Synthetic surfaces to probe human pluripotent stem cell fate decisions

The field of regenerative medicine promises to restore function to failing organs and tissues in patients with debilitating degenerative diseases or injuries. One branch of the field focuses on discovering methods to induce endogenous tissues to regenerate or repair themselves. The other branch seeks to produce replacement tissues and organs in vitro to serve as a limitless source for transplants. Human pluripotent stem cells---with the ability to self-renew indefinitely in culture and differentiated into virtually any functional cell type---hold promise as source material for regenerative medicine. In order to achieve this promise, however, we must develop methods to control the proliferation and differentiation of the cells and the methods must keep them free of contaminative immunogens or pathogens. This thesis focuses on the juncture of these two goals. Initially, we developed of the first synthetic surface capable of supporting human pluripotent stem cells. This synthetic surface interacts with cell surface glycosaminoglycans, and I aimed to determine how insoluble cues influence the signaling mechanisms involved in cell fate determination. First, I developed peptide-presenting surfaces tailored to the changing adhesion needs of pluripotent stem cells as they differentiated into neural progenitor cells and motor neurons (Chapter 2). I demonstrated that cells differentiate more robustly toward endoderm and mesoderm lineages when cultured on GAG-binding surfaces. Further, I probed the molecular mechanisms involved and showed that integrin activation of Akt via integrin-linked kinase inhibits mesendoderm differentiation (Chapter 3). We also analyzed the effects of mechanical cues on stem cell fate. Human pluripotent stem cells require a surface of sufficient stiffness to maintain pluripotency. When cultured on softer surfaces, they rapidly differentiate into neurons, and we demonstrated that this process is governed by the mechanosensitive transcriptional coactivator Yes-associated protein (Chapter 4). Cumulatively, this thesis dissects and reveals mechanisms by which insoluble cues control human fate decisions. The continued research into the crosstalk between insoluble and soluble signaling mechanisms will facilitate the goals of regenerative medicine.