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Investigating the Effect of a Hyaluronic Acid-Laminin Hydrogel on Inflammation After Traumatic Brain Injury

Description
Traumatic brain injury (TBI) is a leading cause of death in individuals under the age of 45, resulting in over 50,000 deaths each year. Over 80,000 TBI patients report long-term deficits consisting of motor or cognitive dysfunctions due to TBI pathophysiology. The biochemical secondary injury triggers a harmful inflammatory cascade,…

Traumatic brain injury (TBI) is a leading cause of death in individuals under the age of 45, resulting in over 50,000 deaths each year. Over 80,000 TBI patients report long-term deficits consisting of motor or cognitive dysfunctions due to TBI pathophysiology. The biochemical secondary injury triggers a harmful inflammatory cascade, gliosis, and astrocyte activation surrounding the injury lesion, and no current treatments exist to alleviate these underlying pathologies. In order to mitigate the negative inflammatory effects of the secondary injury, we created a hydrogel comprised of hyaluronic acid (HA) and laminin, and we hypothesized that the anti-inflammatory properties of HA will decrease astrocyte activation and inflammation after TBI. C57/BL6 mice were subjected to mild-to-moderate CCI. Three days following injury, mice were treated with injection of vehicle or HA-Laminin hydrogel. Mice were sacrificed at three and seven days post injection and analyzed for astrocyte and inflammatory responses. In mice treated with vehicle injections, astrocyte activation was significantly increased at three days post-transplantation in the injured cortex and injury lesion. However, mice treated with the HA-Laminin hydrogel experienced significantly reduced acute astrocyte activation at the injury site three days post transplantation. Interestingly, there were no significant differences in astrocyte activation at seven days post treatment in either group. Although the microglial and macrophage response remains to be investigated, our data suggest that the HA-Laminin hydrogel demonstrates potential for TBI therapeutics targeting inflammation, including acute modulation of the astrocyte, microglia, and macrophage response to TBI.
ContributorsGoddery, Emma Nicole (Author) / Stabenfeldt, Sarah (Thesis director) / Addington, Caroline (Committee member) / Harrington Bioengineering Program (Contributor) / Barrett, The Honors College (Contributor)
Created2016-05

The Development of a Microgel/Hydrogel Composite System for TBI Treatment

Description

Traumatic brain injury (TBI), a neurological condition that negatively affects neural capabilities, occurs when a blunt trauma impacts the head. Following the initial injury that immediately impacts neural cell function and survival, a series of secondary injury events lead to substantial sustained inflammation for weeks to years post-injury. To develo…

Traumatic brain injury (TBI), a neurological condition that negatively affects neural capabilities, occurs when a blunt trauma impacts the head. Following the initial injury that immediately impacts neural cell function and survival, a series of secondary injury events lead to substantial sustained inflammation for weeks to years post-injury. To develop TBI treatments that may stimulate regenerative processes, a novel drug delivery system that efficiently delivers the appropriate drug/payload to injured tissue is crucial. Hyaluronic acid (HA) hydrogels are attractive when developing a biomaterial for tissue reparation and regeneration. HA is a natural polymer with physicochemical properties that can be tuned to match the properties of the extracellular matrix (ECM) of the many tissues including the central nervous system (CNS). Here, the project objective was to develop a HA hydrogel system for local delivery of a biological payload; this objective was completed by employing a composite system with two parts. The first part is an injectable, shear-thinning bulk hydrogel, and the second is microgels for loading biological payloads. The bulk hydrogel was composed of cyclodextrin modified HA (Cd-HA) and adamantane modified HA (Ad-HA) that give rise to guest-host interactions that facilitate physical crosslinking. The microgel, composed of norbornene-HA (Nor-HA) and sulfated-HA, crosslink via chemical crosslinks upon activation of a UV photoinitiator. The sulfated-HA microgels facilitate loading of biological payloads by mimicking heparin binding sites via the conjugated sulfated group. Neuregulin I, an epidermal growth factor with neuroprotective properties, is one such protein with a heparin binding domain that may be retained in the sulfated-HA microgels. Specifically, the project focused on mechanical testing of this composite microgel/hydrogel system and also developing protein affinity assays.
ContributorsKylat, Anna (Author) / Stabenfeldt, Sarah (Thesis director) / Holloway, Julianne (Committee member) / Jensen, Gregory (Committee member) / Barrett, The Honors College (Contributor) / School of Mathematical and Statistical Sciences (Contributor) / Harrington Bioengineering Program (Contributor)
Created2023-05
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High-throughput platforms for tumor dormancy-relapse and biomolecule binding using aminoglycoside-derived hydrogels

Description
Relapse after tumor dormancy is one of the leading causes of cancer recurrence that ultimately leads to patient mortality. Upon relapse, cancer manifests as metastases that are linked to almost 90% cancer related deaths. Capture of the dormant and relapsed tumor phenotypes in high-throughput will allow for rapid targeted drug…

Relapse after tumor dormancy is one of the leading causes of cancer recurrence that ultimately leads to patient mortality. Upon relapse, cancer manifests as metastases that are linked to almost 90% cancer related deaths. Capture of the dormant and relapsed tumor phenotypes in high-throughput will allow for rapid targeted drug discovery, development and validation. Ablation of dormant cancer will not only completely remove the cancer disease, but also will prevent any future recurrence. A novel hydrogel, Amikagel, was developed by crosslinking of aminoglycoside amikacin with a polyethylene glycol crosslinker. Aminoglycosides contain abundant amount of easily conjugable groups such as amino and hydroxyl moieties that were crosslinked to generate the hydrogel. Cancer cells formed 3D spheroidal structures that underwent near complete dormancy on Amikagel high-throughput drug discovery platform. Due to their dormant status, conventional anticancer drugs such as mitoxantrone and docetaxel that target the actively dividing tumor phenotype were found to be ineffective. Hypothesis driven rational drug discovery approaches were used to identify novel pathways that could sensitize dormant cancer cells to death. Strategies were used to further accelerate the dormant cancer cell death to save time required for the therapeutic outcome.

Amikagel’s properties were chemo-mechanically tunable and directly impacted the outcome of tumor dormancy or relapse. Exposure of dormant spheroids to weakly stiff and adhesive formulation of Amikagel resulted in significant relapse, mimicking the response to changes in extracellular matrix around dormant tumors. Relapsed cells showed significant differences in their metastatic potential compared to the cells that remained dormant after the induction of relapse. Further, the dissertation discusses the use of Amikagels as novel pDNA binding resins in microbead and monolithic formats for potential use in chromatographic purifications. High abundance of amino groups allowed their utilization as novel anion-exchange pDNA binding resins. This dissertation discusses Amikagel formulations for pDNA binding, metastatic cancer cell separation and novel drug discovery against tumor dormancy and relapse.
ContributorsGrandhi, Taraka Sai Pavan (Author) / Rege, Kaushal (Thesis advisor) / Meldrum, Deirdre R (Thesis advisor) / Stabenfeldt, Sarah (Committee member) / Caplan, Michael (Committee member) / Tian, Yanqing (Committee member) / Arizona State University (Publisher)
Created2016

Optimizing Recombinant Protein Production for Domain Antibodies: Proof-of-Concept

Description
Recent studies in traumatic brain injury (TBI) have found a temporal window where therapeutics on the nanometer scale can cross the blood-brain barrier and enter the parenchyma. Developing protein-based therapeutics is attractive for a number of reasons, yet, the production pipeline for high yield and consistent bioactive recombinant proteins remains…

Recent studies in traumatic brain injury (TBI) have found a temporal window where therapeutics on the nanometer scale can cross the blood-brain barrier and enter the parenchyma. Developing protein-based therapeutics is attractive for a number of reasons, yet, the production pipeline for high yield and consistent bioactive recombinant proteins remains a major obstacle. Previous studies for recombinant protein production has utilized gram-negative hosts such as Escherichia coli (E. coli) due to its well-established genetics and fast growth for recombinant protein production. However, using gram-negative hosts require lysis that calls for additional optimization and also introduces endotoxins and proteases that contribute to protein degradation. This project directly addressed this issue and evaluated the potential to use a gram-positive host such as Brevibacillus choshinensis (Brevi) which does not require lysis as the proteins are expressed directly into the supernatant. This host was utilized to produce variants of Stock 11 (S11) protein as a proof-of-concept towards this methodology. Variants of S11 were synthesized using different restriction enzymes which will alter the location of protein tags that may affect production or purification. Factors such as incubation time, incubation temperature, and media were optimized for each variant of S11 using a robust design of experiments. All variants of S11 were grown using optimized parameters prior to purification via affinity chromatography. Results showed the efficiency of using Brevi as a potential host for domain antibody production in the Stabenfeldt lab. Future aims will focus on troubleshooting the purification process to optimize the protein production pipeline.
ContributorsEmbrador, Glenna Bea Rebano (Author) / Stabenfeldt, Sarah (Thesis director) / Plaisier, Christopher (Committee member) / Harrington Bioengineering Program (Contributor, Contributor) / Barrett, The Honors College (Contributor)
Created2019-05