Date of Award

Spring 1-1-2015

Document Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Department

Chemistry & Biochemistry

First Advisor

Kristi S. Anseth

Second Advisor

Amy E. Palmer

Third Advisor

Xuedong Liu

Fourth Advisor

Robert T. Batey

Fifth Advisor

Leslie A. Leinwand

Abstract

Multi-potent human mesenchymal stem cells (hMSCs) are widely explored in clinical trials for their applications in cell-based therapy, due to their multipotency and the ability to secrete trophic factors. hMSCs also play a major role in musculoskeletal tissue homeostasis, as well as the pathophysiology of several inflammatory and degenerative diseases, but the understanding of mechanism is limited. It is important, but challenging to delineate, separate or even identify the source of a specific effect in the complex native extracellular environment (ECM). Therefore, in my thesis, poly (ethylene glycol) hydrogels were applied as synthetic ECM mimics to assist the investigation, which provides a pseudo "blank slate" that can minimize non-specific protein interaction and allow incorporation of specific biochemical and mechanical cues of interest through active chemical handles.

The focus of my thesis is to understand the osteogenic differentiation process of hMSCs in the presence of dynamic biochemical and mechanical cues. Particularly, I synthesized and modified PEG-norbornene hydrogels by thiol-ene chemistry with matrix metalloproteinase degradable peptide to allow cell-mediated enzymatic delivery of potent small molecule (e.g. Dexamethasone), and characterize hMSCs response to these locally delivered small molecules. On the other hand, I studied the mechanotransduction in hMSCs by using nitrobenzyl photodegradable material to dynamically control the stiffness of the extracellular substrate and characterize hMSCs response to these in situ physical modifications by monitoring the localization of YAP, a transcriptional co-activator that mediated cellular mechanosensing. I found that cell fates of hMSCs were biased by the previous physical culture condition, that is, hMSCs possess mechanical memory. Sub-cellular activation of YAP was found to act analogous to a mechanical rheostat and regulate the intensity of the response, by which induced reversible and irreversible osteogenesis. I further explored the mechanism that directed the mechanical signaling and dosing process by exploiting the spatial control of the degradation process of the phototunable hydrogels to investigate hMSCs responses to spatial variations in substrate modulus. We found that spatial presentation of mechanical cues affects adhesion, mechanotransduction and differentiation of hMSCs.

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