Viral Clearance and Neuroinflammation in Acute TMEV Infection Vary by Host Genetic Background.

A wide range of viruses cause neurological manifestations in their hosts. Infection by neurotropic viruses as well as the resulting immune response can irreversibly disrupt the complex structural and functional architecture of the brain, depending in part on host genetic background. The interaction between host genetic background, neurological response to viral infection, and subsequent clinical manifestations remains poorly understood. In the present study, we used the genetically diverse Collaborative Cross (CC) mouse resource to better understand how differences in genetic background drive clinical signs and neuropathological manifestations of acute Theiler's murine encephalomyelitis virus (TMEV) infection. For the first time, we characterized variations of TMEV viral tropism and load based on host genetic background, and correlated viral load with microglial/macrophage activation. For five CC strains (CC002, CC023, CC027, CC057, and CC078) infected with TMEV, we compared clinical signs, lesion distribution, microglial/macrophage response, expression, and distribution of TMEV mRNA, and identified genetic loci relevant to the early acute (4 days post-infection [dpi]) and late acute (14 dpi) timepoints. We examined brain pathology to determine possible causes of strain-specific differences in clinical signs, and found that fields CA1 and CA2 of the hippocampal formation were especially targeted by TMEV across all strains. Using Iba-1 immunolabeling, we identified and characterized strain- and timepoint-specific variation in microglial/macrophage reactivity in the hippocampal formation. Because viral clearance can influence disease outcome, we used RNA in situ hybridization to quantify viral load and TMEV mRNA distribution at both timepoints. TMEV mRNA expression was broadly distributed in the hippocampal formation at 4 dpi in all strains but varied between radiating and clustered distribution depending on the CC strain. We found a positive correlation between microglial/macrophage reactivity and TMEV mRNA expression at 4 dpi. At 14 dpi, we observed a dramatic reduction in TMEV mRNA expression, and localization to the medial portion of field CA1 and field CA2. To better understand how host genetic background can influence pathological outcomes, we identified quantitative trait loci associated with frequency of lesions in a particular brain region and with microglial/macrophage reactivity. These QTL were located near several loci of interest: lysosomal trafficking regulator (Lyst) and nidogen 1 (Nid1), and transmembrane protein 106 B (Tmem106b). Together, these results provide a novel understanding about the influences of genetic variation on the acute neuropathological and immunopathological environment and viral load, which collectively lead to variable disease outcomes. Our findings reveal possible avenues for future investigation which may lead to more effective intervention strategies and treatment regimens.

Resilience in Long-Term Viral Infection: Genetic Determinants and Interactions.

Virus-induced neurological sequelae resulting from infection by Theiler's murine encephalomyelitis virus (TMEV) are used for studying human conditions ranging from epileptic seizures to demyelinating disease. Mouse strains are typically considered susceptible or resistant to TMEV infection based on viral persistence and extreme phenotypes, such as demyelination. We have identified a broader spectrum of phenotypic outcomes by infecting strains of the genetically diverse Collaborative Cross (CC) mouse resource. We evaluated the chronic-infection gene expression profiles of hippocampi and thoracic spinal cords for 19 CC strains in relation to phenotypic severity and TMEV persistence. Strains were clustered based on similar phenotypic profiles and TMEV levels at 90 days post-infection, and we categorized distinct TMEV response profiles. The three most common profiles included "resistant" and "susceptible," as before, as well as a "resilient" TMEV response group which experienced both TMEV persistence and mild neurological phenotypes even at 90 days post-infection. Each profile had a distinct gene expression signature, allowing the identification of pathways and networks specific to each TMEV response group. CC founder haplotypes for genes involved in these pathways/networks revealed candidate response-specific alleles. These alleles demonstrated pleiotropy and epigenetic (miRNA) regulation in long-term TMEV infection, with particular relevance for resilient mouse strains.

The radiographic analysis of a wounded patient questioned of having retained unexploded ordinance.

We present the case of a polytrauma patient brought into a military medical treatment facility in Khandahar, Afghanistan, whom on radiographic evaluation was identified to have multiple retained internal radiodensities, some of which resembled projectiles, not immediately obvious as to type, thus by default necessitating the consideration for retained unexploded ordinance. The ensuing analysis ultimately successfully identified the origin of retained radiodensities as inert fragments from a USSR 50/60 steel cored bullet as opposed to unexploded ordinance.

A case of bilateral ophthalmoplegia while diving.

This case report presents a military diver who became dysphoric and lost consciousness during a routine surface-supplied dive. The patient regained consciousness spontaneously, but the physical exam was notable for bilateral ophthalmoplegia. Full eye movement was regained during hyperbaric oxygen (HBO2) therapy, and the patient subsequently made a full recovery. Equipment and dive profile analysis led to the conclusion of hypercapnia and arterial gas embolism as the probable causes of the diver's symptoms. This is a unique case of isolated bilateral ophthalmoplegia presenting in a diving injury.

The efficacy of vitamin D supplementation during a prolonged submarine patrol.

Submariners spend prolonged periods submerged without sunlight exposure and may benefit from vitamin D supplementation to maintain vitamin D status. The primary objective of this study was to determine the efficacy of daily vitamin D supplementation on maintenance of 25-hydroxyvitamin D (25(OH)D) during a 3-month submarine patrol. Submariners were randomly divided into three groups: placebo (n = 16), 1,000 IU/day (n = 20), or 2,000 IU/day (n = 17). Anthropometrics, self-reported dietary calcium and vitamin D intake, serum markers of vitamin D and bone metabolism, and peripheral quantitative computed tomography (pQCT) parameters of the tibia were determined before and after the patrol. Prior to departure, 49 % of the subjects were vitamin D insufficient (<50 nmol/L). Following the patrol, 25(OH)D increased in all groups (p < 0.001): 3.3 ± 13.1 (placebo), 4.6 ± 11.3 (1,000 IU/day), and 13 ± 14 nmol/L (2,000 IU/day). The changes in 25(OH)D levels were dependent upon the baseline concentration of 25(OH)D and body mass (p < 0.001). Osteocalcin increased by 38 % (p < 0.01), and pQCT analyses revealed small, yet significant increases in indices of tibial structure and strength (p < 0.05) that were independent of supplementation. These data suggest that vitamin D status was low prior to the patrol, and the subsequent changes in vitamin D status were dependent on the baseline 25(OH)D levels and body mass. Furthermore, short-term skeletal health does not appear to be negatively affected by 3 months of submergence in spite of a suboptimal response to vitamin D supplementation.

Hormone and immune system interactions in demyelinating disease.

The immune, endocrine and nervous systems communicate with each other through a myriad of molecules including cytokines, hormones and neurotransmitters. Alterations in the balance of the products of these systems affect susceptibility to autoimmune disease and also the progression of disease. One of the most intensely studied autoimmune diseases is multiple sclerosis (MS). The purpose of this review is to explore the relationships between sex hormones and MS disease progression and to attempt to understand the paradox that although women are more likely to develop MS, female sex hormones appear to be beneficial in symptom amelioration. The proposed mechanisms of the therapeutic action of estrogens will be discussed with respect to T cell polarization and also on CNS cell populations. Investigations into the pathogenesis of multiple sclerosis (MS) and animal models of MS have given insights into the interactions between the neuroendocrine systems and provide important potential therapeutic venues that may be expanded to other autoimmune and neurodegenerative conditions.

An Enigmatic Case of Skeletal Fluorosis: Inhalant-Induced Toxicity.

ABSTRACT: Skeletal fluorosis is more common in the developing world, but is occasionally seen in the United States. We present radiographic, scintigraphic, CT, and clinical images of a 26-year-old woman with rapidly progressive, debilitating, polyostotic periostitis, and diffuse osteosclerosis typical of skeletal fluorosis. Laboratory analyses supported this diagnosis. The source of excess fluoride intake was elusive until a concurrent mental health workup revealed the patient's proclivity for inhaling air-duster cans containing difluoroethane. Difluoroethane inhalant abuse is an increasingly reported cause of skeletal fluorosis that astute clinicians should recognize. Discontinuation and sobriety from this toxic agent are essential for recovery.

C57BL/6J mice exposed to perfluorooctanoic acid demonstrate altered immune responses and increased seizures after Theiler's murine encephalomyelitis virus infection.

Introduction: Neurological diseases can stem from environmental influences such as antecedent viral infections or exposure to potential toxicants, some of which can trigger immune responses leading to neurological symptoms. Theiler's murine encephalomyelitis virus (TMEV) is used to model human neurological conditions associated with prior viral infections, with outcomes partly attributable to improper induction and regulation of the immune response. Perfluorooctanoic acid (PFOA) can alter pathologies known to influence neurological disease such as inflammatory responses, cytokine expression, and glial activation. Co-exposure to TMEV and PFOA was used to test the hypothesis that early life exposure to the potential immunotoxicant PFOA would affect immune responses so as to render TMEV-resistant C57BL/6J (B6) mice susceptible to viral-induced neurological disease. Methods: Neonate B6 mice were exposed to different treatments: non-injected, sham-infected with PBS, and TMEV-infected, with the drinking water of each group including either 70 ppt PFOA or filtered water. The effects of PFOA were evaluated by comparing neurological symptoms and changes in immune-related cytokine and chemokine production induced by viral infection. Immune responses of 23 cytokines and chemokines were measured before and after infection to determine the effects of PFOA exposure on immune response. Results: Prior to infection, an imbalance between Th1, Th2, and Treg cytokines was observed in PFOA-exposed mice, suppressing IL-4 and IL-13 production. However, the balance was restored and characterized by an increase in pro-inflammatory cytokines in the non-infected group, and a decrease in IL-10 in the PFOA + TMEV group. Furthermore, the PFOA + TMEV group experienced an increase in seizure frequency and severity. Discussion: Overall, these findings provide insight into the complex roles of immune responses in the pathogenesis of virus-associated neurological diseases influenced by co-exposures to viruses and immunotoxic compounds.

Serum Cytokines Predict Neurological Damage in Genetically Diverse Mouse Models.

Viral infections contribute to neurological and immunological dysfunction driven by complex genetic networks. Theiler's murine encephalomyelitis virus (TMEV) causes neurological dysfunction in mice and can model human outcomes to viral infections. Here, we used genetically distinct mice from five Collaborative Cross mouse strains and C57BL/6J to demonstrate how TMEV-induced immune responses in serum may predict neurological outcomes in acute infection. To test the hypothesis that serum cytokine levels can provide biomarkers for phenotypic outcomes of acute disease, we compared cytokine levels at pre-injection, 4 days post-injection (d.p.i.), and 14 d.p.i. Each strain produced unique baseline cytokine levels and had distinct immune responses to the injection procedure itself. Thus, we eliminated the baseline responses to the injection procedure itself and identified cytokines and chemokines induced specifically by TMEV infection. Then, we identified strain-specific longitudinal cytokine profiles in serum during acute disease. Using stepwise regression analysis, we identified serum immune markers predictive for TMEV-induced neurological phenotypes of the acute phase, e.g., IL-9 for limb paralysis; and TNF-α, IL-1ß, and MIP-1ß for limb weakness. These findings indicate how temporal differences in immune responses are influenced by host genetic background and demonstrate the potential of serum biomarkers to track the neurological effects of viral infection.

Circulating neutrophil activation in dogs with naturally occurring spinal cord injury secondary to intervertebral disk herniation.

OBJECTIVE: To investigate the time course of circulating neutrophil priming and activity in dogs with spinal cord injury secondary to intervertebral disk herniation that undergo decompressive surgery. ANIMALS: 9 dogs with spinal cord injury and 9 healthy dogs (controls). PROCEDURES: For dogs with spinal cord injury, blood samples were collected on the day of hospital admission and 3, 7, 30, and 90 days after injury and decompressive surgery. A single blood sample was collected from the control dogs. Flow cytometry analysis was performed on isolated neutrophils incubated with antibody against CD11b and nonfluorescent dihydrorhodamine 123, which was converted to fluorescent rhodamine 123 to measure oxidative burst activity. RESULTS: Expression of CD11b was increased in dogs with spinal cord injury 3 days after injury and decompressive surgery, relative to day 7 expression. Neutrophils expressed high oxidative burst activity both 3 and 7 days after injury and decompressive surgery, compared with activity in healthy dogs. CLINICAL RELEVANCE: For dogs with spinal cord injury, high CD11b expression 3 days after injury and decompressive surgery was consistent with findings for rodents with experimentally induced spinal cord injury. However, the high oxidative burst activity 3 and 7 days after injury and decompressive surgery was not consistent with data from other species, and additional studies on inflammatory events in dogs with naturally occurring spinal cord injury are needed.

Altered FDG Biodistribution in Subcutaneous White Fat on PET/CT Following l-Asparaginase Chemotherapy.

ABSTRACT: A 13-year-old boy with mediastinal T-cell lymphoblastic lymphoma demonstrated an altered biodistribution with diffuse activity in subcutaneous white adipose tissue and decreased visceral activity on interim posttreatment FDG PET/CT. This altered biodistribution was attributed to administration of the chemotherapeutic enzyme l-asparaginase 3 hours preceding the PET/CT, altering adipocytes amino acid and glucose metabolism. Treatment response assessment was adversely affected by the altered biodistribution, emphasizing the importance of maximizing the time between chemotherapy and PET/CT during successive oncologic treatment cycles. Because adipocytes protect leukemic cells in culture from l-asparaginase, we hypothesize that white adipose tissue-altered biodistribution may be related to l-asparaginase resistance.

Genetic and immunological contributors to virus-induced paralysis.

Infection by a single virus can evoke diverse immune responses, resulting in different neurological outcomes, depending on the host's genetic background. To study heterogenous viral response, we use Theiler's Murine Encephalomyelitis Virus (TMEV) to model virally induced neurological phenotypes and immune responses in Collaborative Cross (CC) mice. The CC resource consists of genetically distinct and reproducible mouse lines, thus providing a population model with genetic heterogeneity similar to humans. We examined different CC strains for the effect of chronic stage TMEV-induced immune responses on neurological outcomes throughout 90 days post infection (dpi), with a particular focus on limb paralysis, by measuring serum levels of 23 different cytokines and chemokines. Each CC strain demonstrated a unique set of immune responses, regardless of presence or absence of TMEV RNA. Using stepwise regression, significant associations were identified between IL-1α, RANTES, and paralysis frequency scores. To better understand these interactions, we evaluated multiple aspects of the different CC genetic backgrounds, including haplotypes of genomic regions previously linked with TMEV pathogenesis and viral clearance or persistence, individual cytokine levels, and TMEV-relevant gene expression. These results demonstrate how loci previously associated with TMEV outcomes provide incomplete information regarding TMEV-induced paralysis in the CC strains. Overall, these findings provide insight into the complex roles of immune response in the pathogenesis of virus-associated neurological diseases influenced by host genetic background.

Host genetic diversity drives variable central nervous system lesion distribution in chronic phase of Theiler's Murine Encephalomyelitis Virus (TMEV) infection.

Host genetic background is a significant driver of the variability in neurological responses to viral infection. Here, we leverage the genetically diverse Collaborative Cross (CC) mouse resource to better understand how chronic infection by Theiler's Murine Encephalomyelitis Virus (TMEV) elicits diverse clinical and morphologic changes in the central nervous system (CNS). We characterized the TMEV-induced clinical phenotype responses, and associated lesion distributions in the CNS, in six CC mouse strains over a 90 day infection period. We observed varying degrees of motor impairment in these strains, as measured by delayed righting reflex, paresis, paralysis, seizures, limb clasping, ruffling, and encephalitis phenotypes. All strains developed neuroparenchymal necrosis and mineralization in the brain, primarily localized to the hippocampal regions. Two of the six strains presented with axonal degeneration with myelin loss of the nerve roots in the lumbar spinal cord. Moreover, we statistically correlated lesion distribution with overall frequencies of clinical phenotypes and phenotype progression to better understand how and where TMEV targets the CNS, based on genetic background. Specifically, we assessed lesion distribution in relation to the clinical progression of these phenotypes from early to late TMEV disease, finding significant relationships between progression and lesion distribution. Finally, we identified quantitative trait loci associated with frequency of lesions in a particular brain region, revealing several loci of interest for future study: lysosomal trafficking regulator (Lyst) and nidogen 1 (Nid1). Together, these results indicate that the genetic background influences the type and severity of clinical phenotypes, phenotypic resilience to TMEV, and the lesion distribution across strains.

Restraint stress fails to render C57BL/6 mice susceptible to Theiler's virus-induced demyelination.

OBJECTIVES: Multiple sclerosis is a degenerative disease of the CNS with a pathology consistent with immunological mediation. Although its cause is unknown, multiple factors are thought to influence both the onset and exacerbation of the disease, including both genetic background as well as environmental factors. METHODS: We are interested in the effect of psychological stress on the onset and exacerbation of Theiler's virus-induced demyelinating disease (TVID), a murine model of MS in which viral persistence facilitates demyelination. In the current study, we determined whether chronic restraint stress (RS)-induced immunosuppression could result in the establishment of a persistent CNS infection in the normally TVID-resistant C57BL/6 mouse strain, resulting in demyelination. RESULTS: Our data indicated that RS repeated over the course of 7 days was not sufficient to cause decreases in virus-specific adaptive immunity, and did not significantly alter CNS viral levels. Furthermore, chronic repeated RS lasting until 4 weeks after infection altered neither the development of virus-specific IgG nor motor function determined by Rotarod analysis. In addition, histological analysis of the CNS of stressed mice indicated no inflammation or demyelination on day 193 after infection. CONCLUSION: These results suggest that stress alone is not sufficient to overcome genetic resistance to TVID in the C57BL/6 mouse strain.

Effects of stress on the immune response to Theiler's virus--implications for virus-induced autoimmunity.

Psychological stress is an important factor in susceptibility to many diseases. Our laboratory has been investigating the impact of stress on the susceptibility to Theiler's virus-induced demyelination (TVID), a mouse model of multiple sclerosis. Using immunodominant viral peptides specific for identification of either CD4(+) or CD8(+) T cells, stress reduced IFN-gamma-producing virus-specific CD4(+) and CD8(+) T cells in the spleen and CD8(+) T cells in the CNS. Expression of mRNA for the Th1 transcription factor T-bet and the Th2 transcription factor GATA-3 were decreased in spleen cells isolated from stressed mice. Cytokine production by cells isolated from the CNS or spleens following stimulation with virus indicated that stress decreased both type 1 and type 2 responses. The adverse effects of stress were partially reversed by concurrent RU486 administration but mimicked by dexamethasone, indicating a major role for glucocorticoids. Global stress-induced immunosuppression resulted in higher levels of virus replication and dissemination. The higher viral load subsequently led to an earlier disease onset and more severe clinical and histological signs of demyelinating disease. Our results have important implications for understanding the pathogenesis of MS, and suggest that stressful events during early infection with an agent capable of inducing demyelination may result in immunosuppression and failure to eliminate the pathogen, which in turn may lead to the development of MS.

Rapidly lethal secondary hemophagocytic lymphohistiocytosis predicted by fluorodeoxyglucose positron-emission tomography/computed tomography.

Secondary hemophagocytic lymphohistiocytosis (sHLH) is a rare disease with either an indolent or aggressive course. A 29-year-old male presented with fever, polyarthralgias, splenomegaly, retroperitoneal adenopathy, and laboratory findings consistent with Epstein-Barr-mediated sHLH. Consistent with a prior survival analysis by Kim et al., splenic maximum standardized uptake value (SUVmax) >2.52 and bone marrow SUVmax >3.13 on 18F- fuorodeoxyglucose positron emission tomography/computed tomography (18FDG-PET/CT) predicted an aggressive disease with poor treatment response. Despite optimal treatment, the patient rapidly progressed to death within 3 months of symptom onset. This case underscores the potential lethal nature of sHLH, and the evolving role of 18FDG-PET/CT in predicting disease severity and treatment response.

Reduced Axial Scan Length Coronary Calcium Scoring Reduces Radiation Dose and Provides Adequate Clinical Decision-Making Before Coronary CT Angiography.

Extensive coronary artery calcium (CAC) diminishes the accuracy of coronary computed tomography angiography (CCTA). Many imagers adjust CCTA acquisition parameters depending on a preCCTA Agatston CAC score to optimize diagnostic accuracy. Typical preCCTA CAC imaging adds considerably to radiation exposure, partially attributable to imaging beyond the area known for highest CAC, the proximal coronary arteries. We aimed to determine whether a z-axis reduced scan length (RSL) would identify the majority of CAC and provide adequate information to computed tomography angiography providers relative to a standard full-scan length (FSL) preCCTA noncontrast CT. We retrospectively examined 200 subjects. The mean CAC scores detected in RSL and FSL were 77.4 (95% CI 50.6 to 104.3) and 93.9 (95% CI 57.3 to 130.5), respectively. RSL detected 81% of the FSL CAC. Among false negatives, with no CAC detected in RSL, FSL CAC severity was minimal (mean score 2.8). There was high concordance, averaging 88%, between CCTA imaging parameter adjustment decisions made by 2 experienced imagers based on either RSL or FSL. CAC detected and decision concordance decreased with increasing CAC burden. CAC detected was lower, and false negatives were more common in the right coronary artery owing to its anatomic course, placing larger segments outside RSL. Axial scan length and effective dose decreased 59% from FSL (∼14.5 cm/∼1.1 mSv) to RSL (∼5.9 cm/∼0.45 mSv). This retrospective study suggests that RSL identifies most CAC, results in similar CCTA acquisition parameter modifications, and reduces radiation exposure. Our colleagues corroborated these results in a recently published prospective study.

Bilateral End-Organ Endophthalmitis in Setting of Serratia marcescens Urosepsis on 18F-FDG PET/CT.

Endogenous endophthalmitis is an uncommon infection of the internal tissues of the eye resulting from a bloodborne infection. We present the case of an 89-year-old man with Serratia marcescens urosepsis with multifocal end-organ infection on F-FDG PET/CT including aortitis, splenic abscess, septic arthritis, thoracolumbar discitis-osteomyelitis, and culture-confirmed bilateral endogenous endophthalmitis. Endophthalmitis-related intraconal ocular space uptake was clearly distinguishable from normal conal space extraocular muscle activity. Although sepsis responded to treatment, endophthalmitis led to complete vision loss. This case report presents PET/CT description of a rare and difficult-to-treat Serratia marcescens endogenous endophthalmitis, which usually has a poor visual outcome.

Antecedent presentation of neurological phenotypes in the Collaborative Cross reveals four classes with complex sex-dependencies.

Antecedent viral infection may contribute to increased susceptibility to several neurological diseases, such as multiple sclerosis and Parkinson's disease. Variation in clinical presentations of these diseases is often associated with gender, genetic background, or a combination of these and other factors. The complicated etiologies of these virally influenced diseases are difficult to study in conventional laboratory mouse models, which display a very limited number of phenotypes. We have used the genetically and phenotypically diverse Collaborative Cross mouse panel to examine complex neurological phenotypes after viral infection. Female and male mice from 18 CC strains were evaluated using a multifaceted phenotyping pipeline to define their unique disease profiles following infection with Theiler's Murine Encephalomyelitis Virus, a neurotropic virus. We identified 4 distinct disease progression profiles based on limb-specific paresis and paralysis, tremors and seizures, and other clinical signs, along with separate gait profiles. We found that mice of the same strain had more similar profiles compared to those of different strains, and also identified strains and phenotypic parameters in which sex played a significant role in profile differences. These results demonstrate the value of using CC mice for studying complex disease subtypes influenced by sex and genetic background. Our findings will be useful for developing novel mouse models of virally induced neurological diseases with heterogenous presentation, an important step for designing personalized, precise treatments.

Restraint stress modulates virus specific adaptive immunity during acute Theiler's virus infection.

Multiple sclerosis (MS) is a devastating CNS disease of unknown origin. Multiple factors including genetic background, infection, and psychological stress affect the onset or progression of MS. Theiler's murine encephalomyelitis virus (TMEV) infection is an animal model of MS in which aberrant immunity leads to viral persistence and subsequently results in demyelination that resembles MS. Here, we examined how stress during acute TMEV infection altered virus-specific cell mediated responses. Using immunodominant viral peptides specific for either CD4(+) or CD8(+) T cells, we found that stress reduced IFN-gamma producing virus-specific CD4(+) and CD8(+) T cells in the spleen and CD8(+) T cells CNS. Cytokine production by cells isolated from the CNS or spleens following stimulation with virus or viral peptides, indicated that stress decreased both type 1 and type 2 responses. Glucocorticoids were implicated in the decreased T cell function as the effects of stress were partially reversed by concurrent RU486 administration but mimicked by dexamethasone. As T cells mediate viral clearance in this model, our data support the hypothesis that stress-induced immunosuppression may provide a mechanism for enhanced viral persistence within the CNS.