Deletion of PTEN in microglia ameliorates chronic neuroinflammation following repetitive mTBI.

Traumatic brain injury is a leading cause of morbidity and mortality in adults and children in developed nations. Following the primary injury, microglia, the resident innate immune cells of the CNS, initiate several inflammatory signaling cascades and pathophysiological responses that may persist chronically; chronic neuroinflammation following TBI has been closely linked to the development of neurodegeneration and neurological dysfunction. Phosphoinositide 3-kinases (PI3Ks) are a family of lipid kinases that have been shown to regulate several key mechanisms in the inflammatory response to TBI. Increasing evidence has shown that the modulation of the PI3K/AKT signaling pathway has the potential to influence the cellular response to inflammatory stimuli. However, directly targeting PI3K signaling poses several challenges due to its regulatory role in several cell survival pathways. We have previously identified that the phosphatase and tensin homolog deleted on chromosome 10 (PTEN), the major negative regulator of PI3K/AKT signaling, is dysregulated following exposure to repetitive mild traumatic brain injury (r-mTBI). Moreover, this dysregulated PI3K/AKT signaling was correlated with chronic microglial-mediated neuroinflammation. Therefore, we interrogated microglial-specific PTEN as a therapeutic target in TBI by generating a microglial-specific, Tamoxifen inducible conditional PTEN knockout model using a CX3CR1 Cre recombinase mouse line PTENfl/fl/CX3CR1+/CreERT2 (mcg-PTENcKO), and exposed them to our 20-hit r-mTBI paradigm. Animals were treated with tamoxifen at 76 days post-last injury, and the effects of microglia PTEN deletion on immune-inflammatory responses were assessed at 90-days post last injury. We observed that the deletion of microglial PTEN ameliorated the proinflammatory response to repetitive brain trauma, not only reducing chronic microglial activation and proinflammatory cytokine production but also rescuing TBI-induced reactive astrogliosis, demonstrating that these effects extended beyond microglia alone. Additionally, we observed that the pharmacological inhibition of PTEN with BpV(HOpic) ameliorated the LPS-induced activation of microglial NFκB signaling in vitro. Together, these data provide support for the role of PTEN as a regulator of chronic neuroinflammation following repetitive mild TBI.

APOE genotype dependent molecular abnormalities in the cerebrovasculature of Alzheimer's disease and age-matched non-demented brains.

Cerebrovascular dysfunction is a hallmark feature of Alzheimer's disease (AD). One of the greatest risk factors for AD is the apolipoprotein E4 (E4) allele. The APOE4 genotype has been shown to negatively impact vascular amyloid clearance, however, its direct influence on the molecular integrity of the cerebrovasculature compared to other APOE variants (APOE2 and APOE3) has been largely unexplored. To address this, we employed a 10-plex tandem isobaric mass tag approach in combination with an ultra-high pressure liquid chromatography MS/MS (Q-Exactive) method, to interrogate unbiased proteomic changes in cerebrovessels from AD and healthy control brains with different APOE genotypes. We first interrogated changes between healthy control cases to identify underlying genotype specific effects in cerebrovessels. EIF2 signaling, regulation of eIF4 and 70S6K signaling and mTOR signaling were the top significantly altered pathways in E4/E4 compared to E3/E3 cases. Oxidative phosphorylation, EIF2 signaling and mitochondrial dysfunction were the top significant pathways in E2E2 vs E3/E3cases. We also identified AD-dependent changes and their interactions with APOE genotype and found the highest number of significant proteins from this interaction was observed in the E3/E4 (192) and E4/E4 (189) cases. As above, EIF2, mTOR signaling and eIF4 and 70S6K signaling were the top three significantly altered pathways in E4 allele carriers (i.e. E3/E4 and E4/E4 genotypes). Of all the cerebrovascular cell-type specific markers identified in our proteomic analyses, endothelial cell, astrocyte, and smooth muscle cell specific protein markers were significantly altered in E3/E4 cases, while endothelial cells and astrocyte specific protein markers were altered in E4/E4 cases. These proteomic changes provide novel insights into the longstanding link between APOE4 and cerebrovascular dysfunction, implicating a role for impaired autophagy, ER stress, and mitochondrial bioenergetics. These APOE4 dependent changes we identified could provide novel cerebrovascular targets for developing disease modifying strategies to mitigate the effects of APOE4 genotype on AD pathogenesis.

Molecular Pathobiology of the Cerebrovasculature in Aging and in Alzheimers Disease Cases With Cerebral Amyloid Angiopathy.

Cerebrovascular dysfunction and cerebral amyloid angiopathy (CAA) are hallmark features of Alzheimer's disease (AD). Molecular damage to cerebrovessels in AD may result in alterations in vascular clearance mechanisms leading to amyloid deposition around blood vessels and diminished neurovascular-coupling. The sequelae of molecular events leading to these early pathogenic changes remains elusive. To address this, we conducted a comprehensive in-depth molecular characterization of the proteomic changes in enriched cerebrovessel fractions isolated from the inferior frontal gyrus of autopsy AD cases with low (85.5 ± 2.9 yrs) vs. high (81 ± 4.4 yrs) CAA score, aged-matched control (87.4 ± 1.5 yrs) and young healthy control (47 ± 3.3 yrs) cases. We employed a 10-plex tandem isobaric mass tag approach in combination with our ultra-high pressure liquid chromatography MS/MS (Q-Exactive) method. Enriched cerebrovascular fractions showed very high expression levels of proteins specific to endothelial cells, mural cells (pericytes and smooth muscle cells), and astrocytes. We observed 150 significantly regulated proteins in young vs. aged control cerebrovessels. The top pathways significantly modulated with aging included chemokine, reelin, HIF1α and synaptogenesis signaling pathways. There were 213 proteins significantly regulated in aged-matched control vs. high CAA cerebrovessels. The top three pathways significantly altered from this comparison were oxidative phosphorylation, Sirtuin signaling pathway and TCA cycle II. Comparison between low vs. high CAA cerebrovessels identified 84 significantly regulated proteins. Top three pathways significantly altered between low vs. high CAA cerebrovessels included TCA Cycle II, Oxidative phosphorylation and mitochondrial dysfunction. Notably, high CAA cases included more advanced AD pathology thus cerebrovascular effects may be driven by the severity of amyloid and Tangle pathology. These descriptive proteomic changes provide novel insights to explain the age-related and AD-related cerebrovascular changes contributing to AD pathogenesis. Particularly, disturbances in energy bioenergetics and mitochondrial biology rank among the top AD pathways altered in cerebrovessels. Targeting these failed mechanisms in endothelia and mural cells may provide novel disease modifying targets for developing therapeutic strategies against cerebrovascular deterioration and promoting cerebral perfusion in AD. Our future work will focus on interrogating and validating these novel targets and pathways and their functional significance.

Repeated unpredictable stress and social isolation induce chronic HPA axis dysfunction and persistent abnormal fear memory.

The lack of progress in the psychopharmacological treatment of stress-related disorders such as PTSD is an ongoing crisis due to its negative socioeconomic implications. Current PTSD pharmacotherapy relies on a few FDA approved medications used primarily for depression which offer only symptomatic relief and show limited efficacy. As the population of PTSD patients is growing, the identification of effective etiology-based treatments for the condition is a high priority. This requires an in-depth understanding of the neurobiological and behavioral outcomes of stress in translationally relevant animal models. In this study, we use neuroendocrine, biochemical and behavioral measures to assess the HPA axis function and fear-memory deficits in a mouse model of chronic stress. The chronic stress procedures involved exposure to 21 days of repeated unpredictable stress (RUS), including predator stress, restraint and foot shock, followed by chronic social isolation. We show that mice exposed to our stress paradigm demonstrate exaggerated fear memory recall and blunted HPA axis functionality at one month after RUS. Our neuroendocrinal testing suggests that the attenuated stress response in our model may be related to an alteration in the adrenal MC2 receptor reactivity. While there was no noticeable change in pituitary negative feedback regulation mechanisms, CRH and phosphorylated Glucocorticoid receptors levels were altered in the hypothalamus. We also show that chronic supplementation with a peripheral glucocorticoid receptor agonist (low-dose dexamethasone) after RUS partially restores a number of stress-related behavioral deficits in the RUS model. This suggests a direct relationship between HPA axis function and behavior in our model. Our findings emphasize the importance of the adrenal receptors as a target for HPA axis dysfunction in stress and fear-related disorders.

Molecular abnormalities in autopsied brain tissue from the inferior horn of the lateral ventricles of nonagenarians and Alzheimer disease patients.

BACKGROUND: The ventricular system plays a vital role in blood-cerebrospinal fluid (CSF) exchange and interstitial fluid-CSF drainage pathways. CSF is formed in the specialized secretory tissue called the choroid plexus, which consists of epithelial cells, fenestrated capillaries and the highly vascularized stroma. Very little is currently known about the role played by the ventricles and the choroid plexus tissue in aging and Alzheimer's disease (AD). METHODS: In this study, we used our state-of-the-art proteomic platform, a liquid chromatography/mass spectrometry (LC-MS/MS) approach coupled with Tandem Mass Tag isobaric labeling to conduct a detailed unbiased proteomic analyses of autopsied tissue isolated from the walls of the inferior horn of the lateral ventricles in AD (77.2 ± 0.6 yrs), age-matched controls (77.0 ± 0.5 yrs), and nonagenarian cases (93.2 ± 1.1 yrs). RESULTS: Ingenuity pathway analyses identified phagosome maturation, impaired tight-junction signaling, and glucose/mannose metabolism as top significantly regulated pathways in controls vs nonagenarians. In matched-control vs AD cases we identified alterations in mitochondrial bioenergetics, oxidative stress, remodeling of epithelia adherens junction, macrophage recruitment and phagocytosis, and cytoskeletal dynamics. Nonagenarian vs AD cases demonstrated augmentation of oxidative stress, changes in gluconeogenesis-glycolysis pathways, and cellular effects of choroidal smooth muscle cell vasodilation. Amyloid plaque score uniquely correlated with remodeling of epithelial adherens junctions, Fc γ-receptor mediated phagocytosis, and alterations in RhoA signaling. Braak staging was uniquely correlated with altered iron homeostasis, superoxide radical degradation and phagosome maturation. CONCLUSIONS: These changes provide novel insights to explain the compromise to the physiological properties and function of the ventricles/choroid plexus system in nonagenarian aging and AD pathogenesis. The pathways identified could provide new targets for therapeutic strategies to mitigate the divergent path towards AD.

Impact of Repetitive Mild Traumatic Brain Injury on Behavioral and Hippocampal Deficits in a Mouse Model of Chronic Stress.

Clinical studies examining the interaction between traumatic brain injury (TBI) and stress-related disorders (e.g., post-traumatic stress disorder) are often complicated by methodological constraints, such as heterogeneity in injury type and severity, time post-trauma, and predisposing risk factors. Developing relevant animal models whereby many variables can be efficiently controlled is thus essential to understanding this elusive relationship. Here, we use our repeated unpredictable stress (RUS) paradigm, in combination with our established mouse model of repetitive mild TBI (r-mTBI), to assess the impact of repeated exposures to these paradigms on behavioral and neurobiological measures. C57BL/6J male mice were exposed to RUS and r-mTBI at 3 and 6 months of age followed by batteries of behavioral testing. Mice were euthanized 10 days and 3 months post-exposure, with brain and plasma samples collected for molecular profiling. The RUS paradigm involved exposure to a predator odor (trimethylthiazoline; TMT) while under restraint, daily unstable social housing, five inescapable footshocks on separate days, and chronic social isolation. Animals receiving r-mTBI ( × 5) and stress were exposed to a single closed-head injury 1 h after each footshock. Stress-alone mice showed significant weight loss, recall of traumatic memories, and anxiety-like and passive stress-coping behavior when compared with control mice. However, in stress+r-mTBI animals, the changes in cued fear memory, anxiety, and stress-coping tests were diminished, possibly due to TBI-induced hyperactivity. We also report complex brain molecular and neuropathological findings. Stress and r-mTBI, either individually or comorbidly, were associated with a chronic reduction in dendritic spine GluN2A/GluN2B ratio in the hippocampus. While stress augmented the r-mTBI-dependent astrogliosis in the corpus callosum, it mitigated r-mTBI-induced increases in hippocampal pro-brain-derived neurotrophic factor. We anticipate that our model will be a good platform to untangle the complex comorbid pathophysiology in stress disorders and r-mTBI.

Converging and Differential Brain Phospholipid Dysregulation in the Pathogenesis of Repetitive Mild Traumatic Brain Injury and Alzheimer's Disease.

Repetitive mild traumatic brain injury (rmTBI) is a major epigenetic risk factor for Alzheimer's disease (AD). The precise nature of how rmTBI leads to or precipitates AD pathology is currently unknown. Numerous neurological conditions have shown an important role for dysfunctional phospholipid metabolism as a driving factor for the pathogenesis of neurodegenerative diseases. However, the precise role in rmTBI and AD remains elusive. We hypothesized that a detailed phospholipid characterization would reveal profiles of response to injury in TBI that overlap with age-dependent changes in AD and thus provide insights into the TBI-AD relationship. We employed a lipidomic approach examining brain phospholipid profiles from mouse models of rmTBI and AD. Cortex and hippocampal tissue were collected at 24 h, 3, 6, 9, and 12 months post-rmTBI, and at ages representing 'pre', 'peri' and 'post' onset of amyloid pathology (i.e., 3, 9, 15 months-old). Total levels of phosphatidylcholine (PC), phosphatidylethanolamine (PE), LysoPE, and phosphatidylinositol (PI), including their monounsaturated, polyunsaturated and saturated fatty acid (FA) containing species were significantly increased at acute and/or chronic time points post-injury in both brain regions. However, levels of most phospholipid species in PS1/APP mice were nominal in the hippocampus, while in the cortex, levels were significantly decreased at ages post-onset of amyloid pathology. Sphingomyelin and LysoPC levels showed coincidental trends in our rmTBI and AD models within the hippocampus, an increase at acute and/or chronic time points examined. The ratio of arachidonic acid (omega-6 FA) to docosahexaenoic acid (omega-3 FA)-containing PE species was increased at early time points in the hippocampus of injured versus sham mice, and in PS1/APP mice there was a coincidental increase compared to wild type littermates at all time points. This study demonstrates some overlapping and diverse phospholipid profiles in rmTBI and AD models. Future studies are required to corroborate our findings in human post-mortem tissue. Investigation of secondary mechanisms triggered by aberrant downstream alterations in bioactive metabolites of these phospholipids, and their modulation at the appropriate time-windows of opportunity could help facilitate development of novel therapeutic strategies to ameliorate the neurodegenerative consequences of rmTBI or the potential triggering of AD pathogenesis by rmTBI.

APOE Genotype Specific Effects on the Early Neurodegenerative Sequelae Following Chronic Repeated Mild Traumatic Brain Injury.

Repeated mild traumatic brain injury (r-mTBI) can potentially manifest into chronic traumatic encephalopathy (CTE). The apolipoprotein E (APOE4) genotype, a well-recognized potent genetic risk factor in age-related neurodegenerative diseases such as Alzheimer's disease, has been linked to worse outcome after TBI in individuals who carry this allele. The underlying molecular modifications triggered by APOE genotype following r-mTBI remain elusive. We addressed the influence of APOE genotype on TBI dependent tau pathology in middle-aged mice. Using a previously established experimental mTBI protocol in a new repetitive injury paradigm, we report the pathological changes that occurred following one-month of repetitive injuries in APOE3/4 gene targeted mice. Firstly, pathological assessment demonstrated evidence of microgliosis and astrogliosis in the corpus callosum of injured animals, but there was no APOE dependent genotype effect on injury. However, in the parietal cortex Iba1-immunoreactivity was significantly increased in injured versus sham APOE3 mice, but not in APOE4 mice. No effects were observed in soluble amyloid levels with injury or interaction with genotype. APOE4 mice showed significant increases in the tau conformational marker MC1, neurofilament H, brain phospholipids, and endothelial specific oxidized low density lipoprotein receptor in cortical homogenates obtained from injured mice compared to sham counterparts. This pilot work suggests APOE3 and APOE4 specific effects following injury in a mouse model of r-mTBI. These changes may underlie the molecular changes that trigger the vulnerability and increased risk of developing neurodegenerative diseases in aged individuals exposed to repetitive mTBI.

Disruption in Brain Phospholipid Content in a Humanized Tau Transgenic Model Following Repetitive Mild Traumatic Brain Injury.

Repetitive mild traumatic brain injury (mTBI) is a risk factor for the development of neurodegenerative diseases such as chronic traumatic encephalopathy typified by immunoreactive tau aggregates in the depths of the sulci. However, the underlying neurobiological mechanisms involved have not been largely explored. Phospholipids are important molecules which form membrane lipid bilayers; they are ubiquitous to every cell in the brain, and carry out a host of different functions. Imbalance in phospholipid metabolism, signaling and transport has been documented in some neurological conditions. However, not much is currently known about their roles in repetitive mTBI and how this may confer risk for the development of age-related neurodegenerative diseases. To address this question, we designed a longitudinal study (24 h, 3, 6, 9, and 12 months post-injury) to comprehensively investigate mTBI dependent brain phospholipid profiles compared to sham counterparts. We use our established mouse model of repetitive mTBI that has been extensively characterized up to 1-year post-injury in humanized tau (hTau) mice, which expresses all six human tau isoforms, on a null murine background. Our data indicates a significant increase in sphingomyelin, phosphatidylethanolamine (PE), phosphatidylcholine (PC), and derivative lysoPE and lysoPC at acute and/or sub-acute time points post-injury within the cortex and hippocampus. There was also a parallel increase at early time points in monounsaturated, polyunsaturated and saturated fatty acids. Omega-6 (arachidonic acid) to omega-3 (docosahexaenoic acid) fatty acid ratio for PE and PC species was increased also at 24 h and 3 months post-injury in both hippocampus and cortex. The long-term consequences of these early changes in phospholipids on neuronal and non-neuronal cell function is unclear, and warrants further study. Understanding phospholipid metabolism, signaling and transport following TBI could be valuable; they may offer novel targets for therapeutic intervention not only in TBI but other neurodegenerative diseases.

Subchronic Pathobiological Response Following Chronic Repetitive Mild Traumatic Brain Injury in an Aged Preclinical Model of Amyloid Pathogenesis.

Repetitive mild traumatic brain injury (r-mTBI) is a risk factor for Alzheimer disease (AD). The precise nature of how r-mTBI leads to, or precipitates, AD pathogenesis remains unclear. In this study, we explore subchronic effects of chronic r-mTBI (12-impacts) administered over 1-month in aged-PS1/APP mice and littermate controls. We investigate specific mechanisms that may elucidate the molecular link between AD and r-mTBI, focusing primarily on amyloid and tau pathology, amyloid processing, glial activation states, and associated clearance mechanisms. Herein, we demonstrate r-mTBI in aged PS1/APP mice does not augment, glial activation, amyloid burden, or tau pathology (with exception of pS202-positive Tau) 1 month after exposure to the last-injury. However, we observed a decrease in brain soluble Aß42 levels without any appreciable change in peripheral soluble Aß42 levels. This was accompanied by an increase in brain insoluble to soluble Aß42 ratio in injured PS1/APP mice compared with sham injury. A parallel reduction in phagocytic receptor, triggering receptor expressed on myeloid cells 2, was also observed. This study demonstrates very subtle subchronic effects of r-mTBI on a preexisting amyloid pathology background, which may be on a continuum toward a slow and worsening neurodegenerative outcome compared with sham injury, and therefore, have many implications, especially in the elderly population exposed to TBI.

Chronic Hippocampal Abnormalities and Blunted HPA Axis in an Animal Model of Repeated Unpredictable Stress.

Incidence of post-traumatic stress disorder (PTSD) ranges from 3 to 30% in individuals exposed to traumatic events, with the highest prevalence in groups exposed to combat, torture, or rape. To date, only a few FDA approved drugs are available to treat PTSD, which only offer symptomatic relief and variable efficacy. There is, therefore, an urgent need to explore new concepts regarding the biological responses causing PTSD. Animal models are an appropriate platform for conducting such studies. Herein, we examined the chronic behavioral and neurobiological effects of repeated unpredictable stress (RUS) in a mouse model. 12 weeks-old C57BL/6J male mice were exposed to a 21-day RUS paradigm consisting of exposures to a predator odor (TMT) whilst under restraint, unstable social housing, inescapable footshocks and social isolation. Validity of the model was assessed by comprehensive examination of behavioral outcomes at an acute timepoint, 3 and 6 months post-RUS; and molecular profiling was also conducted on brain and plasma samples at the acute and 6 months timepoints. Stressed mice demonstrated recall of traumatic memories, passive stress coping behavior, acute anxiety, and weight gain deficits when compared to control mice. Immunoblotting of amygdala lysates showed a dysregulation in the p75NTR/ProBDNF, and glutamatergic signaling in stressed mice at the acute timepoint. At 6 months after RUS, stressed mice had lower plasma corticosterone, reduced hippocampal CA1 volume and reduced brain-derived neurotrophic factor levels. In addition, glucocorticoid regulatory protein FKBP5 was downregulated in the hypothalamus of stressed mice at the same timepoint, together implicating an impaired hypothalamus-pituitary-adrenal-axis. Our model demonstrates chronic behavioral and neurobiological outcomes consistent with those reported in human PTSD cases and thus presents a platform through which to understand the neurobiology of stress and explore new therapeutic interventions.

Lifelong behavioral and neuropathological consequences of repetitive mild traumatic brain injury.

Objective: Exposure to repetitive concussion, or mild traumatic brain injury (mTBI), has been linked with increased risk of long-term neurodegenerative changes, specifically chronic traumatic encephalopathy (CTE). To date, preclinical studies largely have focused on the immediate aftermath of mTBI, with no literature on the lifelong consequences of mTBI in these models. This study provides the first account of lifelong neurobehavioral and histological consequences of repetitive mTBI providing unique insight into the constellation of evolving and ongoing pathologies with late survival. Methods: Male C57BL/6J mice (aged 2-3 months) were exposed to either single or repetitive mild TBI or sham procedure. Thereafter, animals were monitored and assessed at 24 months post last injury for measures of motor coordination, learning deficits, cognitive function, and anxiety-like behavior prior to euthanasia and preparation of the brains for detailed neuropathological and protein biochemical studies. Results: At 24 months survival animals exposed to r-mTBI showed clear evidence of learning and working memory impairment with a lack of spatial memory and vestibule-motor vestibulomotor deficits compared to sham animals. Associated with these late behavioral deficits there was evidence of ongoing axonal degeneration and neuroinflammation in subcortical white matter tracts. Notably, these changes were also observed after a single mTBI, albeit to a lesser degree than repetitive mTBI. Interpretation: In this context, our current data demonstrate, for the first time, that rather than an acute, time limited event, mild TBI can precipitate a lifelong degenerative process. These data therefore suggest that successful treatment strategies should consider both the acute and chronic nature of mTBI.

Unbiased Proteomic Approach Identifies Unique and Coincidental Plasma Biomarkers in Repetitive mTBI and AD Pathogenesis.

The relationship between repetitive mild traumatic brain injury (r-mTBI) and Alzheimer's disease (AD) is well-recognized. However, the precise nature of how r-mTBI leads to or precipitates AD pathogenesis is currently not understood. Plasma biomarkers potentially provide non-invasive tools for detecting neurological changes in the brain, and can reveal overlaps between long-term consequences of r-mTBI and AD. In this study we address this by generating time-dependent molecular profiles of response to r-mTBI and AD pathogenesis in mouse models using unbiased proteomic analyses. To model AD, we used the well-validated hTau and PSAPP(APP/PS1) mouse models that develop age-related tau and amyloid pathological features, respectively, and our well-established model of r-mTBI in C57BL/6 mice. Plasma were collected at different ages (3, 9, and 15 months-old for hTau and PSAPP mice), encompassing pre-, peri- and post-"onset" of the cognitive and neuropathological phenotypes, or at different timepoints after r-mTBI (24 h, 3, 6, 9, and 12 months post-injury). Liquid chromatography/mass spectrometry (LC-MS) approaches coupled with Tandem Mass Tag labeling technology were applied to develop molecular profiles of protein species that were significantly differentially expressed as a consequence of mTBI or AD. Mixed model ANOVA after Benjamini-Hochberg correction, and a stringent cut-off identified 31 proteins significantly changing in r-mTBI groups over time and, when compared with changes over time in sham mice, 13 of these were unique to the injured mice. The canonical pathways predicted to be modulated by these changes were LXR/RXR activation, production of nitric oxide and reactive oxygen species and complement systems. We identified 18 proteins significantly changing in PSAPP mice and 19 proteins in hTau mice compared to their wild-type littermates with aging. Six proteins were found to be significantly regulated in all three models, i.e., r-mTBI, hTau, and PSAPP mice compared to their controls. The top canonical pathways coincidently changing in all three models were LXR/RXR activation, and production of nitric oxide and reactive oxygen species. This work suggests potential biomarkers for TBI and AD pathogenesis and for the overlap between these two, and warrant targeted investigation in human populations. Data are available via ProteomeXchange with identifier PXD010664.

Chronic Repetitive Mild Traumatic Brain Injury Results in Reduced Cerebral Blood Flow, Axonal Injury, Gliosis, and Increased T-Tau and Tau Oligomers.

Exposure to repetitive mild traumatic brain injury (mTBI) is a risk factor for chronic traumatic encephalopathy, which is characterized by patchy deposition of hyperphosphorylated tau aggregates in neurons and astrocytes at the depths of cortical sulci. We developed an mTBI paradigm to explore effects of repetitive concussive-type injury over several months in mice with a human tau genetic background (hTau). Two injuries were induced in the hTau mice weekly over a period of 3 or 4 months and the effects were compared with those in noninjured sham animals. Behavioral and in vivo measures and detailed neuropathological assessments were conducted 6 months after the first injury. Our data confirm impairment in cerebral blood flow and white matter damage. This was accompanied by a 2-fold increase in total tau levels and mild increases in tau oligomers/conformers and pTau (Thr231) species in brain gray matter. There was no evidence of neurofibrillary/astroglial tangles, neuropil threads, or perivascular foci of tau immunoreactivity. There were neurobehavioral deficits (ie, disinhibition and impaired cognitive performance) in the mTBI animals. These data support the relevance of this new mTBI injury model for studying the consequences of chronic repetitive mTBI in humans, and the role of tau in TBI.

Repetitive head trauma, chronic traumatic encephalopathy and tau: Challenges in translating from mice to men.

Chronic traumatic encephalopathy (CTE) is a neurological and psychiatric condition marked by preferential perivascular foci of neurofibrillary and glial tangles (composed of hyperphosphorylated-tau proteins) in the depths of the sulci. Recent retrospective case series published over the last decade on athletes and military personnel have added considerably to our clinical and histopathological knowledge of CTE. This has marked a vital turning point in the traumatic brain injury (TBI) field, raising public awareness of the potential long-term effects of mild and moderate repetitive TBI, which has been recognized as one of the major risk factors associated with CTE. Although these human studies have been informative, their retrospective design carries certain inherent limitations that should be cautiously interpreted. In particular, the current overriding issue in the CTE literature remains confusing in regard to appropriate definitions of terminology, variability in individual pathologies and the potential case selection bias in autopsy based studies. There are currently no epidemiological or prospective studies on CTE. Controlled preclinical studies in animals therefore provide an alternative means for specifically interrogating aspects of CTE pathogenesis. In this article, we review the current literature and discuss difficulties and challenges of developing in-vivo TBI experimental paradigms to explore the link between repetitive head trauma and tau-dependent changes. We provide our current opinion list of recommended features to consider for successfully modeling CTE in animals to better understand the pathobiology and develop therapeutics and diagnostics, and critical factors, which might influence outcome. We finally discuss the possible directions of future experimental research in the repetitive TBI/CTE field.

Ultrastructural Changes in the White and Gray Matter of Mice at Chronic Time Points After Repeated Concussive Head Injury.

Mild traumatic brain injury is a risk factor for neurodegenerative disease. We recently developed a model of repetitive concussive injury in mice that we have extensively characterized from 24 hours to 24 months after injury. Animals show evidence of progressive spatial memory deficits, thinning of the corpus callosum, axonal injury, and neuroglial activation. Here, we extended our neuropathologic characterization to the ultrastructural level in both a qualitative and a quantitative study. We focused on chronic (3 and 6 months) postinjury time points when the earliest stages of degenerative secondary changes were previously observed. In both C57BL/6 and hTau mice, we found white matter damage typified by axonal degeneration, microglial phagocytosis, and increased neuroglial cell density. In the cerebral cortex, we observed evidence of synaptic degeneration, dark neurons, altered dendritic microfilaments, subtle changes to the microvasculature, a mild augmentation of age-related features such as lipofuscin deposition, and electron-dense inclusions in microglial and perivascular cells. The majority of these ultrastructural features seemed to be more prominent at 3 versus 6 months after injury. Similar patterns were observed in C57BL/6 and hTau mice. These findings further support the relevance of our concussive injury model to the consequences of repetitive mild traumatic brain injury in humans.

Impact of age-related neuroglial cell responses on hippocampal deterioration.

Aging is one of the greatest risk factors for the development of sporadic age-related neurodegenerative diseases and neuroinflammation is a common feature of this disease phenotype. In the immunoprivileged brain, neuroglial cells, which mediate neuroinflammatory responses, are influenced by the physiological factors in the microenvironment of the central nervous system (CNS). These physiological factors include but are not limited to cell-to-cell communication involving cell adhesion molecules, neuronal electrical activity and neurotransmitter and neuromodulator action. However, despite this dynamic control of neuroglial activity, in the healthy aged brain there is an alteration in the underlying neuroinflammatory response notably seen in the hippocampus, typified by astrocyte/microglia activation and increased pro-inflammatory cytokine production and signaling. These changes may occur without any overt concurrent pathology, however, they typically correlate with deteriorations in hippocamapal or cognitive function. In this review we examine two important phenomenons, firstly the relationship between age-related brain deterioration (focusing on hippocampal function) and underlying neuroglial response(s), and secondly how the latter affects molecular and cellular processes within the hippocampus that makes it vulnerable to age-related cognitive decline.

Neurobehavioral, neuropathological and biochemical profiles in a novel mouse model of co-morbid post-traumatic stress disorder and mild traumatic brain injury.

Co-morbid mild traumatic brain injury (mTBI) and post-traumatic stress disorder (PTSD) has become the signature disorder for returning combat veterans. The clinical heterogeneity and overlapping symptomatology of mTBI and PTSD underscore the need to develop a preclinical model that will enable the characterization of unique and overlapping features and allow discrimination between both disorders. This study details the development and implementation of a novel experimental paradigm for PTSD and combined PTSD-mTBI. The PTSD paradigm involved exposure to a danger-related predator odor under repeated restraint over a 21 day period and a physical trauma (inescapable footshock). We administered this paradigm alone, or in combination with a previously established mTBI model. We report outcomes of behavioral, pathological and biochemical profiles at an acute timepoint. PTSD animals demonstrated recall of traumatic memories, anxiety and an impaired social behavior. In both mTBI and combination groups there was a pattern of disinhibitory like behavior. mTBI abrogated both contextual fear and impairments in social behavior seen in PTSD animals. No major impairment in spatial memory was observed in any group. Examination of neuroendocrine and neuroimmune responses in plasma revealed a trend toward increase in corticosterone in PTSD and combination groups, and an apparent increase in Th1 and Th17 proinflammatory cytokine(s) in the PTSD only and mTBI only groups respectively. In the brain there were no gross neuropathological changes in any groups. We observed that mTBI on a background of repeated trauma exposure resulted in an augmentation of axonal injury and inflammatory markers, neurofilament L and ICAM-1 respectively. Our observations thus far suggest that this novel stress-trauma-related paradigm may be a useful model for investigating further the overlapping and distinct spatio-temporal and behavioral/biochemical relationship between mTBI and PTSD experienced by combat veterans.