ABSTRACT
Traumatic brain injury (TBI), a significant global health issue, is affecting â¼69 million annually. To better understand TBI's impact on brain function and assess the efficacy of treatments, this study uses a novel temporal-spatial cross-group approach with a porcine model, integrating resting-state functional magnetic resonance imaging (rs-fMRI) for temporal and arterial spin labeling for spatial information. Our research used 18 four-week-old pigs divided into three groups: TBI treated with saline (SLN, n = 6), TBI treated with fecal microbial transplant (FMT, n = 6), and a sham group (sham, n = 6) with only craniectomy surgery as the baseline. By applying machine learning techniques-specifically, independent component analysis and sparse dictionary learning-across seven identified resting-state networks, we assessed the temporal and spatial correlations indicative of treatment efficacy. Both temporal and spatial analyses revealed a consistent increase of correlation between the FMT and sham groups in the executive control and salience networks. Our results are further evidenced by a simulation study designed to mimic the progression of TBI severity through the introduction of variable Gaussian noise to an independent rs-fMRI dataset. The results demonstrate a decreasing temporal correlation between the sham and TBI groups with increasing injury severity, consistent with the experimental results. This study underscores the effectiveness of the methodology in evaluating post-TBI treatments such as the FMT. By presenting comprehensive experimental and simulated data, our research contributes significantly to the field and opens new paths for future investigations into TBI treatment evaluations.
ABSTRACT
Pediatric traumatic brain injury (TBI) often induces significant disability in patients, including long-term motor deficits. Early detection of injury severity is key in determining a prognosis and creating appropriate intervention and rehabilitation plans. However, conventional magnetic resonance imaging (MRI) scans, such as T2 Weighted (T2W) sequences, do not reliably assess the extent of microstructural white matter injury. Diffusion tensor imaging (DTI) tractography enables three-dimensional reconstruction of specific white matter tracts throughout the brain in order to detect white matter injury based on anisotropic diffusion. The objective of this study was to employ DTI tractography to detect acute changes to white matter integrity within the intersecting fibers of key motor-related brain regions following TBI. Piglets were assigned to either the sham craniectomy group (sham; n = 6) or the controlled cortical impact TBI group (TBI; n = 6). Gait and MRI were collected at seven days post-surgery (DPS). T2W sequences confirmed a localized injury predominately in the ipsilateral hemisphere in TBI animals. TBI animals, relative to sham animals, showed an increased apparent diffusion coefficient (ADC) and decreased fractional anisotropy (FA) in fiber bundles associated with key brain regions involved in motor function. TBI animals exhibited gait deficits, including stride and step length, compared to sham animals. Together these data demonstrate acute reductions in the white matter integrity, measured by DTI tractography, of fibers intersecting key brain regions that strongly corresponded with acute motor deficits in a pediatric piglet TBI model. These results provide the foundation for the further development of DTI-based biomarkers to evaluate motor outcomes following TBI.
ABSTRACT
Pediatric traumatic brain injury (TBI) is a leading cause of death and disability in children. Due to bidirectional communication between the brain and gut microbial population, introduction of key gut bacteria may mitigate critical TBI-induced secondary injury cascades, thus lessening neural damage and improving functional outcomes. The objective of this study was to determine the efficacy of a daily fecal microbial transplant (FMT) to alleviate neural injury severity, prevent gut dysbiosis, and improve functional recovery post TBI in a translational pediatric piglet model. Male piglets at 4-weeks of age were randomly assigned to Sham + saline, TBI + saline, or TBI + FMT treatment groups. A moderate/severe TBI was induced by controlled cortical impact and Sham pigs underwent craniectomy surgery only. FMT or saline were administered by oral gavage daily for 7 days. MRI was performed 1 day (1D) and 7 days (7D) post TBI. Fecal and cecal samples were collected for 16S rRNA gene sequencing. Ipsilateral brain and ileum tissue samples were collected for histological assessment. Gait and behavior testing were conducted at multiple timepoints. MRI showed that FMT treated animals demonstrated decreased lesion volume and hemorrhage volume at 7D post TBI as compared to 1D post TBI. Histological analysis revealed improved neuron and oligodendrocyte survival and restored ileum tissue morphology at 7D post TBI in FMT treated animals. Microbiome analysis indicated decreased dysbiosis in FMT treated animals with an increase in multiple probiotic Lactobacilli species, associated with anti-inflammatory therapeutic effects, in the cecum of the FMT treated animals, while non-treated TBI animals showed an increase in pathogenic bacteria, associated with inflammation and disease such in feces. FMT mediated enhanced cellular and tissue recovery resulted in improved motor function including stride and step length and voluntary motor activity in FMT treated animals. Here we report for the first time in a highly translatable pediatric piglet TBI model, the potential of FMT treatment to significantly limit cellular and tissue damage leading to improved functional outcomes following a TBI.
ABSTRACT
Impaired gut homeostasis is associated with stroke often presenting with leaky gut syndrome and increased gut, brain, and systemic inflammation that further exacerbates brain damage. We previously reported that intracisternal administration of Tanshinone IIA-loaded nanoparticles (Tan IIA-NPs) and transplantation of induced pluripotent stem cell-derived neural stem cells (iNSCs) led to enhanced neuroprotective and regenerative activity and improved recovery in a pig stroke model. We hypothesized that Tan IIA-NP + iNSC combination therapy-mediated stroke recovery may also have an impact on gut inflammation and integrity in the stroke pigs. Ischemic stroke was induced, and male Yucatan pigs received PBS + PBS (Control, n = 6) or Tan IIA-NP + iNSC (Treatment, n = 6) treatment. The Tan IIA-NP + iNSC treatment reduced expression of jejunal TNF-α, TNF-α receptor1, and phosphorylated IkBα while increasing the expression of jejunal occludin, claudin1, and ZO-1 at 12 weeks post-treatment (PT). Treated pigs had higher fecal short-chain fatty acid (SCFAs) levels than their counterparts throughout the study period, and fecal SCFAs levels were negatively correlated with jejunal inflammation. Interestingly, fecal SCFAs levels were also negatively correlated with brain lesion volume and midline shift at 12 weeks PT. Collectively, the anti-inflammatory and neuroregenerative treatment resulted in increased SCFAs levels, tight junction protein expression, and decreased inflammation in the gut.
Subject(s)
Ischemic Stroke , Nanoparticles , Neural Stem Cells , Stroke , Male , Animals , Swine , Tumor Necrosis Factor-alpha , Stroke/therapy , Neural Stem Cells/pathology , Inflammation/pathology , Fatty Acids, VolatileABSTRACT
Induced pluripotent stem cell-derived neural stem cells (iNSCs) are a multimodal stroke therapeutic that possess neuroprotective, regenerative, and cell replacement capabilities post-ischemia. However, long-term engraftment and efficacy of iNSCs is limited by the cytotoxic microenvironment post-stroke. Tanshinone IIA (Tan IIA) is a therapeutic that demonstrates anti-inflammatory and antioxidative effects in rodent ischemic stroke models and stroke patients. Therefore, pretreatment with Tan IIA may create a microenvironment that is more conducive to the long-term survival of iNSCs. In this study, we evaluated the potential of Tan IIA drug-loaded nanoparticles (Tan IIA-NPs) to improve iNSC engraftment and efficacy, thus potentially leading to enhanced cellular, tissue, and functional recovery in a translational pig ischemic stroke model. Twenty-two pigs underwent middle cerebral artery occlusion (MCAO) and were randomly assigned to a PBSâ +â PBS, PBSâ +â iNSC, or Tan IIA-NPâ +â iNSC treatment group. Magnetic resonance imaging (MRI), modified Rankin Scale neurological evaluation, and immunohistochemistry were performed over a 12-week study period. Immunohistochemistry indicated pretreatment with Tan IIA-NPs increased iNSC survivability. Furthermore, Tan IIA-NPs increased iNSC neuronal differentiation and decreased iNSC reactive astrocyte differentiation. Tan IIA-NPâ +â iNSC treatment enhanced endogenous neuroprotective and regenerative activities by decreasing the intracerebral cellular immune response, preserving endogenous neurons, and increasing neuroblast formation. MRI assessments revealed Tan IIA-NPâ +â iNSC treatment reduced lesion volumes and midline shift. Tissue preservation and recovery corresponded with significant improvements in neurological recovery. This study demonstrated pretreatment with Tan IIA-NPs increased iNSC engraftment, enhanced cellular and tissue recovery, and improved neurological function in a translational pig stroke model.
Subject(s)
Abietanes , Ischemic Stroke , Nanoparticles , Neural Stem Cells , Animals , Ischemic Stroke/therapy , Swine , Abietanes/pharmacologyABSTRACT
Dynamic changes in the oral microbiome have gained attention due to their potential diagnostic role in neurological diseases such as Alzheimer's disease and Parkinson's disease. Traumatic brain injury (TBI) is a leading cause of death and disability in the United States, but no studies have examined the changes in oral microbiome during the acute stage of TBI using a clinically translational pig model. Crossbred piglets (4-5 weeks old, male) underwent either a controlled cortical impact (TBI, n = 6) or sham surgery (sham, n = 6). The oral microbiome parameters were quantified from the upper and lower gingiva, both buccal mucosa, and floor of the mouth pre-surgery and 1, 3, and 7 days post-surgery (PS) using the 16S rRNA gene. Faith's phylogenetic diversity was significantly lower in the TBI piglets at 7 days PS compared to those of sham, and beta diversity at 1, 3, and 7 days PS was significantly different between TBI and sham piglets. However, no significant changes in the taxonomic composition of the oral microbiome were observed following TBI compared to sham. Further studies are needed to investigate the potential diagnostic role of the oral microbiome during the chronic stage of TBI with a larger number of subjects.
ABSTRACT
BACKGROUND: The absolute number of new stroke patients is annually increasing and there still remains only a few Food and Drug Administration (FDA) approved treatments with significant limitations available to patients. Tanshinone IIA (Tan IIA) is a promising potential therapeutic for ischemic stroke that has shown success in pre-clinical rodent studies but lead to inconsistent efficacy results in human patients. The physical properties of Tan-IIA, including short half-life and low solubility, suggests that Poly (lactic-co-glycolic acid) (PLGA) nanoparticle-assisted delivery may lead to improve bioavailability and therapeutic efficacy. The objective of this study was to develop Tan IIA-loaded nanoparticles (Tan IIA-NPs) and to evaluate their therapeutic effects on cerebral pathological changes and consequent motor function deficits in a pig ischemic stroke model. RESULTS: Tan IIA-NP treated neural stem cells showed a reduction in SOD activity in in vitro assays demonstrating antioxidative effects. Ischemic stroke pigs treated with Tan IIA-NPs showed reduced hemispheric swelling when compared to vehicle only treated pigs (7.85 ± 1.41 vs. 16.83 ± 0.62%), consequent midline shift (MLS) (1.72 ± 0.07 vs. 2.91 ± 0.36 mm), and ischemic lesion volumes (9.54 ± 5.06 vs. 12.01 ± 0.17 cm3) when compared to vehicle-only treated pigs. Treatment also lead to lower reductions in diffusivity (-37.30 ± 3.67 vs. -46.33 ± 0.73%) and white matter integrity (-19.66 ± 5.58 vs. -30.11 ± 1.19%) as well as reduced hemorrhage (0.85 ± 0.15 vs 2.91 ± 0.84 cm3) 24 h post-ischemic stroke. In addition, Tan IIA-NPs led to a reduced percentage of circulating band neutrophils at 12 (7.75 ± 1.93 vs. 14.00 ± 1.73%) and 24 (4.25 ± 0.48 vs 5.75 ± 0.85%) hours post-stroke suggesting a mitigated inflammatory response. Moreover, spatiotemporal gait deficits including cadence, cycle time, step time, swing percent of cycle, stride length, and changes in relative mean pressure were less severe post-stroke in Tan IIA-NP treated pigs relative to control pigs. CONCLUSION: The findings of this proof of concept study strongly suggest that administration of Tan IIA-NPs in the acute phase post-stroke mitigates neural injury likely through limiting free radical formation, thus leading to less severe gait deficits in a translational pig ischemic stroke model. With stroke as one of the leading causes of functional disability in the United States, and gait deficits being a major component, these promising results suggest that acute Tan IIA-NP administration may improve functional outcomes and the quality of life of many future stroke patients.
ABSTRACT
Vitamin E is an essential antioxidant that may benefit athletes by reducing oxidative stress and influencing cytokine expression. Supplements can be derived from natural or manufactured synthetic sources. This study aimed to determine (1) if supplemental vitamin E is beneficial to exercising horses and (2) if there is a benefit of natural versus synthetic vitamin E. After 2 weeks on the control diet (vitamin E-deficient grain and hay), 18 horses were divided into three groups and fed the control diet plus (1) 1000 IU/d synthetic α-tocopherol (SYN-L), (2) 4000 IU/d synthetic α-tocopherol (SYN-H), or (3) 4000 IU/d RRR-α-tocopherol (natural source [NAT]). On day 7, horses began a 6-week training protocol, with standard exercise tests (SETs) performed before and after the 6-week protocol. Venous blood samples were collected on days 0, 7, 29, and 49. Horses fed NAT had higher α-tocopherol (P < .05) at post-SET1 through post-SET2. Plasma thiobarbituric acid-reactive substance levels were lower in NAT versus SYN-L horses after SET2 (P = .02). Serum aspartate aminotransferase was lower after exercise in NAT horses versus SYN-L and SYN-H (P = .02), and less reduction in stride duration was seen after exercise in NAT as compared with SYN-L and SYN-H (P = .02). Gene expression of tumor necrosis factor α was lower in NAT compared with SYN-H (P = .01) but not SYN-L. In conclusion, feeding higher levels of natural vitamin E source resulted in higher serum α-tocopherol levels as well as some improvement in oxidative and inflammatory response and improved functional outcomes in response to an exercise test.
Subject(s)
Animal Feed , Physical Conditioning, Animal , Animal Feed/analysis , Animals , Dietary Supplements , Horses , Oxidative Stress , Vitamin EABSTRACT
The acute stroke phase is a critical time frame used to evaluate stroke severity, therapeutic options, and prognosis while also serving as a major tool for the development of diagnostics. To further understand stroke pathophysiology and to enhance the development of treatments, our group developed a translational pig ischemic stroke model. In this study, the evolution of acute ischemic tissue damage, immune responses, and functional deficits were further characterized. Stroke was induced by middle cerebral artery occlusion in Landrace pigs. At 24 h post-stroke, magnetic resonance imaging revealed a decrease in ipsilateral diffusivity, an increase in hemispheric swelling resulting in notable midline shift, and intracerebral hemorrhage. Stroke negatively impacted white matter integrity with decreased fractional anisotropy values in the internal capsule. Like patients, pigs showed a reduction in circulating lymphocytes and a surge in neutrophils and band cells. Functional responses corresponded with structural changes through reductions in open field exploration and impairments in spatiotemporal gait parameters. Characterization of acute ischemic stroke in pigs provided important insights into tissue and functional-level assessments that could be used to identify potential biomarkers and improve preclinical testing of novel therapeutics.
Subject(s)
Ischemic Stroke/metabolism , Ischemic Stroke/physiopathology , White Matter/pathology , Animals , Brain Ischemia/pathology , Depression/etiology , Depression/physiopathology , Disease Models, Animal , Gait , Gait Analysis/methods , Infarction, Middle Cerebral Artery/pathology , Intracranial Hemorrhages/metabolism , Intracranial Hemorrhages/physiopathology , Lymphocytes/metabolism , Magnetic Resonance Imaging/methods , Male , Neutrophils/metabolism , Stroke/physiopathology , SwineABSTRACT
The Stroke Therapy Academic Industry Roundtable (STAIR) has recommended that novel therapeutics be tested in a large animal model with similar anatomy and physiology to humans. The pig is an attractive model due to similarities in brain size, organization, and composition relative to humans. However, multiple pig breeds have been used to study ischemic stroke with potentially differing cerebral anatomy, architecture and, consequently, ischemic stroke pathologies. The objective of this study was to characterize brain anatomy and assess spatiotemporal gait parameters in Yucatan (YC) and Landrace (LR) pigs pre- and post-stroke using magnetic resonance imaging (MRI) and gait analysis, respectively. Ischemic stroke was induced via permanent middle cerebral artery occlusion (MCAO). MRI was performed pre-stroke and 1-day post-stroke. Structural and diffusion-tensor sequences were performed at both timepoints and analyzed for cerebral characteristics, lesion diffusivity, and white matter changes. Spatiotemporal and relative pressure gait measurements were collected pre- and 2-days post-stroke to characterize and compare acute functional deficits. The results from this study demonstrated that YC and LR pigs exhibit differences in gross brain anatomy and gait patterns pre-stroke with MRI and gait analysis showing statistical differences in the majority of parameters. However, stroke pathologies in YC and LR pigs were highly comparable post-stroke for most evaluated MRI parameters, including lesion volume and diffusivity, hemisphere swelling, ventricle compression, caudal transtentorial and foramen magnum herniation, showing no statistical difference between the breeds. In addition, post-stroke changes in velocity, cycle time, swing percent, cadence, and mean hoof pressure showed no statistical difference between the breeds. These results indicate significant differences between pig breeds in brain size, anatomy, and motor function pre-stroke, yet both demonstrate comparable brain pathophysiology and motor outcomes post-stroke. The conclusions of this study suggest pigs of these different breeds generally show a similar ischemic stroke response and findings can be compared across porcine stroke studies that use different breeds.
ABSTRACT
Traumatic brain injury (TBI) is a leading cause of death and disability in children. Pediatric TBI patients often suffer from crippling cognitive, emotional, and motor function deficits that have negative lifelong effects. The objective of this study was to longitudinally assess TBI pathophysiology using multi-parametric magnetic resonance imaging (MRI), gait analysis, and histological approaches in a pediatric piglet model. TBI was produced by controlled cortical impact in Landrace piglets. MRI data, including from proton magnetic resonance spectroscopy (MRS), were collected 24 hours and 12 weeks post-TBI, and gait analysis was performed at multiple time-points over 12 weeks post-TBI. A subset of animals was sacrificed 24 hours, 1 week, 4 weeks, and 12 weeks post-TBI for histological analysis. MRI results demonstrated that TBI led to a significant brain lesion and midline shift as well as microscopic tissue damage with altered brain diffusivity, decreased white matter integrity, and reduced cerebral blood flow. MRS showed a range of neurochemical changes after TBI. Histological analysis revealed neuronal loss, astrogliosis/astrocytosis, and microglia activation. Further, gait analysis showed transient impairments in cadence, cycle time, % stance, step length, and stride length, as well as long-term impairments in weight distribution after TBI. Taken together, this study illustrates the distinct time course of TBI pathoanatomic and functional responses up to 12 weeks post-TBI in a piglet TBI model. The study of TBI injury and recovery mechanisms, as well as the testing of therapeutics in this translational model, are likely to be more predictive of human responses and clinical outcomes compared to traditional small animal models.
Subject(s)
Brain Injuries, Traumatic/diagnostic imaging , Brain/diagnostic imaging , Disease Models, Animal , Motor Skills Disorders/diagnostic imaging , Animals , Animals, Newborn , Brain/metabolism , Brain Injuries, Traumatic/metabolism , Male , Motor Skills Disorders/metabolism , SwineABSTRACT
Traumatic brain injury (TBI) is a leading cause of death and disability in the United States, with children who sustain a TBI having a greater risk of developing long-lasting cognitive, behavioral, and motor function deficits. This has led to increased interest in utilizing large animal models to study pathophysiologic and functional changes after injury in hopes of identifying novel therapeutic targets. In the present study, a controlled cortical impact (CCI) piglet TBI model was utilized to evaluate cognitive, motor, and histopathologic outcomes. CCI injury (4 m/sec velocity, 9 mm depression, 400 msec dwell time) was induced at the parietal cortex. Compared with normal pigs (n = 5), TBI pigs (n = 5) exhibited appreciable cognitive deficiencies, including significantly impaired spatial memory in spatial T-maze testing and a significant decrease in exploratory behaviors followed by marked hyperactivity in open field testing. Additionally, gait analysis revealed significant increases in cycle time and stance percent, significant decreases in hind reach, and a shift in the total pressure index from the front to the hind limb on the affected side, suggesting TBI impairs gait and balance. Pigs were sacrificed 28 days post-TBI and histological analysis revealed that TBI lead to a significant decrease in neurons and a significant increase in microglia activation and astrogliosis/astrocytosis at the perilesional area, a significant loss in neurons at the dorsal hippocampus, and significantly increased neuroblast proliferation at the subventricular zone. These data demonstrate a strong relationship between TBI-induced cellular changes and functional outcomes in our piglet TBI model that lay the framework for future studies that assess the ability of therapeutic interventions to contribute to functional improvements.