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Synthetic Surfaces to Probe Human Pluripotent Stem Cell Fate Decisions

2015
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Release : 2015
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Book Synopsis Synthetic Surfaces to Probe Human Pluripotent Stem Cell Fate Decisions by :

Download or read book Synthetic Surfaces to Probe Human Pluripotent Stem Cell Fate Decisions written by . This book was released on 2015. Available in PDF, EPUB and Kindle. Book excerpt: 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.

Tailored Surfaces for Investigating Human Pluripotent Stem Cells

2012
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Release : 2012
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Book Synopsis Tailored Surfaces for Investigating Human Pluripotent Stem Cells by :

Download or read book Tailored Surfaces for Investigating Human Pluripotent Stem Cells written by . This book was released on 2012. Available in PDF, EPUB and Kindle. Book excerpt: Human embryonic and induced pluripotent stem cells, collectively known as hPS cells, hold great promise for the fields of regenerative medicine, developmental biology, and drug discovery. Elucidating the molecular mechanisms underlying their proliferation and differentiation is vital for realizing their potential. My research has focused on defining the molecular interactions that mediate hPS cell adhesion and self- renewal. To accomplish this goal, we assayed surfaces that mimic soluble signaling factors, mediate cell-cell interactions, or display peptides derived from extracellular matrix (ECM) proteins. I focused initially on the adhesion receptor E-cadherin, but my latter work used surfaces displaying synthetic peptides to engage other cell surface molecules. A key feature of these tailored surfaces is our ability to attribute cellular responses to the specific interactions between the cell and underlying substratum. The major accomplishment of this research was the development of a synthetic substrate for hPS cell propagation. The effective substrates displayed heparin-binding peptides that can interact with cell surface glycosaminoglycans (GAGs). My results also reveal that adhesion is not the sole prerequisite for self-renewal; integrin-binding surfaces did not maintain hPS cells in their pluripotent state. The chemically defined substratum highlights the utility of engaging cell surface GAGs and provides the foundation for investigating the influence of cellular adhesion on cell fate. To complement this work, I expanded our tailored surface strategy to target differentiated cells. In principle, surfaces modified with synthetic peptides can support adhesion by mimicking the ECM, but synthetic peptides typically lack affinity or selectivity for specific adhesion receptors. In contrast, organic chemistry can afford compounds that are highly selective. We hypothesized that selective small molecule integrin antagonists, when immobilized, would support adhesion and activate signaling. Indeed, surfaces decorated with a selective alphavbeta3 integrin-targeting molecule promoted both cellular adhesion and integrin activation. I anticipate that peptidomimetics will add to our arsenal of building blocks for generating tailored surfaces. Surfaces displaying highly selective ligands can illuminate the contributions of specific adhesion receptors to signaling cascades and perhaps even control stem cell differentiation. In conclusion, my research has afforded new, practical methods for propagating and manipulating pluripotent cells and their derivatives.

Synthetic Substrata to Instruct Human Pluripotent Stem Cell Fate

2012
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Release : 2012
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Book Synopsis Synthetic Substrata to Instruct Human Pluripotent Stem Cell Fate by :

Download or read book Synthetic Substrata to Instruct Human Pluripotent Stem Cell Fate written by . This book was released on 2012. Available in PDF, EPUB and Kindle. Book excerpt: Human pluripotent stem (hPS) cells have the remarkable capacity to self-renew indefinitely and differentiate into desired cell types. They can serve as a virtually unlimited supply of cells for applications ranging from drug screening to cell therapies to understanding human development. Reaping the promise of hPS cells hinges on effective defined culture and differentiation conditions. Efforts to generate chemically-defined environments for hPS cell propagation and directed differentiation have been hindered by access to only a handful of ligands to target hPS cells. Additionally, progress has been limited also by lack of knowledge regarding the relevant functional properties of the cell culture substratum. To address these problems, I first employed forward-chemical-genetics coupled with self-assembled monolayer technology to identify novel peptides that bind to hPS cell-surface receptors. I then developed a controlled synthesis of hydrogels with tailored peptide display and mechanical properties. This approach yielded synthetic hydrogels with specific mechanical properties that function in a defined medium to robustly support hPS cell self-renewal. Finally, by starting from molecular level understanding that matrix elasticity regulates developmental pathways, I generated a highly efficient hydrogel platform that restricts hPS cell differentiation to neurons, even without soluble inductive factors. These results indicate that insoluble cues can be important information conduits to guide hPS cell fate decisions. I envision that the blueprint provided by this work can be utilized to devise new materials to guide hPS cell fate.

A Study of the Effect of Physical Cues on Stem Cell Fate Determination

2007 Stem cells
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Release : 2007
Genre : Stem cells
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Book Synopsis A Study of the Effect of Physical Cues on Stem Cell Fate Determination by :

Download or read book A Study of the Effect of Physical Cues on Stem Cell Fate Determination written by . This book was released on 2007. Available in PDF, EPUB and Kindle. Book excerpt: Stem cell research has been fueled by increasing evidence of their great promise in clinical regenerative therapy. Conventionally, stem cell fate determination can be attributed to genetic and biochemical factors. However, the field has started to recognize the importance of the stem cell microenvironments that provide physical cues to influence cell fate decision. From a tissue engineer's point of view, introducing physical factors to the differentiation process could be an approach to direct stem cell fate. This concept is supported by a growing body of evidence showing the responsiveness of stem cells to physical stimuli. I thus develop two platforms to study the effect of (a) static environmental cues and (b) dynamic mechanical loading on stem cell fate decisions. Carbon nanotubes (CNTs), which possess relevant features such as (1) dimension analogous to that of natural extracellular matrix (ECM) molecules, (2) large surface area, and (3) ability to serve as nano-heaters to convert absorbed near-infrared (NIR) radiation into heat, were used to fabricate artificial stem cell niches. I exploit properties (1) and (2) of CNTs to make a biocompatible thin film with large surface area, to promote growth factor adsorption and preferential stem cell differentiation. Enhanced neuron differentiation from human embryonic stem cells (hESCs) was observed in poly(methacrylic acid) (PMAA)-functionalized CNT (PMAA-g-CNT) thin films. Polarized expression of motor neuron-specific marker, synapsin 1, was also detected in cells differentiatedon PMAA-g-CNT surfaces. Cells survive in this platform, with no detrimental effects observed. The improved differentiation can be attributed to the increased surface area created by the nanofibrillar structure, leading to enhanced growth factor adsorption. This is the first study to indicate that increasing surface area by use of CNT substrates leads to enhanced growth factor adsorption and stem cell differentiation. To shed light on how mechanical stimulation instructs cell fate decision, preliminary work has also been done to develop a remote-controlled nanohybrid actuator system to apply dynamic mechanical stimulation to stem cells. This is achieved by employing the thermal responsive nature of poly(N-isopropylacrylamide) (PNIPAM), and the unique ability of CNTs to absorb NIR. The actuation of PNIPAM hydrogel can be triggered by temperature change, while the cells on the PNIPAM actuator change in cell shape and size upon sensing the mechanical stimulation. CNTs embedded in the polymer matrix convert photon energy into heat and initiate the contraction of PNIPAM gel. The novel device can sense NIR inputs and showed noticeable shrinkage after NIR stimulation. It can therefore be used to apply remotely controlled mechanical loads to stem cells for cell behavior study. Cytosolic calcium fluctuations, which play an important role in cell differentiation and are sensitive to mechanical stimulation, may thus be tuned by using the novel actuator to achieve controlled cell differentiation. My work described above paves a way for further studies to investigate the effect of mechanical inputs on stem cell fate decision.

Control of Human Pluripotent Stem Cell Fate by Engineering Signaling Pathways

2012
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Release : 2012
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Book Synopsis Control of Human Pluripotent Stem Cell Fate by Engineering Signaling Pathways by :

Download or read book Control of Human Pluripotent Stem Cell Fate by Engineering Signaling Pathways written by . This book was released on 2012. Available in PDF, EPUB and Kindle. Book excerpt:

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