Browsing by Author "VandeVord, Pamela J."

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    A 3-D Finite-Element Minipig Model to Assess Brain Biomechanical Responses to Blast Exposure
    Sundaramurthy, Aravind; Kote, Vivek Bhaskar; Pearson, Noah; Boiczyk, Gregory M.; McNeil, Elizabeth M.; Nelson, Allison J.; Subramaniam, Dhananjay Radhakrishnan; Rubio, Jose E.; Monson, Kenneth; Hardy, Warren N.; VandeVord, Pamela J.; Unnikrishnan, Ginu; Reifman, Jaques (Frontiers, 2021-12-17)
    Despite years of research, it is still unknown whether the interaction of explosion-induced blast waves with the head causes injury to the human brain. One way to fill this gap is to use animal models to establish “scaling laws” that project observed brain injuries in animals to humans. This requires laboratory experiments and high-fidelity mathematical models of the animal head to establish correlates between experimentally observed blast-induced brain injuries and model-predicted biomechanical responses. To this end, we performed laboratory experiments on Göttingen minipigs to develop and validate a three-dimensional (3-D) high-fidelity finite-element (FE) model of the minipig head. First, we performed laboratory experiments on Göttingen minipigs to obtain the geometry of the cerebral vasculature network and to characterize brain-tissue and vasculature material properties in response to high strain rates typical of blast exposures. Next, we used the detailed cerebral vasculature information and species-specific brain tissue and vasculature material properties to develop the 3-D high-fidelity FE model of the minipig head. Then, to validate the model predictions, we performed laboratory shock-tube experiments, where we exposed Göttingen minipigs to a blast overpressure of 210 kPa in a laboratory shock tube and compared brain pressures at two locations. We observed a good agreement between the model-predicted pressures and the experimental measurements, with differences in maximum pressure of less than 6%. Finally, to evaluate the influence of the cerebral vascular network on the biomechanical predictions, we performed simulations where we compared results of FE models with and without the vasculature. As expected, incorporation of the vasculature decreased brain strain but did not affect the predictions of brain pressure. However, we observed that inclusion of the cerebral vasculature in the model changed the strain distribution by as much as 100% in regions near the interface between the vasculature and the brain tissue, suggesting that the vasculature does not merely decrease the strain but causes drastic redistributions. This work will help establish correlates between observed brain injuries and predicted biomechanical responses in minipigs and facilitate the creation of scaling laws to infer potential injuries in the human brain due to exposure to blast waves.
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    Advancing Transcranial Focused Ultrasound for Noninvasive Neuromodulation of Human Cortex
    Mueller, Jerel Keith (Virginia Tech, 2015-09-09)
    Ultrasound waves are mechanical undulations above the threshold for human hearing, and have been used widely in both the human body and brain for diagnostic and therapeutic purposes. Ultrasound can be controlled using specially designed transducers into a focus of a few millimeters in diameter. Low intensity ultrasound, such as used in imaging applications, appears to be safe in adults. It is also known that ultrasound waves can penetrate through the skull and be focused within the brain for ablation purposes, employing the heat generation properties of high intensity focused ultrasound. High intensity focused ultrasound is thus used to irreversibly ablate brain tissue in localized areas without observable damage to intermediate tissue and vasculature. Ablation with high intensity focused ultrasound guided by magnetic resonance imaging is used for abolishing brain tumors, and experimentally for pain. Low intensity ultrasound can be utilized beyond imaging in neuroscience and neurology by focusing the ultrasound beam to investigate the structure and function of discrete brain circuits. In contrast to high intensity focused ultrasound, the effects of low intensity focused ultrasound on neurons are reversible. Considering the volume of work on high intensity focused ultrasound, low intensity focused ultrasound remains decidedly underdeveloped. Given the great potential for impact of low intensity focused ultrasound in both clinical and scientific neuromodulation applications, we sought to advance the use of low intensity focused ultrasound for noninvasive, transcranial neuromodulation of human cortex. This dissertation contains novel research on the use of low intensity transcranial focused ultrasound for noninvasive neuromodulation of human cortex. The importance of mechanical forces in the nervous system is highlighted throughout to expand beyond the stigma that nervous function is governed chiefly by electrical and chemical means. Methods of transcranial focused ultrasound are applied to significantly modulate human cortical function, shown using electroencephalographic recordings and behavioral investigations of sensory discrimination performance. This dissertation also describes computational models used to investigate the insertion behavior of ultrasound across various tissues in the context of transcranial neuromodulation, as ultrasound's application for neuromodulation is relatively new and crudely understood. These investigations are critical for the refinement of device design and the overall advancement of ultrasound methods for noninvasive neuromodulation.
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    Age-relevant in vitro models may lead to improved translational research for traumatic brain injury
    Dickerson, Michelle R.; Guilhaume-Correa, Fernanda; Strickler, Jessica; VandeVord, Pamela J. (Elsevier, 2022-06)
    Traumatic brain injury (TBI) is a major health problem affecting both children and adults. Although TBI studies have been focused on neurons, glial cells play an important role in neuropathology following injury. As the consequences of TBI are age-dependent, it is essential that in vitro and in vivo models are fully representative of clinical outcomes. Traditionally, in vitro models that focused on TBI-induced glial cell dysfunction use primary cells isolated from neonatal rodents, or cell lines. These models are widely used to elucidate molecular pathways affected by the injury; however, they fail to account for age-related differences. As glial characteristics are known to change during maturation, it is important to explore new age-relevant in vitro models leading to improved translation research and advancements in therapeutic strategies for TBI.
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    Applying Proteomics and Computational Approaches to Identify Novel Targets in Blast-Associated Post-Traumatic Epilepsy
    Browning, Jack L.; Wilson, Kelsey A.; Shandra, Oleksii; Wei, Xiaoran; Mahmutovic, Dzenis; Maharathi, Biswajit; Robel, Stefanie; VandeVord, Pamela J.; Olsen, Michelle L. (MDPI, 2024-03-01)
    Traumatic brain injury (TBI) can lead to post-traumatic epilepsy (PTE). Blast TBI (bTBI) found in Veterans presents with several complications, including cognitive and behavioral disturbances and PTE; however, the underlying mechanisms that drive the long-term sequelae are not well understood. Using an unbiased proteomics approach in a mouse model of repeated bTBI (rbTBI), this study addresses this gap in the knowledge. After rbTBI, mice were monitored using continuous, uninterrupted video-EEG for up to four months. Following this period, we collected cortex and hippocampus tissues from three groups of mice: those with post-traumatic epilepsy (PTE+), those without epilepsy (PTE−), and the control group (sham). Hundreds of differentially expressed proteins were identified in the cortex and hippocampus of PTE+ and PTE− relative to sham. Focusing on protein pathways unique to PTE+, pathways related to mitochondrial function, post-translational modifications, and transport were disrupted. Computational metabolic modeling using dysregulated protein expression predicted mitochondrial proton pump dysregulation, suggesting electron transport chain dysregulation in the epileptic tissue relative to PTE−. Finally, data mining enabled the identification of several novel and previously validated TBI and epilepsy biomarkers in our data set, many of which were found to already be targeted by drugs in various phases of clinical testing. These findings highlight novel proteins and protein pathways that may drive the chronic PTE sequelae following rbTBI.
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    Astrocyte Mechano-Activation by High-Rate Overpressure Involves Alterations in Structural and Junctional Proteins
    Hlavac, Nora; VandeVord, Pamela J. (Frontiers, 2019-02-22)
    Primary blast neurotrauma represents a unique injury paradigm characterized by high-rate overpressure effects on brain tissue. One major hallmark of blast neurotrauma is glial reactivity, notably prolonged astrocyte activation. This cellular response has been mainly defined in primary blast neurotrauma by increased intermediate filament expression. Because the intermediate filament networks physically interface with transmembrane proteins for junctional support, it was hypothesized that cell junction regulation is altered in the reactive phenotype as well. This would have implications for downstream transcriptional regulation via signal transduction pathways like nuclear factor kappa-light-chain-enhancer of activated B cells (NF-kB). Therefore, a custom high-rate overpressure simulator was built for in vitro testing using mechanical conditions based on intracranial pressure measurements in a rat model of blast neurotrauma. Primary rat astrocytes were exposed to isolated high-ratemechanical stimulation to study cell junction dynamics in relation to their mechano-activation. First, a time course for “classical” features of reactivity was devised by evaluation of glial fibrillary acidic protein (GFAP) and proliferating cell nuclear antigen (PCNA) expression. This was followed by gene and protein expression for both gap junction (connexins) and anchoring junction proteins (integrins and cadherins). Signal transduction analysis was carried out by nuclear localization of two molecules, NF-kB p65 and mitogen-activated protein kinase (MAPK) p38. Results indicated significant increases in connexin-43 expression and PCNA first at 24 h post-overpressure (p < 0.05), followed by structural reactivity (via increased GFAP, p < 0.05) corresponding to increased anchoring junction dynamics at 48 h post-overpressure (p < 0.05). Moreover, increased phosphorylation of focal adhesion kinase (FAK) was observed in addition to increased nuclear localization of both p65 and p38 (p < 0.05) during the period of structural reactivity. To evaluate the transcriptional activity of p65 in the nucleus, electrophoretic mobility shift assay was conducted for a binding site on the promoter region for intracellular adhesion molecule-1 (ICAM-1), an antagonist of tight junctions. A significant increase in the interaction of nuclear proteins with the NF-kB site on the ICAM-1 corresponded to increased gene and protein expression of ICAM-1 (p < 0.05).
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    Attributes of Astrocyte Response to Mechano-Stimulation by High-Rate Overpressure
    Hlavac, Nora (Virginia Tech, 2018-11-29)
    Blast neurotrauma represents a significant mode of traumatic injury to the brain. The incidence of blast neurotrauma is particularly high amongst military combat personnel and can be debilitating and endure clinically for years after injury is sustained. Mechanically, blast represents a unique and complex loading paradigm associated with compressive shock waves that propagate out from an explosive event and interact with the head and other organs through high-rate loading. When subjected to such insult, brain cells undergo characteristic injury responses which include neuroinflammation, oxidative stress, edema and persistent glial activation. These features of the injury have emerged as important mediators of the chronic brain damage that results from blast. Astrocytes have emerged as a potential therapeutic target because of their ubiquitous roles in brain homeostasis, tissue integrity and cognitive function. This glial subtype has a characteristic reactive response to mechanical trauma of various modes. In this work, custom in vitro injury devices were used to characterize functional models of astrocyte reactivity to high-rate insult to study mechano-stimulation mechanisms associated with the reactive phenotype. The working hypothesis was that high-rate overpressure exposure would cause metabolic aberrations, cell junction changes, and adhesion signal transduction activation, all of which would contribute to astrocyte response and reactivity. Astrocyte cultures were exposed to a 20 psi high-rate overpressure scheme using an underwater explosion-driven device. Astrocytes experienced dynamic energetic fluctuations in response to overpressure which were followed by the assumption of a classically defined reactive phenotype. Results indicated specific roles for cationic transduction, cell junction dynamics (gap junction and anchoring junctions) and downstream signal transduction mechanisms associated with adhesion alterations in onset of the astrocyte reactive phenotype. Investigation into adhesion signaling regulation by focal adhesion kinase in 2D and 3D cultures was also explored to better understand cellular reactivity as a function of extracellular environment. Additionally, another underwater in vitro device was built to study combination effects from overpressure and fluid shear associated with insult. Overall, the combined studies offer multiple mechanisms by which to explore molecular targets for harnessing astrocytes' potential for repair after traumatic injury to the brain.
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    Atypical Neurogenesis, Astrogliosis, and Excessive Hilar Interneuron Loss Are Associated with the Development of Post-Traumatic Epilepsy
    Gudenschwager-Basso, Erwin Kristobal; Shandra, Oleksii; Volanth, Troy; Patel, Dipan C.; Kelly, Colin; Browning, Jack L.; Wei, Xiaoran; Harris, Elizabeth A.; Mahmutovic, Dzenis; Kaloss, Alexandra M.; Correa, Fernanda Guilhaume; Decker, Jeremy; Maharathi, Biswajit; Robel, Stefanie; Sontheimer, Harald; VandeVord, Pamela J.; Olsen, Michelle L.; Theus, Michelle H. (MDPI, 2023-04-25)
    Background: Traumatic brain injury (TBI) remains a significant risk factor for post-traumatic epilepsy (PTE). The pathophysiological mechanisms underlying the injury-induced epileptogenesis are under investigation. The dentate gyrus—a structure that is highly susceptible to injury—has been implicated in the evolution of seizure development. Methods: Utilizing the murine unilateral focal control cortical impact (CCI) injury, we evaluated seizure onset using 24/7 EEG video analysis at 2–4 months post-injury. Cellular changes in the dentate gyrus and hilus of the hippocampus were quantified by unbiased stereology and Imaris image analysis to evaluate Prox1-positive cell migration, astrocyte branching, and morphology, as well as neuronal loss at four months post-injury. Isolation of region-specific astrocytes and RNA-Seq were performed to determine differential gene expression in animals that developed post-traumatic epilepsy (PTE+) vs. those animals that did not (PTE−), which may be associated with epileptogenesis. Results: CCI injury resulted in 37% PTE incidence, which increased with injury severity and hippocampal damage. Histological assessments uncovered a significant loss of hilar interneurons that coincided with aberrant migration of Prox1-positive granule cells and reduced astroglial branching in PTE+ compared to PTE− mice. We uniquely identified Cst3 as a PTE+-specific gene signature in astrocytes across all brain regions, which showed increased astroglial expression in the PTE+ hilus. Conclusions: These findings suggest that epileptogenesis may emerge following TBI due to distinct aberrant cellular remodeling events and key molecular changes in the dentate gyrus of the hippocampus.
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    Automated Rat Grimace Scale for the Assessment of Pain
    Arnold, Brendan Elliot (Virginia Tech, 2023-06-21)
    Pain is a complex neuro-psychosocial experience that is internal and private making it difficult to assess in both humans and animals. In research approximately 95% of animal models use rodents, with rats being among the most common for pain studies [3]. However, traditional assessments of the pain response struggle to demonstrate that the behaviors are a direct measurement of pain. The rat grimace scale (RGS) was developed based on facial action coding systems (FACS) which have known utility in non-verbal humans [6, 9]. The RGS measures facial action units of orbital tightening, ear changes, nose flattening, and whisker changes in an attempt to quantify the pain behaviors of the rat. These action units are measured on frontal images of rats with their face in clear view on a scale of 0-2, then summed together. The total score is then averaged to find a final value for RGS between 0-2. Currently, the software program Rodent Face Finder® can extract frontal face images. However, the RGS scores are still manually recorded which is a labor-intensive process, requiring hours of training. Furthermore, the scoring can be subjective, with differences existing between researchers and lab groups. The primary aim of this study is to develop an automated system that can detect action unit regions and generate a RGS score for each image. To accomplish this objective, a YOLOv5 object detector and Vision Transformers (ViT) for classification were trained on a dataset of frontal-facing images extracted using Rodent Face Finder®. Subsequently, the model was then validated using a RGS test for blast traumatic brain injury (bTBI). The validation dataset consisted of 40 control images of uninjured rats, 40 images from the bTBI study on the day of injury, and 40 images 1-month post-injury. All 120 images in the validation set were then manually graded for RGS and tested using the automated RGS system. The results indicated that the automated RGS system accurately and efficiently graded the images with minimal variation in results compared to human graders in just 1/14th of the time. This system provides a fast and reliable method to extract meaningful information of rats' internal pain state. Furthermore, the study presents an avenue for future research into real-time pain monitoring.
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    The Biomechanical Response of the Eye, Face, and Orbit to Primary Blast Overpressure
    Alphonse, Vanessa D. (Virginia Tech, 2015-12-08)
    Combat-related blast injuries are occurring more frequently with the increased use of improvised explosive devices in current military conflicts. Though much research has focused on how the body responds to the relatively low loading rates associated with blunt trauma, little is known regarding the response of the body to the higher loading rates associated with blast trauma. While soldiers are surviving once-lethal blast events due to enhanced protective equipment, injuries such as those to the eye and face that were once considered inconsequential, can now be detrimental to long-term healthcare costs and quality of life. Although it is suggested primary blast overpressure (i.e., the shock wave) can cause severe eye injuries, there remains few empirical data in the literature that confirms this. Adding to this, there are currently no testing standards to assess the effectiveness of personal protective equipment during blast exposure. Expanding upon traditional research techniques within the field of injury biomechanics, the research in this dissertation focuses specifically on developing experimental and physical models of the eye, face, and orbit for blast overpressure exposure. Foremost, a porcine eye model is used to quantify eye injury risk from blast overpressure exposure. Subsequently, these biomechanical data are used to develop a physical model of the eye that can be used in lieu of cadaver specimens for blunt and blast loading. Lastly, military spectacles and goggles are examined for effectiveness at protecting the eye during blast exposure. Combined with detailed computer-aided design geometries, these data can be used to validate computational models of the eye, orbit, and face to blast loading. Results from these tests support one theory that shock waves may enter the skull through the orbit, alluding to future work that is essential to more fully understanding the physiological response of the brain and ocular motor system to blast exposure. Ultimately, the experimental methods and analysis techniques disseminated herein serve as a framework for future experimental work related to blast and other high-rate loading scenarios.
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    Blasted Flies and Nanoparticles for TBI
    Hockey, Kevin S.; Sholar, Christopher A.; Sajja, Venkata Siva Sai Sujith; Hubbard, W. Brad; Thorpe, Chevon; VandeVord, Pamela J.; Rzigalinski, Beverly A. (Brain Injuries and Biomechanics Symposium, 2013-09-19)
    This presentation briefly summaries two major areas of work in our lab, development of a Drosophila model of blast injury and treatment of traumatic brain injury (TBI) with cerium oxide nanoparticles. First, we discuss the design, methodology, and results for the Drosophila blast model, and its relevance to human head injury. Briefly, we found that the Drosophila model was able to reproduce the decreased lifespan and early death seen in military personnel exposed to repetitive mild blast and NFL players exposed to repeated mild head injury. Next we discuss our in vitro and in vivo work with cerium oxide nanoparticles as neuroprotective and regenerative agents for treatment of TBI. Using a tissue culture model for TBI, we found that cerium oxide nanoparticles, delivered up to 6 hrs. post-injury, improved neuronal survival and maintained near-normal glutamate signaling in neurons of mixed organotypic brain cell cultures. In vivo, we found that delivery of cerium oxide nanoparticles prior to lateral fluid percussion brain injury in the rat, improved motor performance, learning and memory.
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    Cerium Oxide Nanoparticles Improve Outcome after In Vitro and In Vivo Mild Traumatic Brain Injury
    Bailey, Zachary S.; Nilson, Eric; Bates, John A.; Oyalowo, Adewole; Hockey, Kevin S.; Sajja, Venkata Siva Sai Sujith; Thorpe, Chevon N.; Rogers, Heidi; Dunn, Bryce; Frey, Aaron S.; Billings, Marc J.; Sholar, Christopher A.; Hermundstad, Amy; Kumar, Challa; VandeVord, Pamela J.; Rzigalinski, Beverly A. (2020-06-15)
    Mild traumatic brain injury results in aberrant free radical generation, which is associated with oxidative stress, secondary injury signaling cascades, mitochondrial dysfunction, and poor functional outcome. Pharmacological targeting of free radicals with antioxidants has been examined as an approach to treatment, but has met with limited success in clinical trials. Conventional antioxidants that are currently available scavenge a single free radical before they are destroyed in the process. Here, we report for the first time that a novel regenerative cerium oxide nanoparticle antioxidant reduces neuronal death and calcium dysregulation after in vitro trauma. Further, using an in vivo model of mild lateral fluid percussion brain injury in the rat, we report that cerium oxide nanoparticles also preserve endogenous antioxidant systems, decrease macromolecular free radical damage, and improve cognitive function. Taken together, our results demonstrate that cerium oxide nanoparticles are a novel nanopharmaceutical with potential for mitigating neuropathological effects of mild traumatic brain injury and modifying the course of recovery.
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    Chronic Anxiety- and Depression-Like Behaviors Are Associated With Glial-Driven Pathology Following Repeated Blast Induced Neurotrauma
    Dickerson, Michelle R.; Murphy, Susan F.; Urban, Michael J.; White, Zakar; VandeVord, Pamela J. (Frontiers, 2021-12-10)
    Long-term neuropsychiatric impairments have become a growing concern following blast-related traumatic brain injury (bTBI) in active military personnel and Veterans. Neuropsychiatric impairments such as anxiety and depression are common comorbidities that Veterans report months, even years following injury. To understand these chronic behavioral outcomes following blast injury, there is a need to study the link between anxiety, depression, and neuropathology. The hippocampus and motor cortex (MC) have been regions of interest when studying cognitive deficits following blast exposure, but clinical studies of mood disorders such as major depressive disorder (MDD) report that these two regions also play a role in the manifestation of anxiety and depression. With anxiety and depression being common long-term outcomes following bTBI, it is imperative to study how chronic pathological changes within the hippocampus and/or MC due to blast contribute to the development of these psychiatric impairments. In this study, we exposed male rats to a repeated blast overpressure (~17 psi) and evaluated the chronic behavioral and pathological effects on the hippocampus and MC. Results demonstrated that the repeated blast exposure led to depression-like behaviors 36 weeks following injury, and anxiety-like behaviors 2-, and 52-weeks following injury. These behaviors were also correlated with astrocyte pathology (glial-fibrillary acid protein, GFAP) and dendritic alterations (Microtubule-Associated Proteins, MAP2) within the hippocampus and MC regions at 52 weeks. Overall, these findings support the premise that chronic glial pathological changes within the brain contribute to neuropsychiatric impairments following blast exposure.
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    Development of a Minipig Model of BINT From Blast Exposure Using a Repeatable Mobile Shock Expansion Tube
    McNeil, Elizabeth M.; Walilko, Timothy; Hulbert, Lindsey E.; VanMeter, John W.; LaConte, Stephen M.; VandeVord, Pamela J.; Zai, Laila; Bentley, Timothy B. (Oxford University Press, 2021-10-22)
    Introduction: The Office of Naval Research (ONR) sponsored the Blast Load Assessment Sense and Test (BLAST) program to provide an approach to operationally relevant monitoring and analysis of blast exposure for optimization of service member performance and health. Of critical importance in this effort was the development of a standardized methodology for preclinical large animal studies that can reliably produce outcome measures that cannot be measured in human studies to support science-based guidelines. The primary advantage of this approach is that, because animal studies report physiological measures that correlate with human neuropathology, these data can be used to evaluate potential risks to service members by accounting for the anatomical and physiological differences between humans and large animal models. This article describes the methodology used to generate a comprehensive outcome measure dataset correlated with controlled blast exposure. Methods and materials: To quantify outcomes associated with a single exposure to blast, 23 age- and weight-matched Yucatan minipigs were exposed to a single blast event generated by a large-bore, compressed gas shock tube. The peak pressure ranged from 280 to 525 kPa. After a post-exposure 72-hour observation period, the physiological response was quantified using a comprehensive set of neurological outcome measures that included neuroimaging, histology, and behavioral measures. Responses of the blast-exposed animals were compared to the sham-treated cohort to identify statistically significant and physiologically relevant differences between the two groups. Results: Following a single exposure, the minipigs were assessed for structural, behavioral, and cellular changes for 3 days after exposure. The following neurological changes were observed: Structural- Using Diffusion Tensor Imaging, a statistically significant decrement (P < .001) in Fractional Anisotropy across the entire volume of the brain was observed when comparing the exposed group to the sham group. This finding indicates that alterations in brain tissue following exposure are not focused at a single location but instead a diffuse brain volume that can only be observed through a systematic examination of the neurological tissue. Cellular- The histopathology results from several large white matter tract locations showed varied cellular responses from six different stains. Using standard statistical methods, results from stains such as Fluoro-Jade C and cluster of differentiation 68 in the hippocampus showed significantly higher levels of neurodegeneration and increased microglia/macrophage activation in blast-exposed subjects. However, other stains also indicated increased response, demonstrating the need for multivariate analysis with a larger dataset. Behavioral- The behavior changes observed were typically transient; the animals' behavior returned to near baseline levels after a relatively short recovery period. Despite behavioral recovery, the presence of active neurodegenerative and inflammatory responses remained. Conclusions: The results of this study demonstrate that (1) a shock tube provides an effective tool for generating repeatable exposures in large animals and (2) exposure to blast overpressure can be correlated using a combination of imaging, behavioral, and histological analyses. This research demonstrates the importance of using multiple physiological indicators to track blast-induced changes in minipigs. The methodology and findings from this effort were central to developing machine-learning models to inform the development of blast exposure guidelines.
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    Development of MRI-based Yucatan Minipig Brain Template
    Norris, Caroline N. (Virginia Tech, 2019-04-05)
    Yucatan minipigs have become increasingly common animal models in neuroscience where recent studies, investigating blast-induced traumatic brain injury, stroke, and glioblastoma, aim to uncover brain injury mechanisms [1-3]. Magnetic Resonance Imaging (MRI) has the potential to validate and optimize unknown parameters in controlled populations. The key to group-level MRI analysis within a species is to align (or register) subject scans to the same volumetric space using a brain template. However, large animal brain templates are lacking, which limits the use of MRI as an effective research tool to study group effects. The objective of this study was to create an MRI-based Yucatan minipig brain template allowing for uniform group-level analysis of this animal model in a standard volumetric space to characterize brain mechanisms. To do this, 5-7 month old, male Yucatan minipigs were scanned using a 3 Tesla whole-body scanner (Siemens AG, Erlangen) in accordance with IACUC. T1-weighted anatomical volumes (resolution = 1×1×1 mm3; TR = 2300 ms; TE= 2.89 ms; TI = 900 ms; FOV = 256 mm2 ; FA = 8 deg) were collected with a three-dimensional magnetization prepared rapid acquisition gradient echo (MPRAGE) pulse sequence [4]. The volumes were preprocessed, co-registered, and averaged using both linear and non-linear registration algorithms in AFNI [5] to create four templates (n=58): linear brain, non-linear brain, linear head, and non-linear head. To validate the templates, tissue probability maps (TPMs) and variance maps were created, and landmark variation was measured. TPMs computed in FSL [6] and AFNI show enhanced tissue probability and contrast in the non-linear template. Additionally, variance maps showed a more uniform spatial variance in the non-linear template compared to the linear. Registration variation within the brain template was within 1.5 mm and displayed improved landmark variation in the non-linear brain template. External evaluation subjects (n=12), not included in the template, were registered to the four templates to assess functionality. The results indicate that the developed templates provide acceptable registration accuracy to enable population comparisons. With these templates, researchers will be able to use MRI as a tool to further neurological discovery and collaborate in a uniform space.
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    Distinguishing the Unique Neuropathological Profile of Blast Polytrauma
    Hubbard, W. Brad; Greenberg, Shaylen; Norris, Carly; Eck, Joseph; Lavik, Erin; VandeVord, Pamela J. (Hindawi, 2017-03-23)
    Traumatic brain injury sustained after blast exposure (blast-induced TBI) has recently been documented as a growing issue for military personnel. Incidence of injury to organs such as the lungs has decreased, though current epidemiology still causes a great public health burden. In addition, unprotected civilians sustain primary blast lung injury (PBLI) at alarming rates. Often, mild-to-moderate cases of PBLI are survivable with medical intervention, which creates a growing population of survivors of blast-induced polytrauma (BPT) with symptoms from blast-induced mild TBI (mTBI). Currently, there is a lack of preclinical models simulating BPT, which is crucial to identifying unique injury mechanisms of BPT and its management. To meet this need, our group characterized a rodent model of BPT and compared results to a blast-induced mTBI model. Open field (OF) performance trials were performed on rodents at 7 days after injury. Immunohistochemistry was performed to evaluate cellular outcome at day seven following BPT. Levels of reactive astrocytes (GFAP), apoptosis (cleaved caspase-3 expression), and vascular damage (SMI-71) were significantly elevated in BPT compared to blast-induced mTBI. Downstream markers of hypoxia (HIF-1α and VEGF) were higher only after BPT. This study highlights the need for unique therapeutics and prehospital management when handling BPT.
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    Effects of Keratin Biomaterial Therapeutics on Cellular and Inflammatory Mechanisms in Injury and Disease Models
    Waters, Michele (Virginia Tech, 2018-06-11)
    Keratins are fibrous structural proteins found in human hair that have been used to develop bioactive and biocompatible constructs for a wide variety of tissue engineering and healthcare applications. Their ubiquity, capacity for self-assembly, ease of use and versatility in blended materials, and ability to modulate cell behavior and promote tissue ingrowth have made keratins well-suited for the development of regenerative therapies. In particular, keratins have demonstrated bioactivity in both in-vivo and in-vitro studies, by altering immune and stem cell phenotype and function and promoting an anti-inflammatory/wound healing environment. This work seeks to build on previous research by investigating the ability of low and high molecular weight keratins to augment anti-inflammatory primary macrophage phenotypes and examining the influence of keratin biomaterials on cellular and inflammatory mechanisms in two models of injury and disease. Rodent models of blast induced neurotrauma (BINT) and severe osteoporosis were used to inform the development of 2D and 3D in-vitro models of macrophage/endothelial cell injury and osteogenic differentiation respectively. Keratin biomaterials exhibited some potential to alter macrophage and endothelial cell dynamics following blast, specifically by promoting anti-inflammatory (M2c-like) macrophage polarization and diminishing endothelial cell injury responses (i.e. endothelial glycocalyx shedding). A more clinically relevant model of osteoporosis found that stem cells harvested from older, osteoporotic animals demonstrated limited proliferative and bone differentiation potential compared to healthy cells. However, 3D constructs (especially keratin-based materials) were able to enhance calcification and osteogenic gene expression of diseased cells. These results highlight the complexity of macrophage phenotypic switching and cellular dynamics in these systems. However, keratin-based therapeutics may prove useful for facilitating tissue regeneration and limiting detrimental inflammatory and cellular responses in various models of injury and disease.
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    Enduring deficits in memory and neuronal pathology after blast-induced traumatic brain injury
    Sajja, Venkata Siva Sai Sujith; Hubbard, W. Brad; Hall, Christina S.; Ghoddoussi, Farhad; Galloway, Matthew P.; VandeVord, Pamela J. (Nature Publishing Group, 2015-11-05)
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    Epigenetic Mechanisms in Blast-Induced Neurotrauma
    Bailey, Zachary S. (Virginia Tech, 2017-09-06)
    Blast-induced neurotrauma (BINT) is a prevalent brain injury within both military and civilian populations due to current engagement in overseas conflict and ongoing terrorist events worldwide. In the early 2000s, 78% of injuries were attributable to an explosive mechanism during overseas conflicts, which has led to increased incidences of BINT [1a]. Clinical manifestations of BINT include long-term psychological impairments, which are driven by the underlying cellular and molecular sequelae of the injury. Development of effective treatment strategies is limited by the lack of understanding on the cellular and molecular level [2a]. The overall hypothesis of this work is that epigenetic regulatory mechanisms contribute to the progression of the BINT pathology and neurological impairments. Epigenetic mechanisms, including DNA methylation and histone acetylation, are important processes by which cells coordinate neurological and cellular response to environmental stimuli. To date, the role of epigenetics in BINT remains largely unknown. To test this hypothesis, an established rodent model of BINT was employed [3a]. Analysis of DNA methylation, which is involved in memory processes, showed decreased levels one week following injury, which was accompanied by decreased expression of the enzyme responsible for facilitating the addition of methyl groups to DNA. The one week time point also showed dramatic decreases in histone acetylation which correlated to decline in memory. This change was observed in astrocytes and may provide a mechanistic understanding for a hallmark characteristic of the injury. Treatment with a specific enzyme inhibitor was able to mitigate some of the histone acetylation changes. This corresponded with reduced astrocyte activation and an altered behavioral phenotype, which was characterized by high response to novelty. The diagnostic efficacy of epigenetic changes following blast was elucidated by the accumulation of cell-free nucleic acids in cerebrospinal fluid one month after injury. Concentrations of these molecules shows promise in discriminating between injured and non-injured individuals. To date, the diagnostic and therapeutic efforts of BINT have been limited by the lack of a mechanistic understanding of the injury. This work provides novel diagnostic and therapeutic targets. The clinical potential impact on diagnosis and therapeutic intervention has been demonstrated.
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    Evaluating Microglia Dynamics in Blast and Impact-Induced Neurotrauma and Assessing the Role of Hemostatic Nanoparticles in Microglia Activation
    White, Michelle Renee (Virginia Tech, 2022-10-03)
    Traumatic brain injury (TBI) is a major medical concern that has demonstrated to be particularly challenging to treat because of the disparity amongst injury modes and severities. Increased use of explosive devices during combat has caused blast TBI (bTBI) to become a widespread consequence in military and Veteran populations, and impact-related trauma from contact-related sports or motor vehicle accidents has made mild impact-induced TBIs (concussion) a major health problem. There is a high risk for those who have sustained a TBI to develop behavioral and cognitive disorders following injury, and these symptoms can present as delayed onset, causing diagnosis to be a major feat when planning for treatment and long-term healthcare. Both preclinical and clinical studies report the neuropathological changes following TBI, yet investigating the distinct mechanistic changes in blast and impact trauma that contribute to pathological disparities has yet to be elucidated. Microglia dynamics play a key role in initiating the inflammatory response after injury, as microglia become activated by undergoing morphological changes that influence their function in the injured brain, and unique signaling pathways influence their functional inflammatory states. While previous literature report on the unique responses of microglia, their mediated-inflammatory responses are still not well defined. This work aimed to investigate the acute and subacute responses of microglia to injury through their diverse activation states following blast and impact trauma. The work herein employed rodent models to investigate these changes, finding that microglia activation was spatially and temporally heterogeneous within and across injury paradigms. Three days following bTBI, activated microglia in the cortex displayed morphologies similar to microglia that are known to increase their interactions with dysfunctional synapses, while dystrophic microglia were prevalent in the hippocampus seven days following injury. Moreover, transhemispheric changes in microglia activation were noted following impact TBI, with stressed/primed microglia responding to immune challenges of the cortex at three days, whereas a unique morphological state that was markedly different from those traditionally reported in CNS injury and disease was present within the hippocampus three- and seven-days following injury. State-of-the-art cell sorting techniques were used for in vivo analysis of microglia, which also exhibited that functional changes of microglia vary between injury paradigms, providing insight into how differences in primary insult may elicit distinct signaling pathways involved in microglia-mediated inflammatory responses. These in vivo studies were then crucial in understanding the malleable responses of microglia to complex injuries such as "blast plus impact" TBI, indicating that phenotypic changes in microglia following this injury are also unique and spatially heterogeneous. To date, therapeutic efforts for TBI are limited due to the lack of understanding the underlying mechanisms that influence TBI pathology. This work also investigated novel therapeutic targets, noting that administration of polyester nanoparticles restored microglia to baseline levels following impact. The fundamental research presented in this study is innovative and advantageous as it can provide essential data into targeted and personalized treatments that can improve long-term healthcare and ultimately, the quality of life for those suffering from a TBI.
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    Extracting Feature Vectors From Event-Related fMRI Data to Enable Machine Learning Analysis
    Soldate, Jeffrey S. (Virginia Tech, 2022-10-05)
    Linear models are the dominant means of extracting summaries of events in fMRI for feature vector based machine learning. While they are both useful and robust, they are limited by the assumptions made in modeling. In this work, we examine a number of feature extraction techniques adjacent to linear models that account for or allow wider variation. Primarily, we construct mixed effects models able to account for variation between stimuli of the same class and perform empirical tests on the resulting feature extraction – classifier system. We extend this analysis to spatial temporal models as well as summary models. We find that mixed effects models increase classifier performance at the cost of increased uncertainty in prediction estimates. In addition, these models identify similar regions of interest in separating classes. While they currently require knowledge hidden during testing, we present these results as an optimum to be reached in additional works.