Organizational Unit:
Wallace H. Coulter Department of Biomedical Engineering

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https://ror.org/02j15s898
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Now showing 1 - 10 of 17
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    Immuno-suppressive hydrogels for stem cell therapy after traumatic brain injury
    (Georgia Institute of Technology, 2019-11-20) Alvarado-Velez, Melissa
    During a traumatic brain injury (TBI) an external force disrupts the brain tissue and the proper functioning of neuronal pathways. This initial insult activates multiple cellular mechanisms that further propagate the tissue damage causing a secondary injury that exacerbates neurological deficits. This phase, known as the secondary injury, opens a therapeutic window in which neuroprotective treatments that successfully contain the propagation of the initial damage could significantly reduce neurological deficits associated with TBI. Mesenchymal stem cell transplantation (MSC) after TBI has been found to ameliorate neurological deficits due to the ability of the stem cells to modulate inflammation and immune cells and to increase the expression of neurotrophic factors that promote the survival of the neuronal tissue surrounding the injury site. However, the active rejection of the transplanted MSC by the host immune system could strongly diminish the stem cell's survival and therapeutic effect. In this thesis, we used immunosuppressive hydrogels, specifically designed to induce the apoptosis of cytotoxic CD8+ T cells, to enhance the survival of transplanted MSC in the injured brain. We demonstrated that creating localized immunosuppression near the MSC transplantation site resulted in a higher presence of MSC near the injury site. We also demonstrate that enhancing MSC survival by using immunosuppressive hydrogels increased the protein expression of the neurotrophic factors, which could lead to reduced neuronal damage. Therefore, the development of immune-suppressive hydrogels for stem cell transplantation could be a successful approach to enhance stem cell therapy after TBI.
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    Investigating differential electrotaxis of glioblastoma and medulloblastoma spheroidal aggregates
    (Georgia Institute of Technology, 2017-11-02) Lyon, Johnathan G.
    Treatment of brain and nervous system cancers remains a daunting clinical challenge, with one of the most common brain malignancies, glioblastoma, incurring a mere 5% five-year survival rate. In the search for new therapeutic means—specifically, new ways to direct or curb the invasion of brain cancers—we investigate the directed invasion of brain cancer cellular aggregates by an electrical field. This property is known as electrotaxis (or galvanotaxis), and is known to be involved in a variety of endogenous phenomena including tissue development and wound healing, and is purported to be involved in cancer invasion and metastasis. In this study, we explore electrotaxis in the context of brain cancer, and provide new insights into the effect’s underlying mechanisms. Here, we have developed an electrotaxis assay that adapts existing cancer invasion assays and allows us to study electrotaxis on aggregated populations of cells. We characterize glioblastoma and medulloblastoma cell lines in these assays and report on their electrotactic properties. We then investigate the transcriptome for two cell lines that were found to have opposing electrotactic responses, followed by pharmacological inhibition studies to further validate some of our findings. Overall, the hypotheses explored in this work advance our understanding of the pathways that underlie electrotactic sensing and steering and may lead to new means for guiding or managing brain malignancies.
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    Strategies for controlling the innate immune responses to in vitro transcribed messenger RNAs
    (Georgia Institute of Technology, 2016-03-15) Loomis, Kristin Helene
    Synthetic messenger RNA (mRNA) produced via in vitro transcription (IVT mRNA) has emerged as an appealing tool for the transient introduction of genes, particularly for vaccination applications. The interaction that IVT mRNA has with the innate immune system is centrally important to its performance as a vaccine. These innate immune responses can both interfere with the expression of the encoded antigenic protein and direct development of adaptive immunity. The objective of this thesis is to investigate the innate immune responses to IVT mRNA and to identify strategies to modulate these immune responses. We first demonstrated that substitution of the modified bases 5-methylcytosine and pseudouridine in IVT mRNA consistently reduces antiviral cytokine responses but affects transgene expression in a gene-specific manner. To assess the pathogen recognition receptors involved in detection of IVT mRNA, we developed proximity ligation assays, which allowed histological identification of PRR signaling complexes. We used these assays to identify that nanoparticle-mediated delivery modified PRR-activation following intramuscular delivery compared to delivery of the naked IVT mRNA molecule. Lastly, we developed a strategy to program the immunostimulatory properties of IVT mRNA by tethering adjuvants directly to the molecule. We show that upon intramuscular injection, the combination delivery of a TLR7 adjuvant and IVT mRNA lead to heightened local antiviral responses when delivered tethered, rather than as a cocktail. This work provides a foundation for the modulation and systematic study of IVT mRNA’s interaction with the innate immune system. Insights gained from this work may help direct and advance the design of IVT mRNA sequences for vaccination applications.
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    The Georgia Tech regenerative electrode - A peripheral nerve interface for enabling robotic limb control using thought
    (Georgia Institute of Technology, 2015-07-20) Srinivasan, Akhil
    Amputation is a life-changing event that results in a drastic reduction in quality of life including extreme loss of function and severe mental, emotional and physical pain. In order to mitigate these negative outcomes, there is great interest in the design of ‘advanced/robotic’ prosthetics that cosmetically and functionally mimic the lost limb. While the robotics side of advanced prosthetics has seen many advances recently, they still provide only a fraction of the natural limbs’ functionality. At the heart of the issue is the interface between the robotic limb and the individual that needs significant development. Amputees retain significant function in their nerves post-amputation, which offers a unique opportunity to interface with the peripheral nerve. Here we evaluate a relatively new approach to peripheral nerve interfacing by using microchannels, which hold the intrinsic ability to record larger neural signals from nerves than previously developed peripheral nerve interfaces. We first demonstrate that microchannel scaffolds are well suited for chronic integration with amputated nerves and promote highly organized nerve regeneration. We then demonstrate the ability to record neural signals, specifically action potentials, using microchannels permanently integrated with electrodes after chronic implantation in a terminal study. Together these studies suggest that microchannels are well suited for chronic implantation and stable peripheral nerve interfacing. As a next step toward clinical translation, we developed fully-integrated high electrode count microchannel interfacing technology capable of functioning while implanted in awake and freely moving animal models as needed for pre-clinical evaluation. Importantly, fabrication techniques were developed that apply to a broad range of flexible devices/sensors benefiting from flexible interconnects, surface mount device (SMD) integration, and/or operation in aqueous environments. Examples include diabetic glucose sensors, flexible skin based health monitors, and the burgeoning flexible wearable technology industry. Finally, we successfully utilized the fully integrated microchannel interfaces to record action potentials in the challenging awake and freely moving animal model validating the microchannel approach for peripheral nerve interfacing. In the end, the findings of these studies help direct and give significant credence to future technology development enabling eventual clinical application of microchannels for peripheral nerve interfacing.
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    In-vivo study of brain tumor migration via electrospun nanofiber implants
    (Georgia Institute of Technology, 2015-01-28) Cho, Jae Sung
    Glioblastoma multiforme (GBM), one of the deadliest forms of human brain cancer, migrates to different parts of the brain via the white matter tracts. This behavior is the basis for biomaterial research currently done to mimic white matter tracts so that GBM migration can be investigated .While there have been many in-vitro studies done on GBM migration with electrospun nanofiber films, only one in-vivo study has been done on GBM migration. Encouraged by our findings on GBM cell migration on aligned fiber films published in Nature Materials, we proposed to make two new implant designs, the aligned conduit implant and the silicone tube implant and utilize these nanofiber films to investigate GBM cell migration from inside the brain to outside of the brain. It was found that the silicone tube implants had a design flaw that hindered GBM cell migration from the tumor. The aligned conduit implant facilitated GBM migration significantly with a p-value of 2.01×10-4. Quantification of migration was done using a recently introduced SeeDB protocol, which greatly expedited analysis time. The results from in this investigation show that it is possible to design a brain implant that is able to remove GBM tumor non-invasively and will add to the advancement to biomedical technology in this field.
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    Probabilistic encoding and feature selectivity in the somatosensory pathway
    (Georgia Institute of Technology, 2014-06-30) Gollnick, Clare Ann
    Our sensory experiences are encoded in the patterns of activity of the neurons in our brain. While we know we are capable of sensing and responding to a constantly changing sensory environment, we often study neural activity by repeatedly presenting the same stimulus and analyzing the average neural response. It is not understood how the average neural response represents the dynamic neural activity that produces our perceptions. In this work, we use functional imaging of the rodent primary somatosensory cortex, specifically the whisker representations, and apply classic signal-detection methods to test the predictive power of the average neural response. Stimulus features such as intensity are thought to be perceptually separable from the average representation; however, we show that stimulus intensity cannot be reliably decoded from neural activity from only a single experience. Instead, stimulus intensity was encoded only across many experiences. We observed this probabilistic neural code in multiple classic sensory paradigms including complex temporal stimuli (pairs of whisker deflections) and multi-whisker stimuli. These data suggest a novel framework for the encoding of stimulus features in the presence of high-neural variability. Specifically we suggest that our brains can compensate for unreliability by encoding information redundantly across cortical space. This thesis predicts that a somatosensory stimulus is not encoded identically each time it is experienced; instead, our brains use multiple redundant pathways to create a reliable sensory percept.
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    Understanding the role topographical features play in stimulating the endogenous peripheral nerve regeneration across critically sized nerve gaps
    (Georgia Institute of Technology, 2011-11-11) Mukhatyar, Vivek
    Severe traumatic injuries and surgical procedures like tumor resection often create peripheral nerve gaps, accounting for over 250,000 injuries in the US annually. The clinical "gold standard" for bridging peripheral nerve gaps is autografts, with which 40-50% of patients regain useful function. However, issues including their limited availability and collateral damage at the donor site limit the effectiveness and use of autografts. Therefore, it is critical to develop alternative bioengineered approaches that match or exceed autograft performance. With the use of guidance channels, the endogenous regeneration process spontaneously occurs when successful bridging of short gaps (< 10mm) occurs, but fails to occur in the bridging of longer gaps (≥15mm). Several bioengineered strategies are currently being explored to bridge these critical size nerve gaps. Other labs and ours have shown how filler materials that provide topographical cues within the nerve guides are able to enhance nerve growth and bridge critical length gaps in rats. However, the mechanism by which intra-luminal fillers enhance nerve regeneration has not been explored. The main goal of this dissertation was to explore the interplay between intra-luminal scaffolds and orchestrated events of provisional fibrin matrix formation, glial cell infiltration, ECM deposition and remodeling, and axonal infiltration - a sequence we term the 'regenerative' sequence. We hypothesized that the mechanism by which thin films with topographical cues enhance regeneration is by serving as physical 'organizing templates' for Schwann cell infiltration, Schwann cell orientation, extra-cellular matrix deposition/organization and axon infiltration. We demonstrate that aligned topographical cues mediate their effects to the neuronal cells through optimizing fibronectin adsorption in vitro. We also demonstrate that aligned electrospun thin films are able to enhance bridging of a critical length nerve gap in vivo by stabilizing the provisional matrix, creating a pro-inflammatory environment and influencing the maturation of the regenerating cable leading to faster functional recovery compared to smooth films and random fibers. This research will advance our understanding of the mechanisms of peripheral nerve regeneration, and help develops technologies that are likely to improve clinical outcomes after peripheral nerve injury.
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    A nanoencapsulated visible dye for intraoperative delineation of brain tumor margins
    (Georgia Institute of Technology, 2011-10-24) Roller, Benjamin Thomas
    Brain and central nervous cancer presents a significant clinical burden, accounting for 2.4% of all cancer deaths. High grade glioma is particularly deadly, with 5 year survival times of 35% or less. Traditional treatment includes tumor resection followed by radiation therapy or chemotherapy. Aggressive resection is essential in order to prolong patient life. In fact, several studies have shown that life expectancy increases with increased extent of resection. Extent of resection is burdened by the fact that surgeons must be careful not to remove functional brain tissue. Resection is incomplete more often than not due to lack of visual cues for the surgeon. He must rely on tactile sensation to distinguish tumor from healthy tissue. Methods such as intraoperative MRI and CT exist, but these require expensive equipment and special training that is not available in all surgical environments. Some laboratories have proposed small molecule dyes to solve this problem, but these are insufficient when used in an invasive tumor model. It was the goal of this research to provide an objective cue in the form of a nanoencapsulated visible dye without the need for additional equipment of changes to the surgery process itself other than injection of the dye. We hypothesized that the nanocarrier would allow staining of the tumor through passive targeting by taking advantage of the enhanced permeability and retention effect. Once the nanocarriers have reached the desired target, they would not diffuse out into healthy tissue due to their large size compared to small molecule dyes, which readily diffuse out and stain healthy tissue. To test this hypothesis, we prepared and characterized a liposomal nanocarrier encapsulating Evans blue dye. The nanocarrier was tested for safety in vitro and in vivo, then used to delineate tumor margins in an invasive rat glioma model in vivo. Microscopic analysis was then conducted to ensure only tumor tissue was stained by the nanocarrier. This thesis presents a successful method of tumor border delineation to provide surgeons with positive visual cues without the need for changes in surgical environment or techniques.
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    Topographic guidance scaffolds for peripheral nerve interfacing
    (Georgia Institute of Technology, 2010-11-22) Clements, Isaac Perry
    In response to high and rising amputation rates, significant advances have been made in the field of prosthetic limb design. Unfortunately, there exists a lag in the neural interfacing technology required to provide an adequate link between the nervous system and this emerging generation of advanced prosthetic devices. Novel approaches to peripheral nerve interfacing are required to establish the stable, high channel count connections necessary to provide natural, thought driven control of an external prosthesis. Here, a tissue engineering-based approach has been used to create a device capable of interfacing with a regenerated portion of amputated nerve. As part of this work, a nerve guidance channel design, in which small amounts of interior scaffolding material could be precisely positioned, was evaluated. Guidance channels containing a single thin-film sheet of aligned scaffolding were shown to support robust functional nerve regeneration across extended injury gaps by minimally supplementing natural repair mechanisms. Significantly, these "thin-film enhanced nerve guidance channels" also provided the capability to guide the course of axons regenerating from a cut nerve. This capability to control axonal growth was next leveraged to create "regenerative scaffold electrodes (RSEs)" able to interface with axons regenerating from an amputated nerve. In the RSE design, low-profile arrays of interfacing electrodes were embedded within layers of aligned scaffolding material, such that regenerating axons were topographically guided by the scaffolding through the device and directly across the embedded electrodes. Chronically implanted RSEs were successfully used to record evoked neural activity from amputated nerves in an animal model. These results demonstrate that the use of topographic cues within a nerve guidance channel might offer the potential to influence the course of nerve regeneration to the advantage of a peripheral nerve interface suitable for limb amputees.
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    Delivery of thermostabilized chondroitinase ABC enhances axonal sprouting and functional recovery after spinal cord injury
    (Georgia Institute of Technology, 2009-11-10) Lee, Hyun-Jung
    Chondroitin sulfate proteoglycans (CSPGs) are one major class of axon growth inhibitors that are upregulated and accumulated around the lesion site after spinal cord injury (SCI), and result in regenerative failure. To overcome CSPG-mediated inhibition, digestion of CSPGs with chondroitinase ABC (chABC) has been explored and it has shown promising results. chABC digests glycosaminoglycan chains on CSPGs and can thereby enhance axonal regeneration and promote functional recovery when delivered at the site of injury. However, chABC has a crucial limitation; it is thermally unstable and loses its enzymatic activity rapidly at 37 ºC. Therefore, it necessitates the use of repeated injections or local infusions with a pump for days to weeks to provide fresh chABC to retain its enzymatic activity. Maintaining these infusion systems is invasive and clinically problematic. In this dissertation, three studies are reported that demonstrate our strategy to overcome current limitations of using chABC and develop a delivery system for facilitating chABC treatment after SCI: First, we enhanced the thermostability of chABC by adding trehalose, a protein stabilizer, and developed a system for its sustained local delivery in vivo. Enzymatic activity was assayed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and dimethylmethylene blue (DMMB), and conformational change of the enzyme was measured via circular dichroism (CD) with and without trehalose. When stabilized with trehalose, chABC remained enzymatically active at 37 ºC for up to 4 weeks in vitro. We developed a lipid microtube-agarose hydrogel delivery system for a sustained release and showed that chABC released from the delivery system is still functionally active and slowly released over 2 weeks in vitro. Second, the hydrogel-microtube system was used to locally deliver chABC over two weeks at the lesion site following a dorsal over hemisection injury at T10. The scaffold consisting of hydrogel and chABC loaded lipid microtubes was implanted at the top of the lesion site immediately following injury. To determine effectiveness of topical delivery of thermostabilized chABC, animal groups treated with single injection or gel scaffold implantation of chABC and penicillinase (P'ase) were included as controls. Two weeks after surgery, the functionality of released chABC and the cellular responses were examined by immunohistological analysis with 3B3, CS-56, GFAP and Wisteria floribunda agglutinin (WFA). The results demonstrated that thermostabilized chABC was successfully delivered slowly and locally without the need for an indwelling catheter by using the hydrogel-microtube delivery system in vivo. The results demonstrated that released chABC from the gel scaffold effectively digested CSPGs, and therefore, there were significant differences in CSPG digestion at the lesion site between groups treated with chABC loaded microtube-hydrogel scaffolds and controls. Third, a long term in vivo study (45 days) was conducted to examine axonal sprouting/regeneration and functional recovery with both a single treatment each of microtube loaded chABC or Neurotrophin-3 (NT-3), and a combination of them by using the hydrogel-microtube delivery system. Over the long term study period, the treated animals showed significant improvement in locomotor function and more sprouting of cholera toxin B subunit (CTB)-positive ascending dorsal column fibers and 5-HT serotonergic fibers around the lesion site. We demonstrated that this significant improvement of chABC thermostability facilitates the development of a minimally invasive method for sustained, local delivery of chABC that is potentially a useful and effective approach for treating SCI. In addition to that, we demonstrated that combinatorial therapy with chABC and neurotrophic factors could provide a synergistic effect on axonal regrowth and functional recovery after SCI.