significant pro-fibrotic cell source which
resides in the bone marrow. These mesenchymal progenitor cells are recruited to the heart in response to
inflammation in a variety of cardiovascular diseases and have been directly implicated in extracellular
matrix protein deposition. The fibrotic extracellular matrix environment significantly influences the
differentiation of mesenchymal progenitor cells. These cells continuously deposit extracellular matrix
proteins worsening fibrosis and further recruiting circulating bone marrow-derived mesenchymal
progenitor cells. This hostile fibrotic environment has also stunted and further complicated the success of
regenerative cell therapy which aims to stimulate cardiac repair and healing. In the many cardiovascular
disorders that lead to fibrosis, these endogenous fibroblasts and bone marrow-derived progenitor cells
differentiate rapidly to the myofibroblast phenotype in response to injury. This pro-fibrotic differentiation
process has been shown to be facilitated and guided by microRNA. The putative role of microRNA has
been implied in both the differentiation of bone marrow-derived stem cells and interstitial fibroblasts to
myofibroblasts. The human body has a myriad of different environments that contribute to cell
differentiation, which includes stiff pathological fibrotic environments. Pathological fibrosis occurs in
response to injury which alters the microenvironment experienced by mesenchymal cells by promoting a
pro-fibrotic phenotype which is influenced, in part, by miRNA expression. The significant potential for a
link between miRNA, the extracellular matrix environment, and the differentiation of various progenitor
cell-types and their contribution to cardiac fibrosis will be explored in this thesis. We hypoethsize that
mesenchymal cells, both bone marrow-derived progenitor cells and atrial fibroblasts, are susceptible to
changes in their surrounding environment that influences their behaviour. As decellularization techniques
can be utilized to isolate native extracellular matrix (ECM), it is possible to evaluate mesenchymal cell
phenotypic changes on an endogenous pro-fibrotic environment. Therefore, this hypothesis will be tested
iiiusing physiological, native ECM isolated from human left ventricular cardiac tissue. This pro-fibrotic
influence may be attenuated by over-expression of miRNA-301a.
This shall be accomplished by first evaluating the effects of statin therapy on the mesenchymal
progenitor cell population to establish how they interact with potential cell sources that would be utilized
in clinical applications. It is recognized that mesenchymal cells that contribute to pathological fibrosis
undergo a proliferative phenotype change upon exposure to inflammation as a result of injury. We identified
a miRNA involved in this phenotype switch and determined how it influences the progression of
mesenchymal cells as they exhibit myofibroblast-like properties under normal culture conditions.
Understanding that progenitor cell differentiation is determined by their niche microenvironment, including
extracellular stiffness, we evaluated how human mesenchymal progenitor cells (hMPCs) respond to varying
surface tensions while measuring possible changes in miRNA expression. In order to observe
physiologically accurate phenomena, it is possible to decellularize native tissue to isolate ECM. With access
to human cardiac tissue, both diseased and healthy, we focused on the influence of pathologically remodeled
ECM on hMPCs and human atrial fibroblasts (hAFs), by using decellularization techniques to isolate
fibrotic-rich areas of left ventricular (LV) myocardium and subsequently recellularizing them with hMPCs
and hAFs. Although studying the effects of hMPCs and hAFs on human LV, we also evaluated the effects
of these cells on commercially available decellularized porcine matrix, called CorMatrix®, which is used
in a variety of cardiac surgery procedures. Finally, in addition to being able to evaluate molecular
differences between ECM products, native diseased and healthy ECM, we assessed the efficacy of miR
301a in attenuating differentiation into a pro-fibrotic phenotype on hAFs and hMPCs seeded in profibrotic
decellularized LV ECM. -
- Graduation date
- Fall 2018
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- Type of Item
- Thesis
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- Degree
- Doctor of Philosophy
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- License
- Permission is hereby granted to the University of Alberta Libraries to reproduce single copies of this thesis and to lend or sell such copies for private, scholarly or scientific research purposes only. Where the thesis is converted to, or otherwise made available in digital form, the University of Alberta will advise potential users of the thesis of these terms. The author reserves all other publication and other rights in association with the copyright in the thesis and, except as herein before provided, neither the thesis nor any substantial portion thereof may be printed or otherwise reproduced in any material form whatsoever without the author's prior written permission.