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 PBS + PBS, PBS + iNSC, or Tan IIA-NP + 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-NP + 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-NP + 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.

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.

Traumatic brain injury (TBI) is a leading cause of death and disability in pediatric patients and often results in delayed neural development and altered connectivity, leading to lifelong learning, memory, behavior, and motor function deficits. Induced pluripotent stem cell-derived neural stem cells (iNSCs) may serve as a novel multimodal therapeutic as iNSCs possess neuroprotective, regenerative, and cell-replacement capabilities post-TBI. In this study, we evaluated the effects of iNSC treatment on cellular, tissue, and functional recovery in a translational controlled cortical impact TBI piglet model. Five days post-craniectomy (n = 6) or TBI (n = 18), iNSCs (n = 7) or PBS (n = 11) were injected into perilesional brain tissue. Modified Rankin Scale (mRS) neurological evaluation, magnetic resonance imaging, and immunohistochemistry were performed over the 12-week study period. At 12-weeks post-transplantation, iNSCs showed long-term engraftment and differentiation into neurons, astrocytes, and oligodendrocytes. iNSC treatment enhanced endogenous neuroprotective and regenerative activities indicated by decreasing intracerebral immune responses, preserving endogenous neurons, and increasing neuroblast formation. These cellular changes corresponded with decreased hemispheric atrophy, midline shift, and lesion volume as well as the preservation of cerebral blood flow. iNSC treatment increased piglet survival and decreased mRS scores. The results of this study in a predictive pediatric large-animal pig model demonstrate that iNSC treatment is a robust multimodal therapeutic that has significant promise in potentially treating human pediatric TBI patients.

Aims: Alcohol is the most commonly abused substance leading to significant economic and medical burdens. Pigs are an attractive model for studying alcohol abuse disorder due to the comparable alcohol metabolism and consumption behavior, which are in stark contrast to rodent models. This study investigates the usage of a porcine model for voluntary binge drinking (BD) and a detailed analysis of gait changes due to motor function deficits during alcohol intoxication. Methods: Adolescent pigs were trained to drink increasing concentration (0–8%) of alcohol mixed in a 0.2% saccharin solution for 1 h in a two bottle choice test for 2 weeks. The training period was followed by a 3-week alcohol testing period, where animals were given free access to 8% alcohol in 0.2% saccharin solution and 0.2% saccharin water solution. Blood alcohol levels were tested and gait analysis was performed pre-alcohol consumption, last day of training, and Day 5 of each testing period. Results: Pigs voluntarily consumed alcohol to intoxication at all timepoints with blood alcohol concentration (BAL) ≥80 mg/dl. Spatiotemporal gait parameters including velocity, cadence, cycle time, swing time, stance time, step time, and stride length were perturbed as a result of intoxication. The stratification of the gait data based on BAL revealed that the gait parameters were affected in a dose-dependent manner. Conclusion: This novel adolescent BD porcine model with inherent anatomical and physiological similarities to humans display similar consumption and intoxication behavior that is likely to yield results that are translatable to human patients.

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.

Background/Objectives: Traumatic brain injury (TBI) is a global leading cause of disability and death, with millions of new cases added each year. Oxidative stress significantly exacerbates primary TBI, leading to increased levels of intracerebral cell death, tissue loss, and long-term functional deficits in surviving patients. Catalase and superoxide dismutase (SOD) mitigate oxidative stress and play a critical role in dampening injury severity. This study examines the neuroprotective effects of the novel antioxidant alpha lipoic acid-based therapeutic, CMX-2043, on antioxidant enzymes in a preclinical TBI model via various drug administration routes. Methods: Piglets (n = 28) underwent cortical controlled impact to induce moderate–severe TBI and were assigned to placebo (n = 10), subcutaneous CMX-2043 (SQ, 10 mg/kg; n = 9), or intravenous CMX-2043 (IV, 9 mg/kg; n = 9) treatment groups. Treatments began 1 h after TBI induction and continued for 5 days. MRI was performed throughout the study period to evaluate brain recovery. Blood was collected at 1, 7, and 42 days post-TBI, and liver and brain tissues were collected at 42 days post-TBI to measure catalase and SOD activity. Results: CMX-2043 IV-treated piglets showed 46.3% higher hepatic catalase activity than placebo (p = 0.0038), while the SQ group did not show significant changes in hepatic catalase activity compared to placebo. In the brain, SQ-treated piglets had significantly higher catalase activity than both IV (p = 0.0163) and placebo (p = 0.0003) groups (45.8340 ± 3.0855, 36.4822 ± 1.5558, 31.6524 ± 1.3129 nmol/min/mg protein for SQ, IV, and placebo, respectively), while IV-treated piglets did not show significant changes compared to placebo. IV-treated piglets did exhibit 39.3% higher brain SOD activity than placebo (p = 0.0148), while the SQ group did not show a significant change. CMX-2043 treatment did not alter plasma antioxidant enzyme activity during the study period. Importantly, within CMX-2043 treated TBI groups, piglets with significantly decreased lesion volumes, midline shift, and combined swelling and atrophy had better brain recovery, determined by MRI on day 1, 7, and 42 days post-injury TBI, exhibited higher brain catalase activity at 42 days post-injury TBI regardless of administration route, suggesting a link between improved recovery and sustained local catalase activity. Conclusions: This study highlights the impact of administration route on tissue-specific antioxidant responses, with IV administration enhancing liver catalase and brain SOD activity, while SQ administration primarily elevated brain catalase activity. In addition, this study shows an association between increased brain catalase activity and decreased TBI brain lesioning, midline shift, and combined swelling and atrophy, thus emphasizing the role of antioxidant defenses in neuroprotection post-injury.