Andrés García, PhD (Georgia Institute of Technology)

Co Advisor:

Robert Guldberg, PhD (U of Oregon)

Committee:

Susan Thomas, PhD (Georgia Institute of Technology)

Nick Willett, PhD (Georgia Institute of Technology)

James Dahlman, PhD (Georgia Institute of Technology)

Designing an Injectable Tissue-Specific Small Molecule Drug Delivery System for Osteoarthritis Treatment

Osteoarthritis (OA) is a joint degenerative disease involving different processes in the articular cartilage, the synovium and subchondral bone. Currently there are no approved disease modifying drugs for OA in part because of the scarcity of appropriate drug delivery vehicles and the lack of understanding of drug effects on different intra-articular tissues. Therefore, this project aims to (i) develop poly(ethylene glycol) (PEG) microgels containing drug-loaded poly(lactic-co-glycolic) (PLGA) nanoparticles for the sustained release of small molecule drugs, (ii) to achieve specific cartilage and synovium PEG microgels binding though their functionalization with targeting peptides and (iii) to evaluate the safety and targeting efficiency of the peptide-functionalized PEG microgels in a rat model of OA. This work will provide a platform for the controlled release of small molecule drugs able to target tissues involved in OA progression such as articular cartilage and synovium. Also, considering that PEG microgels could also be used for the encapsulation of proteins and cells, this work may provide a better tool to study the effects of a combination of therapeutic strategies (small molecules, proteins and cells) on specific joint tissues. This work is expected to contribute to the screening of therapy strategies and the development of a tissue-specific treatment to prevent OA progression.

Committee:

Joseph Manns, Ph.D. (Emory University)

Christopher Rozell, Ph.D. (Georgia Institute of Technology)

Samuel Sober, Ph.D. (Emory University)

Garrett Stanley, Ph.D. (Georgia Institute of Technology)

LEARNING INFANT ASSOCIATED SOUNDS: A BEHAVIORAL PARADIGM TO INVESTIGATE SENSORY PLASTICITY IN A SOCIAL CONTEXT

Auditory cortical representations are shaped by diverse and complex experience dependent factors. In the pursuit to understand how these dynamic representations influence social behavior, auditory cortical researchers are turning to ethological paradigms whereby rodents form auditory associations though social interactions. One such ethological paradigm is the maternal mouse communication model, which has been used to investigate plasticity in auditory cortical representations of pup ultrasonic vocalizations. However, questions have recently emerged regarding the degree for which these representations are in fact shaped by social experience. It appears that we now need the ability to ``pair'' pup interaction with a novel sound that is under experimenter control. Additionally, this method of pairing needs to be done in such a way to allow for the characterization of auditory cortical activity. The development and validation of such a behavioral paradigm is the central aim of this thesis. We have developed a pairing paradigm where we use a novel sound to guide mice to a target arm at the end of a maze where they receive a pup reward. We found that early on in training mice follow a strategy that is not random but is based on returning to the last arm they received a pup. Over training mice shift from using this initial location-based strategy to using an auditory one where they use the delivered sound to seek out and retrieve pups. By silencing auditory cortical activity in mice after they had been conditioned to approach a novel sound for pup reward, we demonstrated that performance on the task significantly drops and that the mice become more likely to use their initial location-based strategy. Finally, we found that mother mice can learn this task faster than cocaring mice. From these results, the paradigm we have developed looks to be a valuable tool for investigating how auditory cortical representations can influence behavior in social contexts.

Mark Prausnitz, Ph.D. (Georgia Institute of Technology - CHBE) - Chair

Sven Behrens Ph.D. (Georgia Institute of Technology - CHBE)

Julie Champion Ph.D. (Georgia Institute of Technology - CHBE)

Saad Bhamla Ph.D. (Georgia Institute of Technology - CHBE)

M.G. Finn Ph.D. (Georgia Institute of Technology - CHEM)

"High-Throughput Synthesis of Liquid Salts for Enhanced Transdermal Drug Delivery"

The field of room temperature ionic liquids (ILs) and deep eutectic solvents (DESs), which are together referred to as liquid salts (LS), has emerged as a promising approach for the development of novel drug administration. Ionic liquids consist of bulky cation-anion combinations which render the salt unable to fuse to a solid state at room temperature. Deep eutectics consist of a mixture of anionic/cationic species and a hydrogen bond donor (HBD) which form a liquid at a temperature below the individual melting temperature of the constituent species. DESs are usually obtained by the combination of a quaternary ammonium salt with a metal salt or HBD. Both ILs and DESs have been shown to have interesting solvent properties, and could potentially replace the need for solvents in drug formulations by incorporating the active pharmaceutical ingredient (API) as a counter ion. Because the API is incorporated at higher concentrations than a typical drug solution (50% in the case of ILs), formulating hard-to-dissolve drugs as liquid salts could improve their pharmacokinetics, and the tunability associated with ion selection can aid in minimizing potential toxicity. Currently, the development of novel LS is limited by significant trial and error in the screening of suitable counterions. In this thesis, we will develop an automated high-throughput method for the synthesis of LS to study ion pair combinations that incorporate pharmaceutically relevant species and study counterion influence over their synthetic outcome. This project aims to characterize the physicochemical properties of the developed LSs and use these insights to generate guidelines for future LS design. Furthermore, we will evaluate the potential of the synthesized LSs for transdermal applications and evaluate a lead LS formulation in-vivo as proof of concept. This work has the potential to enhance the bioactivity of previously problematic drug candidates, and expand our knowledge on biocompatible LS design.

BioE Ph.D. Defense Presentation

Monday, May 7th, 2018, 12:00pm

IBB Building, Room 1128

Advisor: 

Susan N. Thomas, Ph.D. (School of Mechanical Engineering, Georgia Institute of Technology)

Committee:

Brandon Dixon, Ph.D. (School of Mechanical Engineering, Georgia Institute of Technology)

Valeria Milam, Ph.D. (School of Materials Science, Georgia Institute of Technology)

Michael Davis, Ph.D. (Department of Biomedical Engineering, Georgia Institute of Technology)

M.G. Finn, Ph.D. (School of Chemistry, Georgia Institute of Technology)

ENGINEERED BIOMATERIAL DRUG DELIVERY SYSTEMS FOR ENHANCED DELIVERY TO LYMPH NODES

State-of-the-art drug delivery currently focuses on delivery vehicle size, surface chemistry, and/or receptor interactions, all with the hope of improving drug accumulation within the tissue target. This work seeks to alter this paradigm by recognizing that tissue targets are not black boxes to which simply achieving accumulation is sufficient, but instead, complex microenvironments that house the cells that actually produce the function of the tissue and are the real targets of drug delivery. For example, while tissues critically involved in the regulation of immune processes, such as the lymphatics and lymph nodes, possess delivery barriers to getting drugs to their anatomical location, due to the complex spatial and temporal regulations of adaptive immune responses, these tissues, more importantly, possess delivery barriers to specific cells within them that must be overcome to achieve the desired immune response. The main innovation of this work, therefore, is that it addresses all drug delivery barriers, from site of injection to site of action, for the lymphatics and lymph nodes. This work has produced two novel nanoparticle-based delivery systems: one which proposes nitric oxide as a therapeutic for lymphatic-related therapies including direct delivery of nitric oxide to the lymphatics to regulate pumping function, and delivery of nitric oxide to lymph node-resident antigen presenting cells to increasing nanoparticle uptake as well as possibly promote tolerance; and the second which proposes a novel mechanism for the delivery of small molecules to deep lymph node cells for enhanced immune responses. Both of these systems while being nanoparticle-based, however, have engineered through either endogenous or synthetic chemistry the ability to release their small molecule payload, and thus are the first multi-stage lymphatic and lymph node drug delivery systems.

Committee:

Cyrus K. Aidun, PhD (Georgia Institute of Technology)

C. Ross Ethier, PhD (Georgia Institute of Technology)

Shannon L. Meeks, MD (Emory University)

Susan N. Thomas, PhD (Georgia Institute of Technology)

High Shear Arterial Thrombosis: Diagnosis and Therapy

Atherothrombosis is a common event in myocardial infarction and stroke. In the critical care of patients, it is vital to properly diagnose and provide appropriate therapy. An ideal diagnostic tool would be able to determine patient-specific drug regimens; however, there currently does not exist a reliable tool to do so. Current platelet function tests do not have relevant fluid mechanics and, along with current microfluidic models, have variable diagnostic end-points. Therefore, there is still a need for a diagnostic device.

Antiplatelet therapies, such as aspirin and Plavix, have been developed to irreversibly inhibit platelet activation or binding. However, both do not work as intended for a large percentage of the population, as 5-60% of patients exhibit resistance to therapy. Resistance persists even with dual antiplatelet therapy (DAPT). The therapeutic resistance and bleeding risks indicate the need for both a diagnostic device and improved antithrombotic agents.

The overall goal of this proposal is to develop a low variability device for diagnostic and research use. I hypothesize that the main sources of variability observed in previous microfluidic assays are due to three fundamental design factors. Further, I hypothesize that the end-point of occlusion time will provide diagnostic utility for individual antiplatelet responsiveness. Finally, I hypothesize that such a device can be utilized to develop antithrombotic nanoparticle therapies.

BioE M.S. Thesis Defense

Tuesday, March 27th, 2018

12:00 pm, 2110 Whitaker

Advisor:

Robert E. Gross, Ph.D., M.D. (Department of Neurosurgery, Emory University)

Committee:

Lily S. Cheung, Ph.D. (ChBE, Georgia Institute of Technology)

Kostas Konstantinidis, Ph.D. (CEE, Georgia Institute of Technology)

Bioluminescence Calcium Indicator Protein and Gold Nanoparticle-enhanced Bioluminescence

Bioluminescent proteins have been extensively used as a light emission source of many fusion proteins and have a wide range of application in bio-imaging and cell signaling studies. In our study, we focused on Gaussia luciferase (GLuc) from the marine copepod Gaussia princeps, which is the smallest known luciferase. The first part, we developed two ultrasensitive calcium sensing fusion proteins composed of a variant of GLuc, "slow-burn" Gluc (sbGLuc) and the calcium-dependent fluorescent sensor protein, GCaMP6s. Our design exhibited excellent sensitivity at physiological calcium concentration and can be explained by bioluminescence resonance energy transfer (BRET). The second part, we represented a bioluminescence enhancement strategy, which conjugates Ni-NTA (nitrilotriacetic acid) coated gold nanoparticle with 6 histidine tagged sbGLuc. One version exhibited 75% enhancement of bioluminescence compared with the non-treated sbGLuc. The new sensors together with the gold nanoparticle-enhanced bioluminescence should be useful for various studies in various fields such as neuroscience and cell biology.

Committee:

Johnna S. Temenoff, Ph.D. (Georgia Institute of Technology)

Krishnendu Roy, Ph.D. (Georgia Institute of Technology)

Greg Gibson, Ph.D. (Georgia Institute of Technology)

Steven L. Stice, Ph.D. (University of Georgia)

Patient-specific Approaches to Bone Regeneration

Bone is the second-most transplanted tissue after blood with more than 1.6 million bone grafting procedures performed annually in the US at a cost of over 5 billion dollars. Treatment of large bone defects in particular remains one of the most challenging problems faced by orthopedic surgeons. Current therapies include bone grafts and/or delivery of osteoinductive proteins such as bone morphogenetic protein 2 (BMP-2). Despite advances in surgical technique and medical care, many of these treatment options still exhibit high variability in healing, suggesting that patient-specific factors, such as age, gender, treatment timing, and immune status, may play a much more crucial role in treatment success than previously thought. Thus, the need to account for these patient-specific factors with more sophisticated treatment strategies has become increasingly apparent.

            The main objective of this work was to use preclinical animal models to investigate the influence of patient-specific factors on bone regeneration, with a particular focus on long-term immune profile characterization as it relates to the bone healing response after treatment. The impact of age and dose on large bone defect healing was assessed using a well-established bone injury rat model along with delivery of BMP-2 in a collagen sponge, which is the current clinical standard. These results offer valuable insight on a controversial subject: the use of BMP-2 in pediatric patients. Additionally, this work sought to elucidate some of the key mechanisms that lead to impaired healing following nonunion, a significant clinical problem that still affects up to 10% of patients with long bone injuries. To accomplish this, a chronic nonunion model was established that can potentially serve as a more rigorous and clinically relevant platform for studying nonunion and testing novel therapeutics. Finally, the issue of trauma-induced immune dysregulation was evaluated in this model of nonunion as a potential harbinger of poor healing outcome. Collectively, these studies have advanced our understanding of the factors that affect bone regeneration and represent a pivotal step towards improved, more personalized treatment strategies for bone repair.

Committee:

Andreas Bommarius, Ph.D. (Georgia Institute of Technology)

Michael Leamy, Ph.D. (Georgia Institute of Technology)

Pamela Peralta-Yahya, Ph.D. (Georgia Institute of Technology)

Peng Qiu, Ph.D. (Georgia Institute of Technology)

From Experimental Observations to a Functional Model of the Lignin Pathway: Computational Modeling Reveals New Insights.

Lignin is a natural polymer that is interwoven with cellulose and hemicellulose within plant cell walls. Due to this molecular arrangement, lignin is a major contributor to the recalcitrance of plant materials with respect to the extraction of sugars and their fermentation into ethanol, butanol, and other potential bioenergy crops. The lignin biosynthetic pathway is similar, but not identical in different plant species. It is in each case comprised of a moderate number of enzymatic steps, but its responses to manipulations, such as gene knock-downs, are complicated by the fact that several of the key enzymes are involved in several reaction steps. This feature poses a challenge to bioenergy production, as it renders it difficult to select the most promising combinations of genetic manipulations for the optimization of lignin composition and amount. Moreover, species specific regulatory features and distinct spatial and topological characteristics hinder accuracy of a unified lignin pathway model. In this dissertation a systems biology approach is used to address these challenges by means of computational modeling. Novel mathematical techniques are employed on different types of experimental data in situ, and shed light on complexities of lignin biosynthesis pathway. The developed methods are nevertheless general enough to be used in a wide range of metabolic modeling applications.

Todd Sulchek, PhD

Georgia Institute of Technology, Woodruff School of Mechanical Engineering

Thesis Committee:

Dr. Craig R. Forest, PhD
Georgia Institute of Technology, Woodruff School of Mechanical Engineering

Dr. Wilbur A. Lam, M.D. Ph.D
Georgia Institute of Technology and Emory University, Whitaker Biomedical Engineering

Dr. Manu Platt, Ph.D.

Georgia Institute of Technology and Emory University, Whitaker Biomedical Engineering

Dr. A. Fatih Sarioglu, Ph.D.

Georgia Institute of Technology, Electrical and Computer Engineering

Pulse Width Modulated Periodic Backflush to Improve Dead-End Filtration for Label-Free, Size-Based Sorting

Although label-free dead-end filtration cell sorting offers many advantages over other labelled and label-free approaches, it still falls short when comparing recovery percentage in most cases. Regardless, dead-end filters are used ubiquitously in clinical setting for diagnostic and therapeutic applications ranging from tissue engineering to infectious agent identification and stratification. Flow control has been shown to improve filter flux capacity over time, but no comprehensive breakdown of its waveform analysis has been performed and applied across the board. The central hypothesis of the proposed research is that pulse width modulation of fluid velocity can be used to periodically backflush dead-end membranes to interrupt cake formation, reintegrate fouling layers into the bulk of a sample, and improve permeate flux and recovery percentage. This will inform the design of fluid control dynamics for diagnostic as well as therapeutic application in Cystic Fibrosis, blood fractionation, and scaffold seeding. The effects of varying duty cycle to improve targeted permeate recovery percentage or retentate purity will be assessed through these applications.

Vasilis Babaliaros, MD (Emory University, Medicine)

Rahul Sharma, MD (Cedars Sinai Medical Center, Los Angeles, CA)

Wei Sun, PhD (Georgia Tech, Biomedical Engineering)

John Oshinski, PhD (Georgia Tech, Biomedical Engineering)

Cyrus Aidun, PhD (Georgia Tech, Mechanical Engineering)

A Comprehensive Analysis of Potential Factors for Transcatheter Aortic Valve Thrombosis Risk

Transcatheter aortic valve replacement (TAVR) is indicated for aortic stenosis (AS) patients who are deemed intermediate or greater surgical risk. Recent evidence of leaflet thrombosis and reduced leaflet mobility in TAVR devices has led to concerns of stroke and long-term valve durability. Risk factors for thrombosis in TAVR patients remain poorly defined. While materials and blood chemistry are likely to be contributing factors to thrombosis risk, early clinical evidence and experimental data suggest that the fluid dynamic environment in the specific setting of the transcatheter aortic valve (TAV) is a major factor in the development of leaflet thrombosis. This environment can be altered by anatomical, procedural, and device related parameters. Additionally, it has been demonstrated that the thrombus originates in the “neo-sinus,” which is the pocket formed between the TAV leaflets and the native aortic valve leaflets. This study aims to elucidate the contribution of anatomical, deployment, and fluid dynamic factors to a thrombogenic environment in the neo-sinus region of transcatheter aortic valves via analyses of clinical imaging data, the development of predictive statistical models, and a battery of well controlled benchtop experiments on select commercial TAVs. The findings from these studies could provide a vital tool to help predict which valves, deployments, and/or patient may be at a higher risk of valve thrombosis.

Susan N. Thomas, PhD

Georgia Institute of Technology, Woodruff School of Mechanical Engineering

Committee:

Andrés García, PhD Georgia Institute of Technology, Woodruff School of Mechanical Engineering

Wilbur Lam, PhD Georgia Institute of Technology, Coulter Department of Biomedical Engineering

John McDonald, PhD Georgia Institute of Technology, School of Biological Sciences

Cheng Zhu, PhD Georgia Institute of Technology, Coulter Department of Biomedical Engineering

Elucidating the contributions of cellular and microenvironment characteristics on cell adhesion processes in flow

Circulating cell recruitment is critical to a variety of physiological and pathophysiological processes and occurs amidst the high shear environment of the vasculature via a multistep rolling to firm adhesion cascade. Cells initially engage the vascular endothelium through interactions between endothelial-presented selectins and their corresponding ligands presented by circulating cells. These interactions precede firm adhesion and arrest and eventually transmigration across the endothelium for tissue infiltration. Since many cell subtypes including leukocytes and metastatic cancer cells employ this mechanism to facilitate their escape the vasculature, understanding differences in cell-subtype adhesive behavior can inform the development of targeted therapeutics that interfere with metastatic cell transport, while leaving physiologically important immune cell recruitment mechanisms intact. As such, the overall objective of this work was to explore how selectin-mediated adhesion 1) is regulated by the biochemical and biophysical microenvironment of the vasculature and 2) varies among different cell subtypes and 3) with characteristics of circulating cells. Through the use of in vitro fluidic methodologies in conjunction with innovative single-cell analyses we have begun to elucidate key differences in the selectin-dependent adhesive behavior of metastatic versus leukocytic cells and identified potential molecular regulators of enhanced rolling adhesion behavior, which may serve as targets in the development of pharmacologic agents aimed at reducing selectin-mediated adhesion of clinically challenging cell subtypes.

Dr. J. Brandon Dixon, Chair (ME)

Dr. Babak Mehrara (Memorial Sloan Kettering Cancer Center)

Dr. Krishnendu Roy (BME)

Dr. Wei Sun (BME)

Dr. Susan Thomas (ME)

The Functional and Remodeling Response of Collecting Lymphatic Vessels To Disruption of Lymphatic Drainage Pathways

The lymphatic system is composed of a network of vessels, nodes, and accessory organs that is present in most soft tissues of the body. The lymphatics play a vital role in maintaining tissue fluid homeostasis, trafficking immune cells from the periphery to the nodes, and transporting dietary lipids from the intestines to the blood stream. Unlike the blood vasculature, the lymphatic system does not have a central pump. Instead, transport is promoted by collecting lymphatic vessels that are composed of a series of contractile segments separated by one-way valves. When the normal function of the lymphatics is compromised a disease called lymphedema may develop, which is characterized by tissue fluid retention, fibrosis, and adipose accumulation. Unfortunately, lymphedema is a relatively common complication of cancer therapies that damage the lymphatic vasculature, such as lymph node dissections and radiation treatment.

Despite being integral driver of lymphatic transport, relatively little is known about how collecting lymphatic function and remodeling may influence the development of lymphedema. This work demonstrates the development of novel near-infrared imaging methods with the ability to quantify and phenotype collecting lymphatic failure during lymphatic disease. These methods provide biological insight into the functional and remodeling response of the collecting lymphatic vessels to surgical disruption of lymphatic drainage pathways. Specifically, we demonstrate that diet-induced obesity adversely impacts collecting lymphatic contractility and pump function during lymphedema in a mouse model. Further, in a clinically relevant sheep model, we demonstrate that the uninjured vessel can compensate in vivo by altering its intrinsic functional response and structure; however, through this process, the collecting lymphatic muscle experiences increased oxidative stress due to increased contractility. The results of this work demonstrate that functional adaptations of the collecting lymphatic vessels may influence the development of lymphedema.

Todd Sulchek, PhD

Georgia Institute of Technology, Woodruff School of Mechanical Engineering

Thesis Committee:

Alexander Alexeev, PhD

Georgia Institute of Technology, Woodruff School of Mechanical Engineering

Edmund K. Waller, MD, PhD

Emory University School of Medicine, Department of Medical Oncology

Mark Prausnitz, PhD

Georgia Institute of Technology, School of Chemical and Biomolecular Engineering

Krishnendu Roy, PhD

Georgia Institute of Technology, Coulter Department of Biomedical Engineering

CONVECTIVE INTRACELLULAR MACROMOLECULE DELIVERY BY CELL VOLUME EXCHANGE FOR CELL ENGINEERING APPLICATIONS

The rapidly growing field of cell manufacturing requires robust methods for intracellular delivery of cell engineering reagents. However, this field still lacks an intracellular delivery platform that is cost-effective, maintains high cell viability, and is broadly applicable for diverse cargoes and cell types. In this project, we discovered a unique biophysical phenomenon of transient cell volume exchange induced by rapid and sequential microfluidic cell compression. This behavior consists of iterations of brief, mechanically induced cell volume loss followed by rapid volume recovery. We found that cell volume exchange can convectively transfer a variety of macromolecules and particles into cells, including 2 MDa polysaccharides, 100 nm nanoparticles, and 2.5 MDa plasmids, without significantly impacting cell viability or other tested phenotypic properties. The ease of use and successful proof-of-concept transfections demonstrate great potential to address major challenges in intracellular delivery. The work first aims to understand the mechanisms of this technology, and then leverage the platform to impactfully address two growing needs in cell engineering: (1) test and evaluate a new immunotherapy manufacturing approach by delivering CD19 chimeric antigen receptor (CAR) mRNA to primary T cells and assessing CAR T cell expression and cytotoxicity against B cell lymphoma; (2) demonstrate induced pluripotent stem cell (iPSC) reprogramming by direct delivery of Yamanaka factor protein or mRNA.

C. Ross Ethier, PhD, Co-Chair (Wallace H. Coulter Department of Biomedical Engineering, Georgia Institute of Technology and Emory University)

Machelle Pardue, PhD, Co-Chair (Wallace H. Coulter Department of Biomedical Engineering, Georgia Institute of Technology and Emory University)

Brandon Dixon, PhD (George W. Woodruff School of Mechanical Engineering, Georgia Institute of Technology)

Mark Prausnitz, PhD (Chemical and Biomolecular Engineering, Georgia Institute of Technology)

Brian Samuels, MD, PhD (Department of Ophthalmology, University of Alabama, Birmingham)

Stiffening the Posterior Rat Sclera to Provide Neuroprotection in Glaucoma

Glaucoma is the leading cause of irreversible blindness in the world, expected to affect approximately 80 million people by the year 2020. This degenerative optic neuropathy is characterized by retinal ganglion cell (RGC) death, optic nerve damage, and progressive vision loss. While the exact etiology remains elusive, elevated intraocular pressure (IOP) is a known risk factor and lowering IOP remains the only effective treatment. Elevated IOP causes deformation and remodeling of the optic nerve head (ONH) tissues, which in turn is thought to lead to localized neurodegeneration. Computational and ex vivo studies have shown that scleral stiffness strongly influences deformation of the ONH, and that increasing the stiffness of the portion of the sclera surrounding the ONH (the peripapillary sclera) can significantly reduce these excessive strains. We hypothesize that by crosslinking the collagenous peripapillary sclera, we will reduce mechanical deformation in the ONH, which will in turn provide neuroprotection to mitigate glaucomatous vision loss. To investigate this hypothesis, we will develop and evaluate the efficacy of a nontoxic scleral stiffening treatment in a commonly-used rat model of glaucoma, which will lay the foundation for future glaucoma therapies.

BioE PhD Defense Presentation

Date: Wednesday, November 1^st, 2017

Time: 10:00AM

Location: Parker H. Petit Institute for BioEngineering and Biosciences-Suddath Seminar Room

Advisors:

Robert E. Guldberg, PhD, (School of Mechanical Engineering, Georgia Institute of Technology)

Todd C. McDevitt, PhD, Gladstone Institutes

Committee:

Johnna S. Temenoff, PhD (Department of Biomedical Engineering, Georgia Institute of Technology)

Krishnendu Roy, PhD (Department of Biomedical Engineering, Georgia Institute of Technology)

Tom Koob, PhD (MiMedx Group, Inc.)

ENGINEERING AN IMPROVED CARTILAGE REPAIR STRATEGY 
COMBINING CELLS & ECM-DERIVED MATERIALS

Osteoarthritis (OA) is the leading cause of disability in the United States and one in two people are expected to develop symptomatic knee OA by age 85. The avascularity, low cellularity, and slow proliferation of chondrocytes all limit the natural regenerative capacity of cartilage in addition to the inflammation prevalent in the joint space. Cell therapies, such as autologous chondrocyte implantation (ACI), offer promising options for treating persistent cartilage lesions, but the inability to expand chondrocytes to sufficient numbers without adversely affecting their phenotype remains a significant problem for graft success. ACI is not indicated for cartilage damage associated with osteoarthritis (OA) or other inflammatory diseases, however, and this lack of efficacy is attributed to the inflammatory environment cells are exposed to, since multiple inflammatory mediators have been shown to play a pivotal role in the initiation and perpetuation of OA. Anti-inflammatory therapies with single molecular inhibitors are unable to effectively modulate the complex inflammatory environment presented in OA. Thus, novel therapies that are capable of modulating multiple signaling pathways and cell types are an attractive alternative to address OA-associated inflammation.

Therefore, the objective of this proposal was to engineer an improved cartilage repair strategy by combining cells and ECM materials to address problems with both cartilage repair and OA-associated inflammation. We developed decellularized cartilage microcarriers that retain endogenous extracellular matrix proteins to both expand and deliver chondrocytes while retaining their phenotype. We also characterize the effects of aggregation, culture conditions, and donor variability on the ability of mesenchymal stem cell (MSC) immunomodulation of OA. To this end, we quantified MSC paracrine factor production, suppression of activated synoviocyte inflammation, and therapeutic efficacy in the rat medial meniscal transection (MMT) rat model of OA. Furthermore, we investigated the interaction between MSCs and human amniotic membrane and the influence of cell-cell and cell-ECM therein on the modulation of inflammation, both in vitro and in vivo. Overall, this work broadens current understanding of cartilage tissue engineering and immunomodulation via ECM and stem cell-based therapies, providing valuable information that can be used to develop strategies to improve efficacy of osteoarthritis treatments. 

Karl I. Jacob (Georgia Institute of Technology, School of Materials Science and Engineering)

Committee

Luke Brewster, MD, PhD (Emory University School of Medicine)

J. Brandon Dixon, PhD (Georgia Institute of Technology, George Woodruff School of Mechanical Engineering)

Mostafa A. El-Sayed, PhD (Georgia Institute of Technology, School of Chemistry & Biochemistry)

Krishnendu Roy, PhD (Georgia Institute of Technology, Wallace H. Coulter Department of Biomedical Engineering)

Microfluidic generation of cancer nanomedicines

Cancer diagnosis and therapy are perhaps the most promising areas for nanotechnology in medicine, and are expected to soon have applications in the market. The goal of this dissertation was to develop a technological foundation for synthesis and evaluation of polymer-based cancer therapeutic. This was performed using a microfluidic platform for optimization, and characterization of resulted particles for controlled drug release within the tumor environment. This technique was first optimized to show the feasibility of making drug-nanoparticles (NPs) out of different synthetic and natural polymers with it. The biophysical properties of these particles were also investigated at nano-biointerface. The process was then adjusted to develop pH-responsive core-shell NPs enabling oral administration of hydrophobic cancer therapeutics. This technique was also adjusted to fabricate complex NPs via controlled self-assembly of several components in a single step. Resulted particles can be used for theranostics applications or provide ultra-high drug loading capacity. Tuning the surface properties was also possible via this system and it was used to control immune-NPs and cancer cell-NPs interactions. To prolong blood circulation and enhance cell internalization, feasibility of making one-dimensional nanocarriers by template-based self-assembly approach was also confirmed. These nanocarriers can serve as suitable candidates for combinatorial cancer therapy as they can load and deliver substantial amounts of drugs while allowing for hyperthermia effect thanks to their carbon nanotube core. Mechanical properties of nanocarriers can also influence a broad range of NPs’ biological behaviors. Here, inspired by viruses, systematic investigation of the mechanobiological properties of NPs are done to determine the optimized range for in vitro and in vivo targeting. NPs with switchable mechanical properties are proposed capable of switching from soft to stiff state in the site of action and provide enhanced therapeutic efficiencies. Overall, we hope that this research provides broad information on how NP design can affect and control the efficacy of cancer nanomedicine. These findings point to the high potential of microfluidic platforms as engineering toolboxes that enable design of complex multifunctional nanomaterials via controlled bottom-up approach for various biomedical applications.

Committee:

Michael Davis, Ph.D. (Georgia Institute of Technology & Emory University)

Charles Searles Jr., M.D. (Emory School of Medicine)                                                  

Loren Williams, Ph.D. (Georgia Institute of Technology)                                                   

Younan Xia, Ph.D. (Georgia Institute of Technology)                                                  

miR-744 Modulation by Disturbed Flow and its Role in Endothelial Inflammation and Atherosclerosis

 Atherosclerosis is the leading cause of death in developed nations as it is the underlying cause of many cardiovascular diseases (CVD) such as myocardial infarction, ischemic stroke, and peripheral arterial disease. Atherosclerotic plaques preferentially develop in areas with curved or branched geometries due to the effects of low magnitude, oscillating, disturbed blood flow (d-flow) on the endothelium. The mechanisms by which d-flow induces pro-atherogenic responses predominantly involves changes in the endothelial gene expression, in part due to differential microRNA (miRNA) expression. Here, we report the identification of a novel, flow-sensitive miR-744 in endothelial cells that stimulates endothelial inflammation in vitro. Furthermore, we found LIMS2 is a novel, mechanosensitive, conserved target of miR-744. Finally, inhibition of the specific interaction of miR-744 and LIMS2 by target site blockers significantly reduced the development of plaque in a d-flow-induced murine model of atherosclerosis. The work presented here has resulted in the discovery of a novel, atherogenic miRNA, a novel, atheroprotective gene, and underscores the importance of the specificity of the miRNA-gene interaction. This work also provides a foundation for future studies to develop more targeted therapeutic strategies for CVD.

Committee:

               Rudolph “Rudy” Gleason, Ph.D. (Georgia Institute of Technology)

               Vasilis Babaliaros, M.D. (Emory University School of Medicine)

Impact of Simulated MitraClip on Forward Flow Obstruction in the Setting of Mitral Leaflet Tethering

Mitral regurgitation (MR) is the most common valvular disease; significant levels of MR are found in 6.4% of the population between 65 and 74 years of age, and 9.3% of the population over 75 years of age. Surgical repair or replacement of the mitral valve (MV) is the current gold-standard for treatment of MR, and requires open-heart surgery. Many patients with MR in the older population subgroups have co-morbidities and are considered high risk for open heart surgery. In a recent study, among patients with severe symptomatic MR, nearly 50% were denied surgery based on comorbidities. The need for solutions to treat these high-risk patients has inspired a host of new minimally invasive endovascular repair devices. The catheter-based MitraClip device (Abbott Vascular, Santa Clara, CA) is currently the only widely-used and FDA approved endovascular mitral repair device, with over 35,000 patients treated worldwide to-date. MitraClip works by clipping the mitral leaflets together at the site of MR. This closes the regurgitant orifice, while still allowing the leaflets to open to either side of the device, thereby creating a “double orifice”.

Although the device is widely used, it is not without flaw. A fundamental issue with MitraClip is the risk of obstructing forward flow of blood into the left ventricle (LV). By clipping the mitral leaflets together, the MitraClip inherently reduces the MV orifice area, and, in certain cases, can elevate the diastolic pressure gradient across the valve. This is known as Mitral Stenosis (MS), and can lead to increased mortality in patients. Recent limited-size studies have shown that risk of creating MS can be greater in certain patients with restricted mitral leaflet motion, or leaflet tethering. Leaflet tethering is often seen in Functional Mitral Regurgitation (FMR), where underlying disease and remodeling of the LV causes papillary muscle displacement and increased tension on the chordae and leaflets. MitraClip is currently not indicated for use in this large population of MR patients, but ongoing large-scale clinical studies are seeking to determine its safety and effectiveness in FMR.

The purpose of this study is to quantify MS severity under different levels of leaflet tethering, and with placement of MitraClip in different locations. In the first specific aim of this study, an in vitro model capable of simulating MV function and leaflet tethering on excised ovine MV samples was developed, along with custom-designed and machined MitraClip prototypes. In the second specific aim, these tools were used to evaluate the forward flow performance of the MitraClip prototype devices in the setting of mitral leaflet tethering. This first-of-its-kind study will provide valuable information on MitraClip performance, and aid cardiologists in determining the suitability of different MitraClip treatments for the large population of FMR patients.

Advisor:

Stephen H. Sprigle, PhD, PT (School of Mechanical Engineering, Georgia Institute of Technology)

Committee:

Aldo A. Ferri, PhD (School of Mechanical Engineering, Georgia Institute of Technology)
Jun Ueda, PhD (School of Mechanical Engineering, Georgia Institute of Technology)
Young-Hui Chang, PhD (School of Biological Sciences, Georgia Institute of Technology)

Maysam Ghovanloo, PhD (School of Electrical and Computer Engineering, Georgia Institute of Technology)

Mark Greig (Vice President of R&D Engineering, Sunrise Medical LLC)

DEVELOPMENT OF COMPONENT AND SYSTEM-LEVEL TEST METHODS TO CHARACTERIZE MANUAL WHEELCHAIR PROPULSION COST

The current approach to manual wheelchair design lacks a sound and objective connection to metrics for wheelchair performance.  Wheelchair performance directly impacts propulsion effort, which is a strong determinant of user health and mobility.  The objective of this thesis is three-fold: 1) to characterize the inertial and resistive properties of different wheelchair components and configurations, 2) to characterize the systems-level wheelchair propulsion cost, and 3) to model wheelchair propulsion cost as a function of measured component and configuration properties.  To this end, this defense presents the development of 1) a series of instruments and methodologies to evaluate the rotational inertia, rolling resistance, and scrub torque of wheelchair casters and drive wheels on various surface types, and 2) a wheelchair-propelling robot capable of measuring propulsion cost across a collection of maneuvers representative of everyday wheelchair mobility.  Using this collection of devices, I demonstrate the variance manifested in the resistive properties of 8 casters and 4 drive wheels, and the impact of these components (as well as mass and weight distribution) on system-level wheelchair propulsion cost.  Coupling these findings with a theoretical framework describing wheelchair dynamics, I define two empirical models linking system propulsion cost to component resistive properties.  The outcomes of this research empower clinicians and users to make a more informed choice in wheelchair selection by means of a standard, scientifically-motivated performance metric.  Furthermore, the empirical models offer manufacturers a basis by which to optimize their future wheelchair designs, thus motivating a better product for all wheelchair stakeholders.

Dr. Cyrus Aidun (Advisor)

Dr. Gilda Barabino(Advisor)

Dr. Edward Botchwey

Dr. Brandon Dixon

Dr. Wilbur Lam

Title: MICROFLUIDIC DEVICES FOR STIFFNESS DEPENDENT ENRICHMENT OF

RED BLOOD CELL SUBPOPULATION

Red blood cells being the most dominant cell type of blood are often the target of many

hematologic diseases such as sickle cell disease, malaria, spherocytosis and some types of

cancers. In addition to affecting biological properties, these diseases also alter biomechanical

properties such as morphology, size and stiffness of red blood cells. Separating or enriching

the cellular components of blood into subpopulation based on their bio-mechanical

properties and analyzing them have the potential to lead to enhanced strategies for assessment

and treatment of these diseases. Current techniques and equipment for diseased cell

sample enrichment are time consuming, expensive and need well trained professionals to

be conducted. Microfluidic platform based red blood cell enrichment device is one of the

most promising technologies that are currently the subject of considerable interest among

researchers because of its low cost, high throughput, easy operation and the potential to

do enrichment within the physiological flow condition. In this research work, microfluidic

devices were designed, fabricated and tested for enriching red blood cell subpopulations

based on their stiffness from a mixture of stiff and normal red blood cells. In the first portion

of the work, lab developed numerical simulation tools were deployed to study stiffness

dependent margination pattern of red blood cells in high aspect ratio straight microchannels

with rectangular cross-section. Stiff red blood cells were observed to marginate near the

channel walls whereas normal (and hence more deformable) red blood cells were observed

to marginate around the center line of the channel regardless whether cell-cell interaction

was significant or not. Cells of different stiffness reached to their equilibrium locations

faster in channels with smaller cross sections. Increasing flow Reynolds number and hence

the flow rate resulted in stronger segregation between normal and stiff red blood cells for

the whole range of Reynolds numbers for which simulations were run. Increasing cell volume

fraction in solution also boosted separation between cells of different stiffness. Based

on the findings of the simulations, two types of cell enrichment devices were designed and

fabricated, simple straight channel device and multistep device. The simple straight channel

device was tested for a wide range of flow Reynolds number and cell volume fractions.

Simple straight channels were observed to perform better with increasing flow Reynolds

number and cell volume fraction up to certain threshold for each of them, and after that

threshold there was no significant improvement of performance. Numerical simulations

were conducted with parameters matching with some of the experiments and the results

obtained were remarkably close to those from the experiments. Statistical analysis on experimental

data found the effect of individual parameters, flow Reynolds number and cell

volume fraction, to be significant. It also revealed that there was significant interaction between

the factors flow Reynolds number and volume fraction. This implies that the extent

of the effect of one factor (e.g. flow Reynolds number) changes when the value of the

other factor (e.g. volume fraction) varies. The multistep device was also tested for different

combinations of flow Reynolds number and cell volume fraction and, was observed

to perform 1.6 times to 3.15 times better in enriching stiff cells from a mixture of stiff and

normally deformable red blood cells. To our knowledge this is the first study that incorporated

such rigorous multiphysics simulations to support experimental study on stiffness

dependent margination of red blood cells in straight micro-channels. This research work

revealed previously unreported information about stiffness dependent cell enrichment with

simple straight channel microfluidic device and proposed a new device that performed significantly

better than the simple straight channel device.

Ajit Yoganathan, PhD (Georgia Tech, BME)

Committee:

Shriprasad Deshpande, MD (Emory University, Department of Pediatrics)

J. Brandon Dixon, PhD (Georgia Tech, Mechanical Engineering)

Mark Fogel, MD (Children’s Hospital of Philadelphia, Division of Cardiology)

John Oshinski, PhD (Georgia Tech, Biomedical Engineering)

Timothy Slesnick, MD (Emory University, Department of Pediatrics)

Hemodynamic Assessment of Proposed Solutions for Fontan Failure

Congenital heart defects are the most common types of birth defects and are responsible for an estimated 300,000 newborn deaths per year. The most severe of these defects can result in a “single ventricle” physiology. Thankfully, over the last 40 years surgeons have pioneered a set of 3 staged surgeries to palliate single ventricle heart defects, which results in a total cavopulmonary connection. Short term outcomes of these “Fontan” patients are very promising, with a 1 year survival rate around 95%. However, as these patients age, long term complications are inevitable. The central purpose of this thesis is to investigate the effectiveness of current, clinically implemented “solutions” for two of the most common modes of Fontan failure including pulmonary arteriovenous malformations (PAVMs) and liver disease.  Specific Aim 1 will test if surgical planning can be used to accurately predict post-operative hepatic flow distribution (a factor in PAVM formation), and if Y-grafts can provide more balanced hepatic flow distribution than traditional Fontan connections. Specific Aim 2 will test if the extent of liver fibrosis in Fontan patients is associated with poor hemodynamics, and if ventricular assist devices can decrease Fontan hepatic congestion by augmenting flow and decreasing inferior vena cava pressure.

Robert Liu, PhD (Emory University, Biology)

Committee:

Joseph Manns, PhD (Emory University, Psychology)

Christopher Rozell, PhD (Georgia Institute of Technology, Electrical and Computer Engineering)

Sam Sober, PhD (Emory University, Biology)

Garrett Stanley, PhD (Georgia Institute of Technology, Biomedical Engineering)

 A Behavioral Paradigm for Investigating Auditory Learning in Social versus Nonsocial Contexts

 The ability to detect, discriminate, and recognize communication sounds is important for basic social interactions and is a precursor for more complex abilities such as language learning. A central interest in auditory neuroscience is to understand the mechanisms by which the brain learns to do this from experience. Recent evidence suggests that social context may play a significant role in influencing these mechanisms within auditory cortex. One proposed element of social context that may be important is the rewarding effect of interactions between individuals. However, other nonauditory and nonsocial factors that are part of the learning paradigm have also been demonstrated to influence learning mechanisms. Consequently, explicitly testing the role of social versus nonsocial reward while animals learn to perform an auditory task can be hindered by an inability to control for these other factors. The goal of this proposal is to (1) develop such a paradigm by using the reinforcing nature of mouse pups as a social reward and water as a nonsocial reward to condition mice to approach a target sound, (2) characterize the rate of learning in these tasks, and (3) test the necessity of auditory cortex in this learning.

Committee:

Brandon Dixon, Ph.D. (Georgia Institute of Technology)

Todd Sulchek, Ph.D. (Georgia Institute of Technology)

Susan Thomas, Ph.D. (Georgia Institute of Technology)

Hua Wang, M.D., Ph.D. (Emory University)

DEVELOPING MICROFLUIDIC SYSTEMS TO RESOLVE LONGSTANDING QUESTIONS in hematology

Recent research has revealed that cells dynamically sense and respond to their physical microenvironments. In hematology, it has been shown that shear mediated red blood cell (RBC) deformation results in ATP release, and that platelets attenuate contraction force based on substrate stiffness. The objective of this thesis is to create microfluidic systems in which the biophysical and biochemical aspects of hematological processes are independently investigated toward the aim of discovering new solution spaces for diagnostics and therapeutics. To that end, this defense presents novel microfluidic systems: 1) an “endothelial”-ized, T-junction fluidic to elucidate the biophysical processes that define the mechanism of action of the ferric chloride thrombosis model and 2) microfluidic devices with single-micron features (pillars and canals) to examine the effects of physical interactions between blood cells—RBCs, platelets, and neutrophils—and geometrically relevant, non-biological matrices at the single cell level. Using this suite of devices, I resolve the mechanism of action of the FeCl₃- thrombosis model, begin to characterize RBC fragmentation parameters, and give new insight into the pathological role of neutrophils in thrombosis. Microfluidic platforms allow for real-time, microscopic evaluation of cell response (via brightfield morphology and immunostaining) and precise spatiotemporal control of system inputs and flow characteristics, including shear stress. The mechanistic understandings gained by creating systems that successfully decouple the biophysical and biological aspects of blood cells, as is done in this work, can result in enhanced understanding of the etiology of pathologies, improved diagnostic assays for blood cell activity, and new targets for therapeutics. 

Committee:

Vasilis Babaliaros, M.D. (Emory University)

Gautam Kumar, M.D. (Atlanta VA Medical Center)

Wei Sun, Ph.D. (Georgia Institute of Technology)

Cyrus Aidun, Ph.D. (Georgia Institute of Technology)

A Parametric Investigation of Transcatheter Aortic Valve Replacement Performance

While transcatheter aortic valve replacement (TAVR) has many advantages over surgical replacement, it is a young technology with little evidence of long-term effectiveness. As such, there is an ever-growing need to understand factors which can lead to poor patient outcomes. This work aims to clarify which surgical and transcatheter prosthesis type, size, and deployment positions will yield the most favorable performance while minimizing adverse events. In addition, this study provides possible mechanistic explanations for the presence of TAVR leaflet thrombosis. These phenomena will be investigated through novel analyses of clinical data and innovative in vitro experimental techniques. Understanding the complex interplay between anatomical, deployment, and hemodynamic conditions in such complex scenarios will help inform clinical practice and guide the development of improved next-generation valve replacements.

MS Thesis Presentation

9 AM, Monday, May 22^nd, 2017

IBB, 1128

Advisor: Young-Hui Chang , Ph.D. (Georgia Institute of Technology)

Committee:

Daniel Goldman, Ph.D. (Georgia Institute of Technology)

Aaron Young, Ph.D. (Georgia Institute of Technology)

The Effects of Rehabilitation Interventions on Hind Limb Kinematics in a Rat Model of Osteoarthritis

Osteoarthritis (OA) is a joint disorder and the knee is the most common large joint affected by it, causing many clinical symptoms in elderly populations. Knee OA has effects on gait patterns, such as a varus rotation at the knee, and these effects can change depending on the severity of the disease. Testing rehabilitation interventions for knee OA can provide an understanding of possible preventative measures. Animal models such as the rat medial meniscal tear (MMT) model have been used in testing interventions. Two interventions, exercise and immobilization, were applied to the MMT injury model. A custom biplanar high-speed video radiography system was chosen to measure the kinematics of the experimental rats and quantify the effects of the interventions. Prior to use, workflows needed to be developed for this system. Additionally, the system needed to be validated and an appropriate analysis technique for the knee OA study needed to be chosen. Through extensive testing, an XMALab workflow reliant on manual recognition of joint centers and an Autoscoper workflow using 3D models of subject-specific bones were developed. The system’s accuracy and precision values were measured using phantoms of known length, yielding a system accuracy of 0.087 mm and precision of 0.073 mm. Qualitative and quantitative differences between the two workflows were compared and the Autoscoper workflow was chosen for the knee OA study for its ability to measure more angles. 14 rats were organized into four experimental groups: a non-intervention, an exercise intervention, an immobilization intervention, and a sham surgery group. The kinematic and spatiotemporal parameters were measured at three gait cycle events. Hip abduction results indicate the non-intervention group developed mild OA, while hip abduction and knee varus rotation results indicate the exercise intervention group developed advanced OA. The immobilization group results were indistinguishable from atrophic changes.

Committee:

Krishnendu Roy, Ph.D. (Georgia Institute of Technology)

Edward Botchwey, Ph.D. (Georgia Institute of Technology)

Fredrik Vannberg, Ph.D. (Georgia Institute of Technology)

Edmund Waller, M.D., Ph.D. (Emory University)

ROLE OF VASCULAR REMODELING IN THE ACCUMULATION, CLEARANCE, AND BIODISTRIBUTION OF BIOMOLECULAR FACTORS IN MELANOMA

Local inflammation within the tumor microenvironment is implicated in the systemic effects of disease progression, such as immune suppression and metastasis. Soluble factors (SF) produced within the tumor, including cytokines, exosomes, proteases, and microvesicles, mediate pathological signaling and have emerged as putative therapeutic targets. However, SF bioavailability in distributed tissues and the impact of disease progression on their dissemination profiles and exposure to various immune cell subsets is poorly defined. This stymies progress towards therapeutic amelioration of SF signaling activities to improve disease outcomes and is the critical knowledge gap this thesis seeks to fill. The central hypothesis is that tumor vascular remodeling redirects the organism-wide exposure of SF secreted locally within the tumor microenvironment, which may negatively contribute to disease burden by altering the bioavailability of molecules important to systemic disease progression. In this thesis, the effects of local tissue remodeling in melanoma leading to pathological SF accumulation profiles within distributed tissues are elucidated in order to provide insight into the potential for localized disease to exert systemic effects and inform opportunities to develop better preventive and curative treatment options for advanced melanomas.

Committee:

Jonathan S. Colton, Ph.D. (Georgia Institute of Technology)

Geza Kogler, Ph.D. (Georgia Institute of Technology)

Design and Prototyping of a Granular Foot Orthosis (FootGO)

Foot orthoses are assistive devices that are widely used to treat, alter, and in some cases correct different foot disorders. Inconsistencies during their manufacturing process can lead to ill-fitting orthoses, meaning that the clinician must go through many iterations to arrive at a properly formed insole. Iterative orthosis fabrication leads to extra cost, waste of materials, and increased lead time to help the patient. In addition to these front-end concerns, material durability during use is a major issue. To minimize these issues, we proposed to design a novel orthosis that utilizes granular jamming technology. I hypothesize that the Foot Granular Orthosis (FootGO), will exhibit tunable stiffness properties (by means of adjusting the vacuum pressure on the system) that allow it to mimic the material behavior of a wide range of commercial orthotic foams in a single generic device. Three types of granular media were used in different prototype compositions based on varying volume fill, particulate size, and granular media type. We conducted a series of uniaxial compression tests on a selection of foams as well as the newly designed FootGO prototypes, and obtained stress-strain curves from this data. Most of the prototypes agreed with expected material behavior trends. The energy absorption for each specimen, both foam and prototype, was then determined. The FootGO showed indications of superior durability through consistent results over repeated tests. The range of energy absorption performance for each FootGO prototype was compared to the range for commercial foams. The FootGO prototypes spanned the range over the various negative pressures, some of them extending well beyond the upper limit of the foam performance band.  These findings imply that varying the negative pressure within the FootGO produces corresponding adjustable stiffness properties and that the FootGO can potentially mimic the behavior of a wide range of orthotic materials.

BioE M.S. Defense Presentation

Date: Thursday, April 20^th 2017

Time: 2:00 PM

Location: 4029 EBB

Advisor:

Levi B. Wood, Ph.D. (Georgia Institute of Technology, School of Mechanical Engineering)

Thesis Committee:

Young Jang, Ph.D. (Georgia Institute of Technology, School of Biological Sciences)

Tony Kim, Ph.D. (Georgia Institute of Technology, School of Mechanical Engineering)

Effects of Neuorinflammation on Vascular Dysfunction in Alzheimer’s Disease

Alzheimer’s disease (AD) is the most common form of dementia, affecting more than 35 million people worldwide, and lacks any effective therapy to stop or slow the disease. AD is characterized by progressive appearance of extracellular amyloid beta (Aβ) plaques in affected regions of the brain, which lead to neuronal dystrophy and death. There is increasing evidence, however, that Aβ is not the sole driver of disease progression. A functional MRI study has revealed that the blood-brain barrier (BBB), a vital regulator of molecular transport between the brain and vascular system comprised in part of microvascular endothelial cells, becomes leaky early in disease. Furthermore, analysis of postmortem tissue has revealed reduced angiogenic vascular growth in AD tissues. Thus, vascular defects may promote neuronal death by reducing perfusion and allowing peripheral cells to enter the brain. The mechanisms responsible for BBB breakdown have not been delineated, but may be influenced or driven by Aβ and neuroinflammation. Inflammatory cytokines such as tumor necrosis factor alpha (TNFα) have been previously established by our lab and others to be upregulated in the AD microenvironment. I hypothesize that Aβ and inflammatory cytokines together drive loss of vascular endothelial barrier function and angiogenic sprouting. To test this hypothesis, I have used an integrated cell culture approach combining transwell plates, 2D cell cultures, and 3D microfluidic devices. Dextran permeability assays in microfluidics and transwell plates demonstrate that Aβ conditions increase endothelial permeability. Further, western blotting of 2D cultures demonstrates a dose-dependent down-regulation of both platelet-endothelial cell adhesion molecule (PECAM) and VE-cadherin expression in response to Aβ. Since PECAM is a critical regulator of angiogenesis, I quantified changes in angiogenic sprouting in response to Aβ using our microfluidic platform, thus in total my data will link PECAM dysregulation to dual vascular pathologies in AD: loss of barrier function and reduced angiogenic growth. In total, my data demonstrate that Aβ and key AD cytokines effect endothelial barrier function and angiogenic sprouting.  Moreover, my approach combining 2D and physiologically relevant 3D cell cultures provides great utility for interrogating vascular response to specific components of the AD microenvironment.

Robert J. Butera, Ph.D. (Georgia Institute of Technology, School of ECE and Dept. of BME)

Thesis Committee:

Laura O’Farrell, DVM, Ph.D. (Georgia Institute of Technology, GTRC)

Arthur English, Ph.D. (Emory University, Dept. of Cell Biology)

Chris Rozell, Ph.D. (Georgia Institute of Technology, School of ECE)

Thomas Burkholder, Ph.D. (Georgia Institute of Technology, School of Applied Physiology)

Optimization and Application of Kilohertz Electrical Stimulation to Autonomic Neural Circuits

More than half a century after the first implantation of a cardiac pacemaker (1958), there is a renewed interest in the ability to electrically stimulate biological tissues, specifically peripheral nerves, to relieve clinical conditions. In their most common form, these implanted devices deliver brief bursts of electrical pulses at various frequencies (< 130 Hz) to peripheral nerves, the spinal cord, or the brain to activate nearby excitable tissues. By doing so, these therapies treat symptoms related to disorders such as epilepsy, depression, and obesity. In stark contrast to this burst of neural activation induced by low frequency stimulation is the ability to completely inhibit neural activity using continuous, kilohertz frequency electrical stimulation. When applied to peripheral nerves, kilohertz electrical stimulation creates a localized block of peripheral nerve activity, thus inhibiting propagation of activity between neural elements (e.g., ganglia, spinal circuits, the brain) and end-targets (e.g., organs, muscles).

Although implemented in a handful of clinical devices, a significant number of parameters remain to be investigated for achieving a safe and effective nerve block using kilohertz electrical stimulation. Furthermore, the application of kilohertz electrical stimulation to block propagating activity in autonomic neural circuits, specifically as it relates to maintaining homeostasis, remains an uncharted territory. The goal of this thesis was thus two-fold - i) optimize kilohertz electrical stimulation parameters for safe and effective clinical implementation and ii) investigate the utility of kilohertz electrical stimulation nerve block in autonomic neural circuits involved in homeostatic regulation.

In the first part, I briefly discuss optimized electrode geometries that increase the battery life of implanted devices, as well as the electrochemical behavior of electrodes at kilohertz frequencies. In the second part, I discuss application of kilohertz electrical stimulation nerve block to autonomic neural circuits involved in regulation of systemic inflammation and glycemia. I demonstrate the therapeutic benefits of inhibiting nerve activity in each physiological circuit and discuss potential mechanisms underlying the observed therapeutic benefits.

Blue Jean Stream: https://bluejeans.com/291990472/browser

Meeting ID: 291990472

Committee:

Wilbur Lam M.D., Ph.D. (Georgia Institute of Technology, Emory University)

Julia Babensee, Ph.D. (Georgia Institute of Technology)

Lakshmi Sankar, Ph.D. (Georgia Institute of Technology)

Kevin Maher, M.D. (Emory University, Children's Healthcare of Atlanta)

Shriprasad Deshpande, M.D. (Emory University, Children's Healthcare of Atlanta)

Towards Improved Device Design and Clinical Management: The Thrombogenic Effect of the Fluid Dynamics and Material Surface Relationship

Blood-contacting medical devices, such as stents, grafts, catheters, extracorporeal circuits, and ventricular assist devices (VADs), are used to treat a variety of cardiovascular and cardiopulmonary diseases. Thrombotic complications are frequent sources of failure for these devices, and the balance of thrombosis, anticoagulation, and hemorrhage is currently an immense clinical challenge. Thrombosis not only hinders device function, but also poses direct risk to the patient. Device thrombosis is currently unpredictable due to a gap in understanding of the interaction of the contributing mechanisms: material surface activation and fluid dynamics. The purpose of this thesis was thus to elucidate the understanding of the material-flow relationship and its effects on bulk thrombotic outcomes, and to use a combination of analysis of clinical data and in vitro modeling to tackle thrombogenic issues in current devices and to make recommendations for future design. 

Committee Members:

Dr. Mark Prausnitz, ChBE (Georgia Institute of Technology)

Dr. Krishnendu Roy, BME(Georgia Institute of Technology)

Dr. Jennifer Leavey, Biology (Georgia Institute of Technology)

Dr. Baozhong Wang, (Georgia State University)

Protein Nanoparticle Vaccines

Highly conserved pathogen proteins are ideal components for broadly cross-protective vaccines, but tend to be poorly immunogenic. The presence of particulates in a vaccine has been known for more than 100 years to enhance a vaccine’s efficacy, yet only in recent decades has rational nanoparticle vaccine design emerged from advances in our understanding of immunology. While most vaccine nanoparticles seek to encapsulate or coat antigen onto particles, protein nanoparticles are made entirely of crosslinked antigen. This form of nanoparticulate antigen does not require the addition of stabilizing additives, and the nanoparticle is both the antigen and the immunostimulatory adjuvant. The objectives of this thesis are to (1) translate our initial immunization successes with conserved influenza protein M2e protein nanoparticles into other types influenza antigen nanoparticles, (2) understand the mechanism of nanoparticle adjuvancy, (3) explore extended storage of nanoparticle vaccines for the possibility of cold chain-independent storage, and (4) test the ability of molecular adjuvant coatings to boost protein nanoparticle immunogenicity.

Committee:

Ken Gall, PhD (Duke University)

Johnna Temenoff, PhD (Georgia Institute of Technology)

Meisha Shofner, PhD (Georgia Institute of Technology)

Nick Willett, PhD (Emory University)

Fatigue and Cyclic Loading of 3D Printed Soft Polymers for Orthopedic Applications

The use of soft, synthetic materials for soft tissue replacement in load-bearing, orthopedic applications has been largely unsuccessful due to a lack of adequate materials with sufficient fatigue and wear resistance. Silicone was once purported to be suitable for this purpose, and has been used in applications ranging from radial head implants to intervertebral disc replacements. However, the long term results for these devices demonstrated that there was significant room for improvement, with complications including implant fracture, deformation, and wear. More recently, there has been a surge in devices based on polycarbonate urethane (PCU), which has gained traction due to its relative biocompatibility, compliant nature, viscoelastic properties, as well as it’s durability as seen through preclinical device testing. Despite its promising nature, caution is warranted as the long-term clinical results of PCU devices have yet to be seen. Considering past difficulties, there is a clear need for a better fundamental understanding of the fatigue resistance of soft, synthetic polymers for such applications. Therefore, the purpose of this thesis was to develop useful processing-structure-property relationships for relevant soft polymers under fatigue loading in order to assist in the use and success of such polymers in load-bearing orthopedic applications.

Mark Styczynski Ph.D. (ChBE/Georgia Institute of Technology)

Committee:

Andreas Bommarius, Ph.D. (ChBE/Georgia Institute of Technology) 

Brian Hammer, Ph.D. (Biology/Georgia Institute of Technology)

Hang Lu, Ph.D. (ChBE/Georgia Institute of Technology)

Pamela Peralta-Yahya, Ph.D. (Chemistry/Georgia Institute of Technology)

Engineering a Fast-Responding Bacterial Test for Zinc Deficiency

 Zinc deficiency is estimated to affect millions of children each year, but the high cost and logistical challenges associated with current zinc diagnostic tools prevent adequate surveillance and treatment of zinc deficiency.  To be used in surveillance programs, a zinc status diagnostic test must be inexpensive, easy to administer, nearly equipment-free, and fast-responding. A bacterial biosensor has the potential to meet all of these requirements. Our group recently developed a low-cost E. coli biosensor that can produce different visible outputs based on the concentration of zinc in which the cells grow. However, these initial sensor cells always produce some pigment, which necessitates that they be grown from a very small inoculum so that initial coloration is minimal – thus requiring overnight assay times. To make an easy-to-use, field-deployable assay, we will engineer cells to repress pigmentation during growth and to quickly produce pigments from either the carotenoid or violacein pathways upon induction.  Zinc-responsive elements will also be tested in a lysate based cell-free system, and both the whole cell and cell-free systems will be tuned to respond to physiologically relevant serum zinc concentrations. The resulting biosensor will be a significant step towards a deployable, field-friendly zinc diagnostic tool.

Andrés J. García, Ph.D. (Georgia Institute of Technology)  

Committee:

Cheng Zhu, Ph.D. (Georgia Institute of Technology)

Jennifer E. Curtis, Ph.D. (Georgia Institute of Technology)

Andrew P. Kowalczyk, Ph.D. (Emory University)

Aránzazu del Campo, Ph.D. (Leibniz Institute for New Materials)

Force-Signaling Coupling at Single Focal Adhesions

Cell adhesion plays a critical role in development, physiology, and disease. Despite significant progress in determining the biochemistry driving cell adhesion assembly and signaling, very little is known about how cell adhesive forces are transduced into biochemical signals. The objective of this project is to analyze how focal adhesions (FAs) sense and transmit force. Our central hypothesis is that mechanosensing at a single FA involves a feedback loop in which force regulates FAK phosphorylation, which controls recruitment of vinculin and paxillin to the FA to tune the local force balance between extracellular matrix-integrin forces and cytoskeletal tension. Through this integrated module, force and FA signaling are coupled.

 We will first analyze the effects of local adhesive force and cytoskeletal tension on FAK phosphorylation and its coupling to force generation and vinculin and paxillin recruitment to FAs. We will establish spatiotemporal profiles for force transmission and FA assembly at single FAs by analyzing live cells expressing fluorescent FA proteins on microfabricated pillar-array-detectors (mPADs), which consist of an array of silicone micropillars, and are commonly used to measure cell traction forces, based on the deflections of the micropillars. The micropillars will also present caged RGD peptide, which can be activated with UV light to initiate FAs at prescribed sites and generate coupled spatiotemporal FA assembly and force generation profiles at single adhesions. We will evaluate these force-signaling responses on mPADs with different elastic moduli and in the presence of contractility modulators. By varying substrate stiffness and contractility state, we will perturb the local force balance between ECM-integrin forces and cytoskeletal tension to examine how adhesive force balance regulates FAK phosphorylation and vinculin-paxillin recruitment at single FAs. We will use also FAK inhibitors to determine how FAK phosphorylation regulates force transmission and vinculin and paxillin recruitment. We hypothesize that adhesive forces at FAs regulate FAK phosphorylation, which then drives paxillin localization and vinculin recruitment. We will next evaluate force-FAK phosphorylation coupling for cells expressing mutant vinculin head and tail domains to dissect the contributions of vinculin head and tail domains to regulating force-dependent FAK phosphorylation at FAs. As outcomes of this project, we will establish how the local balance of adhesive force and cytoskeletal tension regulates coupled force-FAK signaling at single FAs. We will also dissect the roles of FAK and vinculin in this mechanosensing. This research will generate insights into how cell adhesive forces are integrated into biochemical signals. This understanding will provide a framework for mechanotransduction events at cell-ECM junctions, such as adhesion assembly at migratory fronts, force-regulated morphogenesis, and stem cell lineage commitment in response to matrix stiffness.

Committee:

Asma Nusrat, M.D. (University of Michigan)

Johnna Temenoff, Ph.D. (Georgia Institute of Technology)

Krishnendu Roy, Ph.D. (Georgia Institute of Technology)

Alberto Fernandez De Las Nieves, Ph.D. (Georgia Institute of Technology)

Synthetic Hydrogels that Recapitulate Epithelial Morphogenesis Programs

Understanding the contributions of the ECM biophysical and biochemical properties to epithelial cell responses has been a major goal for biomaterials scientists in order to engineer materials that can recapitulate ECM-mediated epithelial morphogenesis programs. Although 3D natural matrices have been found suitable for the study of many cellular processes, they are limited by lot-to-lot compositional and structural variability, inability to decouple mechanical and biochemical properties, and in some cases, their tumor-derived nature limits their clinical translational potential. Therefore, there is a significant need for a biomaterial matrix that can recapitulate epithelial morphogenetic programs while overcoming these limitations.

This project aims to develop an engineered synthetic hydrogel matrix that presents independently-tunable ECM-like bioactivity and mechanical properties, and can support epithelial cell survival, proliferation, polarization, and assembly into 3D multicellular structures recapitulating different epithelial morphogenesis program. This synthetic material has the capacity to present adhesive peptides and protease-degradable crosslinks that support cell functions and promote cell engraftment in vivo. As part of this project, we have developed an engineered synthetic hydrogel platform that recapitulates the morphogenetic program of human pluripotent stem cell (hPSC)-derived intestinal organoids (HIOs), and has been established as a delivery vehicle for HIOs to mucosal intestinal wounds in mice. Furthermore, in order to prove the versatility of our hydrogel platform, we aim to engineer a synthetic hydrogel that recapitulates the mouse renal proximal tubular cells (RPTCs) morphogenetic program. We hypothesize that these engineered hydrogels will be superior to naturally-derived materials by supporting these different epithelial morphogenetic programs while overcoming the imitations of natural and other synthetic materials. This synthetic hydrogel technology is innovative as it allows the study of the independent contributions of ECM properties to different epithelial morphogenetic programs, and will form a basis for the adaptation to in vitro generation and in vivo delivery of human PSC-derived organoids for regenerative medicine.

Committee Members:

Julie A. Champion, PhD (Georgia Institute of Technology)

Susan N. Thomas, PhD (Georgia Institute of Technology)

Edward A. Botchwey, PhD (Georgia Institute of Technology)

Krishnendu Roy, PhD (Georgia Institute of Technology)

Conditioning Dendritic Cell Responses using Engineered Biomaterials for Immunotherapy

Pivotal discoveries in the field of immunology over the last five decades have changed the way new therapies are designed for applications as varied as organ transplantation, autoimmune diseases or even cancer. In this regard, dendritic cells (DCs) were identified to play an important role in the orchestration of adaptive immune response. Importantly, the phenotype of DCs is a powerful indicator of their downstream effector functions. In the recent years, parallel advancements made in biomaterial design and biocompatibility considerations are being directly translated into developing improved immunotherapies. Interestingly, biomaterials also elicit differential effects on the host immune response as well as on the phenotypic state of DCs.

 The first objective of this doctoral thesis was to validate the role of DCs in the previously documented in vivo PLGA adjuvant effect in boosting a humoral immune response to co-delivered antigen. Herein, by conditionally ablating DCs in a murine CD11c-DTR model, the adjuvant effect of PLGA towards co-delivered OVA was revisited. The diminished proliferation of adoptively transferred OVA-reactive T-cells in these mice was suggestive of a lowered adjuvant effect due to the absence of CD11c+ DCs. The second objective of this thesis was to design, develop and validate a multicomponent, multifunctional immunomodulatory (MI) scaffold comprised of macroporous agarose as the base scaffold material into which were embedded crosslinked gelatin microparticles (MPs), pre-loaded with immunomodulators, for their controlled release to mimic tolerogenic human or murine DC culture conditions. Aided by empirical modeling, using the Weibull equation, of experimental data using ‘model’ proteins, we identified parameters of gelatin MP crosslinking density and number of embedded MPs in agarose to achieve prescribed temporal controlled release of immunomodulators for induction of tolerogenic DCs.  The prescribed MI scaffold aimed to release granulocyte monocyte colony-stimulating factor (GM-CSF; delivered within 0-3 days) to induce differentiation of monocyte precursors into DCs after dexamethasone (DEX, delivered within 3-6 days) addition would induce regulatory properties to these cells as well as peptidoglycan (PGN, delivered on days 5-6) to induce an alternative activated phenotype in DCs.  Such alternatively activated DCs (aaDCs), are endowed with immunosuppressive as well as directed lymph node migratory properties to effectively exert their tolerogenic effect.  Ability of this MI scaffold to induce tolerogenic phenotype in human blood-derived as well as murine bone marrow-derived cells was demonstrated upon in vitro treatment using a large cadre of immunological assessments.

In summary, this thesis documents the importance of DCs in the biomaterial adjuvant effect in vivo and provides a construct formulation that can be used to generate aaDCs with tolerogenic and migratory properties that are highly relevant in designing future immunotherapies targeting autoimmune diseases as well as in alleviating allograft rejection.

Monday October 24th, 2016
1:00 pm EDT
1128 IBB

950 Atlantic Drive Atlanta, GA 30332

Advisor: Craig R. Forest, PhD (Georgia Institute of Technology)

Committee:
Edward S. Boyden, PhD (Massachusetts Institute of Technology)
Hongkui Zeng, PhD (Allen Institute for Brain Science)
Garrett B. Stanley, PhD (Georgia Institute of Technology)
Todd Sulchek, PhD (Georgia Institute of Technology)
Suhasa B. Kodandaramaiah, PhD (University of Minnesota)

“In Vivo Serial Patch Clamp Robotics for Cell-Type Identification in the Mouse Visual Cortex”

In 2013, President Obama announced the Brain Initiative to fund the development of new tools for studying the brain and to identify the root causes of nervous system disorders.  Our knowledge of the brain is currently limited by our ability to record the dynamic activity of neurons in intact, behaving circuits.  Here we show the development of robotics tools to investigate the unique behaviors of neurons in the visual cortex of mice and transform the highly manual art of obtaining patch clamp recordings into a systematic, automated procedure.

The patch clamp technique is the gold standard for recording the intracellular electrical activity of individual cells and has the highest resolution and specificity of any other technique.  However, the manual methods used to control the position, pressure, and voltage of the glass recording pipette severely limit the throughput and the ability to perform multiple simultaneous recordings in vivo.  This work shows the development of automation systems to precisely and repeatably prepare the recording pipette, position it in the brain, establish the recording, and conduct the entire electrophysiological experiment all without requiring the presence of a human operator.  The robot has autonomously obtained multiple, consecutive recordings in vivo with the same quality and throughput as a human operator.  Robotic hardware and software algorithms enable parallel scaling for increased throughput, systematic operation, and rapid dissemination of challenging techniques.  These tools will increase our capacity to rapidly identify new cell-type classification schemes and understand the in vivo function and dysfunction of cells within the nervous system.

C. Ross Ethier, Ph.D. (BME/Georgia Institute of Technology)

 Committee:

Andrés, J. García, Ph.D. (ME/Georgia Institute of Technology)

Robert E. Guldberg, Ph.D. (ME/Georgia Institute of Technology)

J. Brandon Dixon, Ph.D. (ME/Georgia Institute of Technology)

Ian A. Sigal, Ph.D. (Ophthalmology/University of Pittsburgh)

Finite Element Modeling of Rat Optic Nerve Head Biomechanics

Glaucoma is the second leading cause of blindness and is characterized by the loss of retinal ganglion cells (RGCs). Biomechanical insult, especially occurring in the optic nerve head (ONH), is thought to be a key driver leading to RGC death in glaucoma. However, the link between biomechanics and RGC death is not well understood. The rat has been widely used to study glaucoma, especially RGC pathophysiology. However, rat ONH biomechanics have not been characterized. We plan to characterize rat ONH biomechanics using finite element modeling. We hypothesize that regional patterns of stress and strain in the rat ONH will coincide with regional patterns of biological response seen in rat glaucoma studies conducted by collaborating labs.

Signaling molecules and their activities are well coordinated inspace and time to regulate cellular functions in response to mechanicaland chemical microenvironment. Based on fluorescent resonance energytransfer (FRET), we have developed several genetically encodedbiosensors for detecting the spatiotemporal activities of signalingmolecules, including Src, Rac1, MT1-MMP, and Calcium. A Rac biosensorrevealed that the Rac activity in cells constrained on micropatternedextracellular-matrix surface is polarized with higher activityconcentrated at the leading edge of migrating cells upon PDGFstimulation, whereas Src activities in these cells displayed globalactivation patterns without obvious polarity. Our calcium biosensoralso allowed the revelation that there is a spontaneous Ca2+oscillation in human mesenchymal stem cells (HMSCs) both inside thecytoplasm and endoplasmic reticulum (ER). The substrate stiffness whereHMSCs are seeded can significantly affect this Ca2+ oscillation, in afashion dependent on the RhoA signaling pathway. We have furtherdeveloped a FAK FRET biosensor and targeted it into lipid rafts ornon-rafts of plasma membrane by lipid modifications. Upon cell adhesionon extracellular matrix proteins or stimulation by platelet-derivedgrowth factor, the raft-targeting FAK biosensor showed a surprisinglystronger FRET response than that at non-rafts, suggesting that the FAKactivation mainly occurs at lipid rafts. Further experiments revealedthat the PDGF-induced FAK activation at rafts is mediated by the kinaseactivity of Src, whereas FAK activation induced by adhesion isindependent of, and in fact essential for the Src activation. Theseresults suggest that FAK is activated at rafts with distinct activationmechanisms in response to different physiological stimuli. In summary,our novel FRET biosensors in combination with tools innano-biotechnology and bio-photonics have made it possible to monitorkey signaling cascades in live cells with subcellular and dynamiccharacterization when cells interact with their physical/chemicalmicroenvironment.

Osteoarthritis (OA) is a debilitating degenerative disease thatafflicts an estimated 27 million Americans age 25 and older.  Thisdisease leads to the progressive degradation of the articular layers ofdiarthrodial joints, significantly compromising the main function ofcartilage as a load bearing material, leading to pain and limitingactivities of daily living.  Cartilage functional tissue engineering isa highly promising technology that aims to provide a biologicalreplacement to worn articular layers, as a modality that considerablyexpands the limited options in the treatment of this disease.  Thoughcartilage degeneration is occasionally limited to small focal areaswithin articular layers, OA generally becomes symptomatic whendegradation has spread over much greater surface areas (such as greaterthan 25 percent of the articular layer). Unfortunately, functionaltissue engineering of large cartilage constructs is significantlyconstrained by the balance of nutrient transport and
consumption.  Several studies have shown that matrix deposition andelaboration of functional properties preferentially occurs near theperiphery of constructs, where nutrient supply from the surroundingculture medium is most abundant, whereas cells in the interior receiveless nutrients and produce less matrix, with poorer functionalproperties. In this presentation, we show that dynamic mechanicalloading can enhance
solute transport by up to an order of magnitude, and this enhancementcan be considerably accelerated by placing channels in the constructs.

Abstract:
The threats of disease outbreaks and bioterrorism events demand portable technology capable of rapid, sensitive, and accurate diagnosis for timely treatment and containment. In this seminar, I will present the efforts at Sandia National Labs towards addressing such public health concerns through the development of the SpinDx platform, a portable centrifugal microfluidic device capable of high sensitivity, multi-modal, multi-analyte measurements. This versatile platform uses a disposable microfluidic disc designed for handling an array of small volumes of clinical samples to perform multiplexed diagnostics. A novel sample-to-answer immunoassay approach has been demonstrated for ultrasensitive toxin detection via binding of toxins to antibody-laden capture particles followed by sedimentation of the particles through a density-media in a microfluidic disc and quantification by laser-induced fluorescence. In addition, nucleic acid tests via isothermal amplification have been performed with single cell sensitivity using a non-contact temperature control system. As the recent Ebola outbreak and the current Zika crisis have shown, reliable and field-deployable technology such as SpinDx is urgently needed around the globe for accurate diagnostics in low-resource settings.

Bio:
Christopher Phaneuf earned his Bachelor’s degree in Mechanical Engineering from The Cooper Union in 2008. That same year he began his graduate studies in the Bioengineering program at Georgia Tech where he joined the Precision Biosystems Lab working with Prof. Craig Forest. As a Department of Homeland Security graduate fellow, Christopher developed a microfluidic platform designed for point-of-care molecular diagnostics. In collaboration with the Centers for Disease Control and Prevention (CDC) and Children’s Healthcare of Atlanta, the compact, low-cost platform was used to perform multiplexed detection of viral targets. After completing his doctorate in the Fall of 2014, he joined Sandia National Laboratories in Livermore, CA as a postdoc in the Biotechnology and Bioengineering division to develop microfluidic technologies for biodefense and public health applications.

Committee:

• Cyrus K. Aidun, Ph.D. (Georgia Institute of Technology)
• Wei Sun, Ph.D. (Georgia Institute of Technology)
• F. Levent Degertekin, Ph.D. (Georgia Institute of Technology)
• Joseph H. Gorman, M.D. (University of Pennsylvania)
• Vinod H. Thourani, M.D. (Emory University)

Wilbur Lam, M.D., Ph.D. (Georgia Institute of Technology & Emory University)

Committee:

Brandon Dixon, Ph.D. (Georgia Institute of Technology)

Todd Sulchek, Ph.D. (Georgia Institute of Technology)

Susan Thomas, Ph.D. (Georgia Institute of Technology)

Hua Wang, Ph.D. (Georgia Institute of Technology)

DEVELOPING MICROFLUIDIC SYSTEMS TO DECOUPLE BIOPHYSICAL AND BIOCHEMICAL ASPECTS OF HEMATOLOGICAL PROCESSES

Recent research has revealed that cells dynamically sense and respond to their physical microenvironments. For instance, in hematology specifically, it was shown that shear mediated red blood cell (RBC) deformation results in ATP release, and that platelets attenuate contraction force based on substrate stiffness. The objective of this proposal is thus to create microfluidic systems in which the biophysical and biochemical aspects of hematological processes can be independently investigated. More specifically, this proposal will present novel microfluidic devices: an “endothelial”-ized, T-junction to elucidate the biophysical processes that define the mechanism of action of the ferric chloride thrombosis model; a micropillar array to examine the physical effect of a geometrically relevant, non-biological matrix on platelet and RBC activity; and an electrospun fibrinogen mesh device and a micro-slit device to define the physical parameter space (shear, time of deformation, cell stiffness) that governs RBC fragmentation. Microfluidic platforms allow for real-time, microscopic evaluation of cell response (via brightfield morphology and immunostaining) and precise spatiotemporal control of system inputs and flow characteristics, including shear stress. The knowledge gained by successfully decoupling the biophysical and biological aspects of hematology can result in improved diagnostic assays for blood cell activity and new targets for therapeutics. 

J. Brandon Dixon, Ph.D. (ME/Georgia Institute of Technology)

Committee:

Andrés J. García, PhD (Georgia Institute of Technology)

C. Ross Ethier, PhD (BME/(Georgia Institute of Technology)

Michael Davis, PhD (BME/Georgia Institute of Technology & Emory University)

Mariappan Muthuchamy (Medical Physiology/Texas A&M)

Role of Mechanical Microenvironment on the Regulation of Lymphatic Function and Health

Failure of lymph fluid transport plays an important role in pathologies, such as lymphedema and lymphacele after organ donation. The lymphatic system plays a critical role in maintaining fluid homeostasis in all of the soft tissue of the body. Its ability to transport interstitial fluid is partially dependent on the intrinsic pumping capacity of lymphatic smooth muscle cells. Recently, our lab has shown that lymphatic collecting vessels near an injury never return to pre-injury levels of pumping and lymph transport, but factors that impact collecting vessel phenotype are poorly understood. The central hypothesis of this work is that the components of the extracellular matrix (ECM) and mechanics of the lymphatic microenvironment play a central role in the contractile phenotype of lymphatic muscle cells (LMCs). 

Zhou Yuan

Ph. D Proposal Presentation

Wednesday, January 18^th, 2017, 3pm

3:00pm, IBB Seminar Room 1128

Advisor:

Dr. Zhu (Georgia Institute of Technology)

Committee:

Dr. Susan Thomas (Georgia Institute of Technology)

Dr. Michelle Krogsgaard (New York University)

Dr. Mandy Ford (Emory University)

Dr. Gabe Kwong (Georgia Institute of Technology)

Understanding the effect of tumor microenvironment on T cell antigen recognition

Skin cancer is the most common cancer in the United States, among which melanoma is the most serious type with high mortality rate. Despite the critical role of CD8⁺ T cells in tumor clearance, their functions in the tumor microenvironment (TME) are impaired by immunosuppressive cells/cytokines, inhibitory receptors, and metabolic restriction. Targeting these pathways were shown to promote tumor clearance, yet unknown mechanisms may still exist curtailing the T cell responses. T cell activation has been shown to be largely determined by the in situ mechanokinetic properties of the binding of T cell receptor (TCR) to peptide-major histocompatibility complex (pMHC), which are sensitive to perturbations of the cellular environment. Our preliminary studies have shown that the molecular interactions involved in T cell antigen recognition are altered in the TME. The present thesis will study the extent of this alteration, how such alteration consequentially suppresses T cell effector functions, and what the underlying mechanisms are. This study aims to address these questions with animal models, highly-sensitive biomechanical assays of single molecules, and other cellular and bimolecular approaches. The outcome will greatly enhance our understanding of the impaired anti-tumor T cell responses and inspire novel strategies for cancer immunotherapy.

Devon M. Headen

Thursday, January 26^th, 2017, 3:00PM

Petit Institute Suddath Seminar Room (IBB 1128)

Advisor: Andrés J. García, PhD, ME, Georgia Tech

Committee:

W. Robert Taylor, MD, PhD (Department of Medicine, Emory University)

Hang Lu, PhD,(ChBE, Georgia Tech)

Peter M. Thulé, MD (Department of Medicine, Emory University)

Krishnendu Roy, PhD (BME, Georgia Tech)

Microfluidics-based Microgel Synthesis for Immunoisolation and Immunomodulation in Pancreatic Islet Transplantation.

Encapsulation of islets in hydrogel microspheres (microgels) before transplantation into diabetic recipients can establish an adequate immuno-isolation barrier to mitigate allogeneic rejection. The synthetic hydrogel macromer PEG-4MAL (4-arm polyethylene glycol terminated with maleimides) is an ideal candidate polymer for immunoisolation applications, since it can be easily modified with thiolated bioactive molecules, allowing precise control of islet microenvironment. Alginate microencapsulation dominates in literature even though alginate provides limited control of islet microenvironment, because no technique exists for islet encapsulation in synthetic microgels. Therefore, a microfluidic platform for the encapsulation of islets in size-controlled PEG-4MAL microgels was developed, and hydrogel composition was optimized to support encapsulated islet function. Islets microencapsulated in optimized PEG-4MAL restored glycemic control better than islets microencapsulated in alginate and equally as well as unencapsulated islets when delivered to epididymal fat pads in diabetic syngeneic mice within bulk vasculogenic hydrogels. Improved function was partially attributed to decreased microgel size vs. alginate, and therefore reduced diffusional barrier. Immuno-isolation potential of this strategy is currently being investigated in allogeneic recipients. In a separate scheme, PEG-4MAL microgels were designed which could capture and display the chimeric immunomodulatory protein SA-FasL in its bioactive form. Simple cotransplantation of SA-FasL presenting microgels with unmodified allogeneic islets under the kidney capsule of diabetic mice resulted in long term graft acceptance without long term immunosuppression. Regulatory T cells mediated this acceptance since their ablation on day 50 post-transplantation prompted rapid graft rejection. Effective control or mitigation of immune responses is critical for successful outcomes in islet transplantation, and this work presents the development of two novel strategies for achieving long term function of allogeneic islet grafts.