Theses and Dissertations from UMD

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New submissions to the thesis/dissertation collections are added automatically as they are received from the Graduate School. Currently, the Graduate School deposits all theses and dissertations from a given semester after the official graduation date. This means that there may be up to a 4 month delay in the appearance of a give thesis/dissertation in DRUM

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Subject: search.filters.subject.Alginate

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    Electrically Induced Gelation, Rupture, and Adhesion of Polymeric Materials
    (2017) Gargava, Ankit; Raghavan, Srinivasa R; Chemical Engineering; Digital Repository at the University of Maryland; University of Maryland (College Park, Md.)
    There has been considerable interest in developing stimuli-responsive soft materials for applications in drug delivery, biosensing, and tissue engineering. A variety of stimuli have been studied so far, including temperature, pH, light, and magnetic fields. In this dissertation, we explore the use of electric fields as a stimulus for either creating new soft materials or for rupturing existing ones. Our materials are based typically on biocompatible polymers such as the polysaccharides alginate, chitosan, and agarose. We also discuss the advantages and disadvantages of electric fields over other stimuli. First, we describe the use of electric fields to form transparent and robust alginate gels around an initial mold made of agarose. Moreover, we can melt away the agarose by heat, leaving us with hollow alginate tubes. In our technique, a tubular agarose mold with dissolved calcium chloride (CaCl2) is placed in a solution of sodium alginate. A voltage of ~ 10 V is then applied, with the mold as the anode and the container as the cathode. As the Ca2+ ions migrate from the mold towards the cathode, they contact the alginate chains at the mold surface. In turn, the Ca2+ crosslinks the alginate chains into a gel, and the gel grows outward with time. The technique can be used to grow multiple layers of alginate, each with a different content, and it is also safe for encapsulation of biological species. Complex tubular structures with multiple branches and specific patterns can be created. Next, we report that electric fields can be used to rupture particles formed by ionic complexation. The particles under study are typically in the microscale (~ 200 µm radius) and are either uniformly crosslinked microbeads (e.g., alginate/Cu2+) or microcapsules formed by complexation of oppositely charged polymers (alginate and chitosan). When these particles are placed in aqueous solution and subjected to an electric field of about 10 V/cm (applied remotely, i.e., electrodes not in contact), the particles rupture within about 5 min. A possible mechanism for the electric-field-induced disruption is discussed. We also use the above particles to create electrically actuatable valves, where the flow of a liquid occurs only when the particle blocking the flow is disrupted by the field. In our final study, we show that polyelectrolyte gels and beads can be rapidly induced to adhere by an electric field. We typically work with crosslinked acrylate hydrogels made with cationic co-monomers, and anionic beads made by contacting alginate with Ca2+. When the cationic gel (connected to an anode) is contacted for just a few seconds with the anionic bead (connected to a cathode) under a voltage of ~ 10 V, the two form a strong adhesive bond. When the polarity of the electrodes is reversed, the phenomenon is reversed, i.e., the gel and bead can be easily detached. We suggest that the adhesion is due to electrophoretic migration of polyelectrolyte chains, resulting in the formation of polyion complexes. Applications of this reversible adhesion are discussed for the pick-up and drop-off of soft cargo, and for the sorting of beads.
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    New Concepts for Gelation of Alginate and its Derivatives
    (2013) Javvaji, Vishal; Raghavan, Srinivasa R; Chemical Engineering; Digital Repository at the University of Maryland; University of Maryland (College Park, Md.)
    Bioengineering applications require materials that offer tunable and precise control over material properties. In particular, hydrogels of the polysaccharide, alginate have been widely studied for applications such as drug-delivery vehicles, matrices for encapsulation of cells, and scaffolds for tissue engineering. The ability of alginate to form a physically cross-linked hydrogel under mild conditions is a key factor for many applications. Traditionally, alginate gelation has been induced by the addition of divalent ions like calcium (Ca2+). In this work, we explore new ways to induce gelation of alginate or its derivatives. These new routes are of interest because they can allow researchers to circumvent current limitations and moreover they can also enable new applications. Three new concepts are explored: (1) ionic gelation activated by light; (2) ionic gelation activated by an enzyme and its substrate; (3) gelation of hydrophobically modified alginate mediated by biological cells. In our first study, we demonstrate a concept for ionic gelation of alginate in response to light, which enables us to create chemically erasable and spatially selective patterns of alginate gels. We impart light responsiveness by combining alginate, an insoluble calcium vector (e.g., CaCO3) and a light responsive component, viz. a photoacid generator (PAG). Upon UV irradiation, the PAG dissociates to release H+ ions, which react with the CaCO3 to generate free Ca2+ in-situ. In turn, the Ca2+ ions cross-link the alginate to form a gel. We show photopatterning of alginate gels, which are used to entrap contents (e.g., microparticles) and subsequently release them by a Ca2+ chelator. In our second study, we demonstrate enzymatic gelation of alginate. Here, we use an enzyme/substrate reaction to generate H+ ions. The components of our system are glucose oxidase (GOx, enzyme), glucose (substrate), alginate and CaCO3. First, GOx catalyzes oxidation of glucose to generate H+ ions. These H+ ions solubilize CaCO3 and release free Ca2+ ions in-situ. In turn, Ca2+ ions cross-link alginate chains into a gel. A sol-gel transition is observed only when GOx senses and catalyzes glucose. By exploiting the specificity of the enzyme for its substrate, we use this concept to build a visual test for the presence of glucose in an unknown product. In our final study, we induce gels by combining a hydrophobically modified (hm) derivative of alginate with biological cells. Gelation occurs due to hydrophobic interactions between the grafted hydrophobes and the bilayers of biological cells. The polymer chains thus get attached to the cells and bridge the cells into a three-dimensional network. This gelation can also be reversed (to release the cells) by addition of a supramolecule, α-cyclodextrin, which has a hydrophobic binding pocket that binds to the hydrophobes. Cell gelation by hm-alginate may be useful in cell culture and tissue engineering applications. As a step towards these potential applications, we show that the process of gelation by hm-alginate is benign to the cells.
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    Engineering Zonal Cartilage Through Utilization of a Mesenchymal Stem Cell Population
    (2012) Coates, Emily Elizabeth; Fisher, John; Bioengineering; Digital Repository at the University of Maryland; University of Maryland (College Park, Md.)
    Articular cartilage has a limited ability to repair itself after damage due to injury or disease. Regenerative therapies using chondrocytes, the primary cartilage cell population, result in poor quality repair tissue and often cause further damage at the donor site. Furthermore, there are no current therapies which aim to regenerate the zonal organization and function of the tissue. In an effort to address both cell source limitations and zonal tissue regeneration the goal of the presented work was to utilize a mesenchymal stem cell (MSC) population to generate abundant numbers of chondrocytes with zonal phenotypes. To this end, zonal subpopulations of articular chondrocytes were isolated, characterized for differences in gene and protein expression, and exposed to scaffold environments designed to aid in phenotype retention. From these results, and reports in the literature, it was clear a major functional difference between zones was the production of a lubricating protein, proteoglycan 4 (PRG4), in the superficial zone only. Middle and deep zone cells were found to be phenotypically similar and distinct from superficial zone cells. It was further found that gene expression of PRG4 by superficial zone cells in alginate culture can be significantly enhanced by incorporation of matrix molecules hyaluronic acid (HA) and chondroitin sulfate (CS) to the scaffold environment. HA and CS also had favorable effects on MSC chondrogenesis by upregulating chondrogenic transcription factor Sox9 gene expression, and downregulating type I collagen (fibroblastic marker) gene expression. The potential of soluble signals derived from zonal (superficial or middle/deep) cartilage explants to drive MSC chondrogenesis was also investigated. Results show that signals derived from cartilage explants can induce chondrogenesis to varying degrees, with superficial zone explants inducing robust and sustained differentiation. This differentiation was found to be dependent on the proximity of the MSCs and tissue explants, implying that communication between MSCs and chondrocytes is necessary for chondrogenic induction. Coculture with superficial zone explants also upregulated MSC gene expression of PRG4. This research highlights the important functional differences between zonal chondrocyte populations and identifies MSCs as a progenitor population capable of differentiating into zone-specific chondrocytes.
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    TUBULAR PERFUSION SYSTEM BIOREACTOR FOR THE DYNAMIC CULTURE OF HUMAN MESENCHYMAL STEM CELLS
    (2012) Yeatts, Andrew Bryan; Fisher, John P; Bioengineering; Digital Repository at the University of Maryland; University of Maryland (College Park, Md.)
    In vitro culture techniques must be improved in order to increase the feasibility of cell based tissue engineering strategies. Limitations of current techniques are largely a result of the slow diffusion of molecules such as oxygen into the interior of three dimensional scaffolds in static culture. In order to enhance nutrient transport we have developed a novel bioreactor, the tubular perfusion system (TPS), to culture human mesenchymal stem cells (hMSCs) in three dimensional scaffolds. In our design, hMSCs are cultured on scaffolds tightly packed in a tubular growth chamber. Media is perfused by a peristaltic pump through the growth chamber and around the tightly packed scaffolds. In the first part of the work hMSCs are encapsulated in alginate scaffolds and results demonstrate bioreactor culture enhances late osteoblastic differentiation of hMSCs. An investigation into shear stress in the system revealed that osteogenic markers increase with increasing shear stress and that the differentiation of hMSCs is dependent on cell radial position within scaffolds. In order to enhance the ability to implant these constructs in vivo, a method to create an aggregated cell containing construct in vitro in a bioreactor system was developed. In this part of the work hMSCs are cultured in individual alginate beads in the TPS bioreactor and the beads are aggregated to form one large construct. Following this the TPS bioreactor was investigated to culture synthetic poly-L-lactic acid scaffolds which were fabricated using supercritical carbon dioxide gel drying. In addition to investigating the effects of perfusion on hMSC growth in these scaffolds, the effect of microporosity was investigated. In the final part of the work, a study was completed to determine how TPS culture influenced in vivo bone regeneration. Here alginate beads as well as synthetic PLGA/PCL constructs were used as scaffolds. Results revealed the efficacy of using the tubular perfusion system for bone tissue engineering and demonstrated increased bone formation as a result of hMSC implantation in both alginate and PLGA/PCL scaffolds. These studies highlighted the need for bioreactor culture in vitro as well as scaffolds to support in vivo tissue interaction.
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    Desing of Click Hydrogels for Cell Encapsulation
    (2011) Breger, Joyce; Wang, Nam Sun; Chemical Engineering; Digital Repository at the University of Maryland; University of Maryland (College Park, Md.)
    The long-term stability of ionically crosslinked alginate hinders the development of a bioartificial pancreas for the treatment of Type I Diabetes. Ionically crosslinked alginate with divalent cations is traditionally utilized to encapsulate islets of Langerhans serving as a protective barrier between the host's immune system and the donor islets of Langerhans. However, due to ion exchange with monovalent ions from the surrounding serum, alginate degrades exposing donor tissue to the host's immune system. The overall goal of this dissertation was to explore the possibility of utilizing `click' chemistry to introduce covalent crosslinking in alginate for therapeutic cell encapsulation. `Click' chemistry is customarily defined as the Cu (I) catalyzed reaction between an azide and alkyne to form a 1,2,3 triazole ring. To achieve the goal of covalently crosslinked polysaccharides, the following aims were determined: (1) synthesis and characterization of functionalized polysaccharides (alginate and/or hyaluronic acid) with alkyne or azide end groups; (2) measurement and comparison of the stability and transport properties of covalently crosslinked alginate hydrogels to that of ionically crosslinked alginate hydrogels; (3) determination of the inflammatory potential and cytotoxicity of these functionalized polysaccharides and `click' reagents by employing RAW264.7, a murine macrophage cell line under various simulated inflammatory states (with or without endotoxin, with or with out the inflammatory cytokine gamma-interferon); (4) optimization of the `click' reaction for therapeutic cell encapsulation utilizing RIN-5F, a rat insulinoma cell line, while minimizing cytotoxicity and maintaining insulin production; (5) encapsulation of primary porcine islets of Langerhans in either ionically and/or covalently crosslinked alginate capsulation and comparing insulin response to a glucose challenge. The results of these experiments demonstrate the utility of employing `click' chemistry to increase the overall stability of alginate hydrogels while maintaining therapeutic cell function.