Peak Disease Severity Research Articles

Ferritin loss in astrocytes reduces spinal cord oxidative stress and demyelination in the experimental autoimmune encephalomyelitis (EAE) model.

Demyelinating diseases such as multiple sclerosis (MS) cause myelin degradation and oligodendrocyte death, resulting in the release of toxic iron and iron-induced oxidative stress. Astrocytes have a large capacity for iron transport and storage, however the role of astrocytic iron homeostasis in demyelinating disorders is not completely understood. Here we investigate whether astrocytic iron metabolism modulates neuroinflammation, oligodendrocyte survival, and oxidative stress following demyelination. To this aim, we conditionally knock out ferritin in astrocytes and induce experimental autoimmune encephalomyelitis (EAE), an autoimmune-mediated model of demyelination. Ferritin ablation in astrocytes reduced the severity of disease in both the acute and chronic phases. The day of onset, peak disease severity, and cumulative clinical score were all significantly reduced in ferritin KO animals. This corresponded to better performance on the rotarod and increased mobility in ferritin KO mice. Furthermore, the spinal cord of ferritin KO mice display decreased numbers of reactive astrocytes, activated microglia, and infiltrating lymphocytes. Correspondingly, the size of demyelinated lesions, iron accumulation, and oxidative stress were attenuated in the CNS of ferritin KO subjects, particularly in white matter regions of the spinal cord. Thus, deleting ferritin in astrocytes reduced neuroinflammation, oxidative stress, and myelin deterioration in EAE animals. Collectively, these findings suggest that iron storage in astrocytes is a potential therapeutic target to lessen CNS inflammation and myelin loss in autoimmune demyelinating diseases.

Deletion of voltage-gated calcium channels in astrocytes decreases neuroinflammation and demyelination in a murine model of multiple sclerosis

The experimental autoimmune encephalomyelitis (EAE) model of multiple sclerosis was used in combination with a Cav1.2 conditional knock-out mouse (Cav1.2KO) to study the role of astrocytic voltage-gated Ca++ channels in autoimmune CNS inflammation and demyelination. Cav1.2 channels were specifically ablated in Glast-1-positive astrocytes by means of the Cre-lox system before EAE induction. After immunization, motor activity was assessed daily, and a clinical score was given based on the severity of EAE symptoms. Cav1.2 deletion in astrocytes significantly reduced the severity of the disease. While no changes were found in the day of onset and peak disease severity, EAE mean clinical score was lower in Cav1.2KO animals during the chronic phase of the disease. This corresponded to better performance on the rotarod and increased motor activity in Cav1.2KO mice. Furthermore, decreased numbers of reactive astrocytes, activated microglia, and infiltrating lymphocytes were found in the lumbar section of the spinal cord of Cav1.2KO mice 40 days after immunization. The degree of myelin protein loss and size of demyelinated lesions were also attenuated in Cav1.2KO spinal cords. Similar results were found in EAE animals treated with nimodipine, a Cav1.2 Ca++ channel inhibitor with high affinity to the CNS. Mice injected with nimodipine during the acute and chronic phases of the disease exhibited lower numbers of reactive astrocytes, activated microglial, and infiltrating immune cells, as well as fewer demyelinated lesions in the spinal cord. These changes were correlated with improved clinical scores and motor performance. In summary, these data suggest that antagonizing Cav1.2 channels in astrocytes during EAE alleviates neuroinflammation and protects the spinal cord from autoimmune demyelination.

SARS-CoV-2 antibody responses associate with sex, age and disease severity in previously uninfected people admitted to hospital with COVID-19: An ISARIC4C prospective study.

The SARS-CoV-2 pandemic enables the analysis of immune responses induced against a novel coronavirus infecting immunologically naïve individuals. This provides an opportunity for analysis of immune responses and associations with age, sex and disease severity. Here we measured an array of solid-phase binding antibody and viral neutralising Ab (nAb) responses in participants (n=337) of the ISARIC4C cohort and characterised their correlation with peak disease severity during acute infection and early convalescence. Overall, the responses in a Double Antigen Binding Assay (DABA) for antibody to the receptor binding domain (anti-RBD) correlated well with IgM as well as IgG responses against viral spike, S1 and nucleocapsid protein (NP) antigens. DABA reactivity also correlated with nAb. As we and others reported previously, there is greater risk of severe disease and death in older men, whilst the sex ratio was found to be equal within each severity grouping in younger people. In older males with severe disease (mean age 68 years), peak antibody levels were found to be delayed by one to two weeks compared with women, and nAb responses were delayed further. Additionally, we demonstrated that solid-phase binding antibody responses reached higher levels in males as measured via DABA and IgM binding against Spike, NP and S1 antigens. In contrast, this was not observed for nAb responses. When measuring SARS-CoV-2 RNA transcripts (as a surrogate for viral shedding) in nasal swabs at recruitment, we saw no significant differences by sex or disease severity status. However, we have shown higher antibody levels associated with low nasal viral RNA indicating a role of antibody responses in controlling viral replication and shedding in the upper airway. In this study, we have shown discernible differences in the humoral immune responses between males and females and these differences associate with age as well as with resultant disease severity.

COVID-19 Chest CT Quantification: Triage and Prognostic Value in Different Ages.

Objective: We evaluated the triage and prognostic performance of seven proposed computed tomography (CT)-severity score (CTSS) systems in two different age groups.Design: Retrospective study.Setting: COVID-19 pandemic.Participants: Admitted COVID-19, PCR-positive patients were included, excluding patients with heart failure and significant pre-existing pulmonary disease.Methods: Patients were divided into two age groups: ≥65 years and ≤64 years. Clinical data indicating disease severity at presentation and at peak disease severity were recorded. Initial CT images were scored by two radiologists according to seven CTSSs (CTSS1-CTSS7). Receiver operating characteristic (ROC) analysis for the performance of each CTSS in diagnosing severe/critical disease on admission (triage performance) and at peak disease severity (prognostic performance) was done for the whole cohort and each age group separately.Results: Included were 96 patients. Intraclass correlation coefficient (ICC) between the two radiologists scoring the CT scan images were good for all the CTSSs (ICC=0.764-0.837). In the whole cohort, all CTSSs showed an unsatisfactory area under the curve (AUC) in the ROC curve for triage, excluding CTSS2 (AUC=0.700), and all CTSSs showed acceptable AUCs for prognostic usage (0.759-0.781). In the older group (≥65 years; n=55), all CTSSs excluding CTSS6 showed excellent AUCs for triage (0.804-0.830), and CTSS6 was acceptable (AUC=0.796); all CTSSs showed excellent or outstanding AUCs for prognostication (0.859-0.919). In the younger group (≤64 years; n=41), all CTSSs showed unsatisfactory AUCs for triage (AUC=0.487-0.565) and prognostic usage (AUC=0.668-0.694), excluding CTSS6, showing marginally acceptable AUC for prognostic performance (0.700).Conclusion: Those CTSSs requiring more numerous segmentations, namely CTSS2, CTSS7, and CTSS5 showed the best ICCs; therefore, they are the best when comparison between two separate scores is needed. Irrespective of patients' age, CTSSs show minimal value in triage and acceptable prognostic value in COVID-19 patients. CTSS performance is highly variable in different age groups. It is excellent in those aged ≥65 years, but has little if any value in younger patients. Multicenter studies with larger sample size to evaluate results of this study should be conducted.

Distinct blood inflammatory biomarker clusters stratify host phenotypes during the middle phase of COVID-19

The associations between clinical phenotypes of coronavirus disease 2019 (COVID-19) and the host inflammatory response during the transition from peak illness to convalescence are not yet well understood. Blood plasma samples were collected from 129 adult SARS-CoV-2 positive inpatient and outpatient participants between April 2020 and January 2021, in a multi-center prospective cohort study at 8 military hospitals across the United States. Plasma inflammatory protein biomarkers were measured in samples from 15 to 28 days post symptom onset. Topological Data Analysis (TDA) was used to identify patterns of inflammation, and associations with peak severity (outpatient, hospitalized, ICU admission or death), Charlson Comorbidity Index (CCI), and body mass index (BMI) were evaluated using logistic regression. The study population (n = 129, 33.3% female, median 41.3 years of age) included 77 outpatient, 31 inpatient, 16 ICU-level, and 5 fatal cases. Three distinct inflammatory biomarker clusters were identified and were associated with significant differences in peak disease severity (p < 0.001), age (p < 0.001), BMI (p < 0.001), and CCI (p = 0.001). Host-biomarker profiles stratified a heterogeneous population of COVID-19 patients during the transition from peak illness to convalescence, and these distinct inflammatory patterns were associated with comorbid disease and severe illness due to COVID-19.

A 1-year longitudinal study on COVID-19 convalescents reveals persistence of anti-SARS-CoV-2 humoral and cellular immunity

ABSTRACT The immune memory of over 400 million COVID-19 convalescents is not completely understood. In this integrated study, we recorded the post-acute sequelae symptoms and tested the immune memories, including circulating antibodies, memory B cell, and memory CD4 or CD8 T cell responses of a cohort of 65 COVID-19 patients over 1-year after infection. Our data show that 48% of them still have one or more sequelae symptoms and all of them maintain at least one of the immune components. The chances of having sequelae symptoms or having better immune memory are associated with peak disease severity. We did four-time points sampling per subject to precisely understand the kinetics of durability of SARS-CoV-2 circulating antibodies. We found that the RBD IgG levels likely reach a stable plateau at around 6 months, albeit it is waning at the first 6 months after infection. At 1-year after infection, more than 90% of the convalescents generated memory CD4 or CD8 T memory responses, preferably against the SARS-CoV-2 M peptide pool. The convalescents also have polyfunctional and central memory T cells that could provide rapid and efficient response to SARS-CoV-2 re-infection. Based on this information, we assessed the immune protection against the Omicron variant and concluded that convalescents should still induce effective T cell immunity against the Omicron. By studying the circulating antibodies and memory B or T cell responses to SARS-CoV-2 in an integrated manner, our study provides insight into the understanding of protective immunity against diseases caused by secondary SARS-CoV-2 infection.

Characteristics, Outcomes, and Severity Risk Factors Associated With SARS-CoV-2 Infection Among Children in the US National COVID Cohort Collaborative

Understanding of SARS-CoV-2 infection in US children has been limited by the lack of large, multicenter studies with granular data. To examine the characteristics, changes over time, outcomes, and severity risk factors of children with SARS-CoV-2 within the National COVID Cohort Collaborative (N3C). A prospective cohort study of encounters with end dates before September 24, 2021, was conducted at 56 N3C facilities throughout the US. Participants included children younger than 19 years at initial SARS-CoV-2 testing. Case incidence and severity over time, demographic and comorbidity severity risk factors, vital sign and laboratory trajectories, clinical outcomes, and acute COVID-19 vs multisystem inflammatory syndrome in children (MIS-C), and Delta vs pre-Delta variant differences for children with SARS-CoV-2. A total of 1 068 410 children were tested for SARS-CoV-2 and 167 262 test results (15.6%) were positive (82 882 [49.6%] girls; median age, 11.9 [IQR, 6.0-16.1] years). Among the 10 245 children (6.1%) who were hospitalized, 1423 (13.9%) met the criteria for severe disease: mechanical ventilation (796 [7.8%]), vasopressor-inotropic support (868 [8.5%]), extracorporeal membrane oxygenation (42 [0.4%]), or death (131 [1.3%]). Male sex (odds ratio [OR], 1.37; 95% CI, 1.21-1.56), Black/African American race (OR, 1.25; 95% CI, 1.06-1.47), obesity (OR, 1.19; 95% CI, 1.01-1.41), and several pediatric complex chronic condition (PCCC) subcategories were associated with higher severity disease. Vital signs and many laboratory test values from the day of admission were predictive of peak disease severity. Variables associated with increased odds for MIS-C vs acute COVID-19 included male sex (OR, 1.59; 95% CI, 1.33-1.90), Black/African American race (OR, 1.44; 95% CI, 1.17-1.77), younger than 12 years (OR, 1.81; 95% CI, 1.51-2.18), obesity (OR, 1.76; 95% CI, 1.40-2.22), and not having a pediatric complex chronic condition (OR, 0.72; 95% CI, 0.65-0.80). The children with MIS-C had a more inflammatory laboratory profile and severe clinical phenotype, with higher rates of invasive ventilation (117 of 707 [16.5%] vs 514 of 8241 [6.2%]; P < .001) and need for vasoactive-inotropic support (191 of 707 [27.0%] vs 426 of 8241 [5.2%]; P < .001) compared with those who had acute COVID-19. Comparing children during the Delta vs pre-Delta eras, there was no significant change in hospitalization rate (1738 [6.0%] vs 8507 [6.2%]; P = .18) and lower odds for severe disease (179 [10.3%] vs 1242 [14.6%]) (decreased by a factor of 0.67; 95% CI, 0.57-0.79; P < .001). In this cohort study of US children with SARS-CoV-2, there were observed differences in demographic characteristics, preexisting comorbidities, and initial vital sign and laboratory values between severity subgroups. Taken together, these results suggest that early identification of children likely to progress to severe disease could be achieved using readily available data elements from the day of admission. Further work is needed to translate this knowledge into improved outcomes.

CT-severity score in COVID-19 patients: for whom is it applicable best?

Background:lung involvement in COVID-19 can be quantified by chest CT scan. We evaluated the triage and prognostication performance of seven proposed CT-severity score (CTSS) systems in two age groups of ≥65 and <65 years old.Methods: Confirmed COVID-19 patients by reverse transcriptase polymerase chain reaction (RT-PCR) admitted from February 20th, 2020 to July 22nd were included in a retrospective single center study. Clinical disease severity at presentation and at peak disease severity were recorded. CT images were scored according to seven different scoring systems (CTSS1-CTSS7). The cohort was divided into two age groups of ≥65 and <65 years old. Receiver operator characteristic (ROC) curves for each age group for diagnosis of severe/critical disease on admission (for triage) were plotted. Such curves were also plotted for predicting severe/critical disease at peak disease severity (for prognostication), and critical disease at peak severity (for prognostication). Areas under the curve (AUCs), best thresholds, and corresponding sensitivities (Sens.) and specificities (Spec.) were calculated.Results:96 patients were included with a mean age of 63.6±17.4 years. All CTSSs in 65-year-old or more group (N=55) showed excellent performance (AUC=0.80-0.83, Sens.+Spec.= 155-162%) in triage and excellent or outstanding performance (AUC=0.81-0.92, Sens.+Spec.= 153-177%) in prognostication. In the younger group (N=44), all CTSSs were unsatisfactory for triage (AUC=0.49-0.57) and predicting severe/critical disease (AUC=0.67-0.70), but were acceptable for predicting critical disease (AUC=0.70-0.73, Sens.+Spec.= 132-151%).Conclusion:CTSS is an excellent tool in triage and prognostication in patients with COVID-19 ≥65 years old, but is of limited value in younger patients.

COVID-19 Outcomes and Genomic Characterization of SARS-CoV-2 Isolated From Veterans in New England States: Retrospective Analysis.

BackgroundClinical and virologic characteristics of COVID-19 infections in veterans in New England have not been described. The average US veteran is a male older than the general US population. SARS-CoV-2 infection is known to cause poorer outcomes among men and older adults, making the veteran population an especially vulnerable group for COVID-19.ObjectiveThis study aims to evaluate clinical and virologic factors impacting COVID-19 outcomes.MethodsThis retrospective chart review included 476 veterans in six New England states with confirmed SARS-CoV-2 infection between April and September 2020. Whole genome sequencing was performed on SARS-CoV-2 RNA isolated from these veterans, and the correlation of genomic data to clinical outcomes was evaluated. Clinical and demographic variables were collected by manual chart review and were correlated to the end points of peak disease severity (based on oxygenation requirements), hospitalization, and mortality using multivariate regression analyses.ResultsOf 476 veterans, 274 had complete and accessible charts. Of the 274 veterans, 92.7% (n=254) were men and 83.2% (n=228) were White, and the mean age was 63 years. In the multivariate regression, significant predictors of hospitalization (C statistic 0.75) were age (odds ratio [OR] 1.05, 95% CI 1.03-1.08) and non-White race (OR 2.39, 95% CI 1.13-5.01). Peak severity (C statistic 0.70) also varied by age (OR 1.07, 95% CI 1.03-1.11) and O2 requirement on admission (OR 45.7, 95% CI 18.79-111). Mortality (C statistic 0.87) was predicted by age (OR 1.06, 95% CI 1.01-1.11), dementia (OR 3.44, 95% CI 1.07-11.1), and O2 requirement on admission (OR 6.74, 95% CI 1.74-26.1). Most (291/299, 97.3%) of our samples were dominated by the spike protein D614G substitution and were from SARS-CoV-2 B.1 lineage or one of 37 different B.1 sublineages, with none representing more than 8.7% (26/299) of the cases.ConclusionsIn a cohort of veterans from the six New England states with a mean age of 63 years and a high comorbidity burden, age was the largest predictor of hospitalization, peak disease severity, and mortality. Non-White veterans were more likely to be hospitalized, and patients who required oxygen on admission were more likely to have severe disease and higher rates of mortality. Multiple SARS-CoV-2 lineages were distributed in patients in New England early in the COVID-19 era, mostly related to viruses from New York State with D614G mutation.

Performance of Cytokine Storm Syndrome Scoring Systems in Pediatric COVID‐19 and Multisystem Inflammatory Syndrome in Children

ObjectiveThe objective of this study is to evaluate pediatric patients using existing macrophage activation syndrome (MAS) and hemophagocytic lymphohistiocytosis (HLH) scoring systems to determine how these systems identify patients with cytokine storm syndrome (CSS) in the setting of a multisystem inflammatory syndrome in children (MIS‐C) and active coronavirus disease 2019 (COVID‐19) infection.MethodsHospitalized pediatric patients with MIS‐C and active COVID‐19 infection at a single institution were identified. Infectious data, clinical findings, and laboratory values were collected, and patients were stratified by disease severity. Eight historically used scoring systems for MAS, HLH, and CSS were examined in the cohort of patients with MIS‐C and pediatric COVID‐19.ResultsThe HLH‐2004 criteria and HScore did not identify any patients as having CSS on admission, with only one patient with COVID‐19 meeting criteria at peak disease severity. The 2016 systemic juvenile idiopathic arthritis (sJIA)/MAS criteria, ferritin/erythrocyte sedimentation rate (ESR) ratio, and COVID‐19 CSS Quick Score most frequently identified CSS in this population and distinguished between COVID‐19 and MIS‐C hyperinflammation. The 2019 MAS/sJIA (MS) score and the COVID‐19–associated hyperinflammatory syndrome (cHIS) criteria were less likely to identify CSS, as the MS score overestimated CSS and the cHIS resulted in similar scores regardless of severity or disease type. The Caricchio COVID‐Cytokine Storm (COVID‐CS) criteria identified patients with COVID‐19 frequently but was less useful in MIS‐C because of its COVID‐19‐specific criteria.ConclusionMIS‐C and pediatric COVID‐19 result in relatively unique CSSs and patterns of inflammation. Existing scoring systems for CSSs likely do not capture the full breadth of this disease process in MIS‐C and pediatric COVID‐19.

Improved Assessment of Overall Health in Variably Aged Murine Models of Multiple Sclerosis With a Novel Frailty Index Tool.

The experimental autoimmune encephalomyelitis (EAE) model is the most commonly used animal model of multiple sclerosis (MS). However, phenotypic characterization of mice based on the traditional 5-point clinical paralysis scale does not fully capture disease progression. The frailty index (FI) conceptualizes frailty as the accumulation of health deficits and it is widely used to assess overall health in aging humans and preclinical models. Here, we adapted an established mouse FI tool for use in EAE mice and determined whether this could evaluate general signs of health in variably aged female EAE mice. The EAE-Clinical FI included 34 items related to clinical signs and deficits characteristic of aging and MS. This tool clearly showed more detailed EAE progression and severity at all ages, highlighting changes in systems other than motor paralysis measured with the traditional 5-point paralysis scale. When we induced disease at 3 and 6 months of age, mice showed typical EAE clinical manifestations with peak disease severity between 17 and 19 days post-induction and mean frailty scores of 0.36 ± 0.04 (3-month-old) and 0.43 ± 0.05 (6-month-old). By contrast, disease severity peaked after 14 days in 12-month-old mice. They showed atypical signs including wobbling, early belly drag, and splayed hindlegs that were better captured with the EAE-Clinical FI. Peak frailty scores also were higher than those of younger animals (0.54 ± 0.04). As MS most often develops in young to middle-aged people, this new tool may have significant value for use in EAE animal studies as a first step toward translation to people with MS.

Complement levels at admission as a reflection of coronavirus disease 2019 (COVID-19) severity state.

Complement system hyperactivation has been proposed as a potential driver of adverse outcomes in severe acute respiratory syndrome coronavirus 2 infected patients, given prior research of complement deposits found in tissue and blood samples, as well as evidence of clinical improvement with anticomplement therapy. Its role in augmenting thrombotic microangiopathy mediated organ damage has also been implicated in coronavirus disease 2019 (COVID‐19). This study aimed to examine associations between complement parameters and progression to severe COVID‐19 illness, as well as correlations with other systems. Blood samples of COVID‐19 patients presenting to the emergency department (ED) were analyzed for a wide panel of complement and inflammatory biomarkers. The primary outcome was COVID‐19 severity at index ED visit, while the secondary outcome was peak disease severity over the course of illness. Fifty‐two COVID‐19 patients were enrolled. C3a (p = 0.018), C3a/C3 ratio (p = 0.002), and sC5b‐9/C3 ratio (p = 0.021) were significantly elevated in with severe disease at ED presentation. Over the course of illness, C3a (p = 0.028) and C3a/C3 ratio (p = 0.003) were highest in the moderate severity group. In multivariate regression controlled for confounders, complement hyperactivation failed to predict progression to severe disease. C3a, C3a/C3 ratio, and sC5b‐9/C3 ratio were correlated positively with numerous inflammatory biomarkers, fibrinogen, and VWF:Ag, and negatively with plasminogen and ADAMTS13 activity. We found evidence of complement hyperactivation in COVID‐19, associated with hyperinflammation and thrombotic microangiopathy. Complement inhibition should be further investigated for potential benefit in patients displaying a hyperinflammatory and microangiopathic phenotype.

Alpha coma EEG pattern in patients with severe COVID-19 related encephalopathy

ObjectiveEncephalopathy is a major neurological complication of severe Coronavirus Disease 2019 (COVID-19), but has not been fully defined yet. Further, it remains unclear whether neurological manifestations are primarily due to neurotropism of the virus, or indirect effects, like cerebral hypoxia. MethodsWe analysed the electroencephalograms (EEGs) of 19 consecutive patients with laboratory-confirmed COVID-19, performed at peak disease severity as part of their clinical management. Disease severity, respiratory failure, immune and metabolic dysfunction, sedation status, and neurological examination on the day of the EEG were noted. ResultsSevere encephalopathy was confirmed in 13 patients, all with severe COVID-19; 10 remained comatose off sedation, and five of them had alpha coma (AC). Disease severity, sedation, immune and metabolic dysfunction were not different between those with AC and those without. ConclusionsSevere COVID-19 encephalopathy is a principal cause of persisting coma after sedation withdrawal. The relatively high incidence of the rare AC pattern may reflect direct SARS-CoV-2 neurotropism with a predilection for the brainstem ascending reticular system. SignificanceSystematic early EEG detection of encephalopathy related to severe COVID-19 is important for the acute care and the management of long-term neurological and cognitive sequelae, and may help our better understanding of its pathophysiology.

Aging impairs the cellular immune response to acute C. difficile infection

Abstract Clostridium difficile (C. difficile) is a spore-forming bacterium that produces enterotoxins that damage the intestinal epithelium. The incidence of C. difficile infection (CDI) increased significantly over the last two decades, particularly among those ≥65 years of age. The objective of this study was to determine the role of age in immune responses to C. difficile infection in mice, utilizing strains of varying virulence. We have shown that mice infected with C. difficile strain VPI 10463 (VPI) exhibit more severe disease compared to C. difficile strain 630. Thus, to examine an age-related effect on immune responses in CDI, young mice (2–3 months old) and aged mice (22–24 months old) were rendered susceptible with cefoperazone and then infected with either C. difficile strain 630 or VPI. Cecum and colon were harvested during peak disease severity (day 2–4 post-infection) and processed to assess intestinal immune cell profile by flow cytometry. We found that CD45+ immune cell infiltration occurs early in the cecum of mice infected with VPI, but not in 630, regardless of age. Of the CD45+ immune cells in the cecum and colon of VPI-infected mice, the majority were CD11b+ myeloid cells. While young mice infected with C. difficile strain VPI showed a robust increase in cecal eosinophils, there was a significant lack of an eosinophil response in aged counterparts. In summary, C. difficile strain 630 did not elicit an early cellular intestinal immune response and this was independent of age. Furthermore, aged mice demonstrated a significantly decreased local cellular immune response to C. difficile VPI 10463 infection compared to young mice, suggesting a role for age-associated immune responses in CDI outcomes in older patient populations.

Absence of endothelial \u03b15\u03b21 integrin triggers early onset of experimental autoimmune encephalomyelitis due to reduced vascular remodeling and compromised vascular integrity

Early in the development of multiple sclerosis (MS) and its mouse model experimental autoimmune encephalomyelitis (EAE), vascular integrity is compromised. This is accompanied by a marked vascular remodeling response, though it is currently unclear whether this is an adaptive vascular repair mechanism or is part of the pathogenic process. In light of the well-described angiogenic role for the α5β1 integrin, the goal of this study was to evaluate how genetic deletion of endothelial α5 integrin (α5-EC-KO mice) impacts vascular remodeling and repair following vascular disruption during EAE pathogenesis, and how this subsequently influences clinical progression and inflammatory demyelination. Immunofluorescence staining revealed that fibronectin and α5 integrin expression were strongly upregulated on spinal cord blood vessels during the pre-symptomatic phase of EAE. Interestingly, α5-EC-KO mice showed much earlier onset and faster progression of EAE, though peak disease severity and chronic disease activity were no different from wild-type mice. At the histological level, earlier disease onset in α5-EC-KO mice correlated with accelerated vascular disruption and increased leukocyte infiltration into the spinal cord. Significantly, spinal cord blood vessels in α5-EC-KO mice showed attenuated endothelial proliferation during the pre-symptomatic phase of EAE which resulted in reduced vascular density at later time-points. Under pro-inflammatory conditions, primary cultures of α5KO brain endothelial cells showed reduced proliferation potential. These findings suggest that α5β1 integrin-mediated angiogenic remodeling represents an important repair mechanism that counteracts vascular disruption during the early stages of EAE development.

Detection of Glycan Shedding in the Blood: New Class of Multiple Sclerosis Biomarkers?

Multiple sclerosis (MS) is a devastating autoimmune disease, afflicting people in the prime of their lives. Presently, after initial clinical presentation, there are no reliable markers for whether a patient will develop MS, or whether their prognosis will be aggressive or relapsing-remitting. Furthermore, many MS patients do not respond to treatment. Thus, markers for diagnosis, prognosis, and treatment-responsiveness are lacking for a disease, where a precision medicine approach would be valuable. The glycocalyx (GLX) is the carbohydrate-rich outer surface of the blood vessel wall and is the first interaction between the blood and the vessel. We hypothesized that cleavage of the GLX may be an early stage predictor of immune attack, blood-brain barrier (BBB) breakdown, and disease severity in MS. Two experimental models of MS, experimental autoimmune encephalitis (EAE), were included in this study. EAE was induced in C57BL/6J mice and Lewis rats, which were monitored for weight loss and clinical presentation in comparison to healthy controls. Plasma samples were obtained longitudinally from mice until peak disease severity and at peak disease severity in rats. Soluble GLX-associated glycosaminoglycans (GAG) and proteoglycans (PG) were detected in plasma samples. All animals receiving EAE emulsion developed fulminant EAE (100% penetrance). Increased plasma levels of chondroitin sulfate were detected before the onset of clinical symptoms and remained elevated at peak disease severity. Hyaluronic acid was increased at the height of the disease, whereas heparan sulfate was transiently increased during early stages only. By contrast, syndecans 1, 3, and 4 were detected in EAE samples as well as healthy controls, with no significant differences between the two groups. In this study, we present data supporting the shedding of the GLX as a new class of biomarker for MS. In particular, soluble, sugar-based GLX components are associated with disease severity in two models of MS, molecules that would not be detected in proteomics-based screens of MS patient samples. Patient studies are presently underway.

Glucagon-Like Peptide-1 Analog, Liraglutide, Delays Onset of Experimental Autoimmune Encephalitis in Lewis Rats.

Introduction: Recent findings indicate that metabolic disturbances are involved in multiple sclerosis (MS) pathology and influence the susceptibility to treatment, directing attention toward anti-diabetic drugs such as metformin and pioglitazone. Liraglutide, a drug of the glucagon-like peptide-1 (GLP-1) family, is also anti-diabetic and weight-reducing and is, moreover, directly neuroprotective and anti-inflammatory in a broad spectrum of experimental models of brain disease. In this study we investigate the potential for this FDA-approved drug, liraglutide, as a treatment for MS by utilizing the experimental model, experimental autoimmune encephalitis (EAE).Methods: EAE was induced in 30 female Lewis rats that subsequently received twice-daily liraglutide (200 μg/kg s.c.) or saline. Healthy controls were included (saline, n = 6, liraglutide, n = 7). Clinical score and weight were assessed daily by blinded observers. Animals were killed at peak disease severity (day 11) or if exceeding humane endpoint (clinical score ≥4). Protein levels of manganese superoxide dismutase (MnSOD), amyloid precursor protein (APP), and glial fibrillary acidic protein (GFAP) were determined.Results: Liraglutide treatment delayed disease onset (group clinical score significantly >0) by 2 days and markedly reduced disease severity (median clinical score 2 vs. 5; p = 0.0003). Fourteen of 15 (93%) of vehicle-treated rats reached the humane endpoint (clinical score ≥4) by day 11 compared to 5 of 15 (33%) of liraglutide-treated rats (p = 0.0004). Liraglutide substantially increased the mitochondrial antioxidant MnSOD (p < 0.01) and reduced the neurodegenerative marker APP (p = 0.036) in the brain. GFAP levels were not significantly changed with drug treatment (p = 0.09).Conclusion: We demonstrate, for the first time, that liraglutide treatment delays onset of EAE in Lewis rats and is associated with improved protective capacity against oxidative stress. These data suggest GLP-1 receptor agonists should be investigated further as a potential therapy for MS.

Temporal and Spatial Development of Cercospora Leaf Spot of Faba Bean Influenced by In Situ Inoculum.

The temporal and spatial dynamics of Cercospora leaf spot on susceptible and resistant lines of faba bean grown in or at defined distances from soil with residues infested by Cercospora zonata were examined in South Australia in 2005 and 2006. The disease was first observed on susceptible seedlings 49 days after sowing (DAS) in soil that had been sown with faba bean every 3 years since 1997 (positive soil zone for C. zonata) but was delayed by 1 week in adjacent soil (0 to 16 m away) with no history of cultivation of faba bean (negative soil zone). The incidence of diseased seedlings from 49 to 63 DAS showed a gradient from 4 to 16 m from the infested soil and was significantly greater for susceptible plants grown in the positive versus negative soil zones in field trials conducted in 2005 and 2006 (92 versus 30% in 2005, χ21 = 32.2, P < 0.001; 98 versus 55% in 2006, χ21 = 12.1, P < 0.001). The incidence of Cercospora leaf spot on the resistant line 1322/2 was significantly less (χ26 = 171.7; P < 0.001) than on the susceptible line 'Farah' at that time in both years, with fewer than 5% of the seedlings showing the disease. However, a gradient was shown at 70 to 84 DAS, where disease incidence was significantly greater on line 1322/2 in the positive soil zone than on plants in the negative soil zone in both years (62 and 18%, respectively, with χ21 = 27.9, P < 0.001 in the 2005 trial; and 47 and 6%, respectively, with χ21 = 33.3, P < 0.001 in the 2006 trial). At peak disease severity on Farah, Cercospora leaf spot mean leaf area diseased (%LAD) was severe (85 ± 4.3%) on leaves of the three nodes closest to the soil surface, and much less severe (1 ± 0.6%) in the upper canopy. Defoliation combined with %LAD was used to describe the loss of photosynthetic leaf area (%LPLA) in both cultivars, on both soil zones, in each year. Nonlinear regression analyses using a logistic model described disease development over time on susceptible plants grown in infested soil (e.g., for +12-m blocks within infested soil, y = 2.66 + 46.08/[1 + exp(-0.23 × [X - 40.92])] in 2005 and y = 0.49 + 5.02/[1 + exp(-0.14 × [X - 28.30])] in 2006, where X = DAS and y = %LPLA, with both regressions significant at P < 0.001), whereas an exponential model (e.g., for -12-m blocks from infested soil, y = 0.23 + 0.77 × 1.04X in 2005 and y = 0.44 + 0.56 × 1.04X in 2006, both at P < 0.001) best described disease gradients with increasing distance from the inoculum source. Paired t tests of %LPLA at 77 and 98 DAS showed significant differences in disease severity in the positive versus negative soil zones and a steep gradient in %LPLA from 0 to 4 m from the inoculum source. The role of infested faba bean residue in survival of C. zonata over time was also examined using a pot-bioassay and in situ field assay. When residues were removed from the soil surface or depleted rapidly by animal grazing, the amount of C. zonata inoculum in the soil was significantly less (P < 0.001) than for soil with residue remaining on the soil surface. C. zonata survived in soil and remained infective for at least 30 months after harvest of an infected faba bean crop.

Natural Killer Cells and Innate Interferon Gamma Participate in the Host Defense against Respiratory Vaccinia Virus Infection.

In establishing a respiratory infection, vaccinia virus (VACV) initially replicates in airway epithelial cells before spreading to secondary sites of infection, mainly the draining lymph nodes, spleen, gastrointestinal tract, and reproductive organs. We recently reported that interferon gamma (IFN-γ) produced by CD8 T cells ultimately controls this disseminated infection, but the relative contribution of IFN-γ early in infection is unknown. Investigating the role of innate immune cells, we found that the frequency of natural killer (NK) cells in the lung increased dramatically between days 1 and 4 postinfection with VACV. Lung NK cells displayed an activated cell surface phenotype and were the primary source of IFN-γ prior to the arrival of CD8 T cells. In the presence of an intact CD8 T cell compartment, depletion of NK cells resulted in increased lung viral load at the time of peak disease severity but had no effect on eventual viral clearance, disease symptoms, or survival. In sharp contrast, RAG(-/-) mice devoid of T cells failed to control VACV and succumbed to infection despite a marked increase in NK cells in the lung. Supporting an innate immune role for NK cell-derived IFN-γ, we found that NK cell-depleted or IFN-γ-depleted RAG(-/-) mice displayed increased lung VACV titers and dissemination to ovaries and a significantly shorter mean time to death compared to untreated NK cell-competent RAG(-/-) controls. Together, these findings demonstrate a role for IFN-γ in aspects of both the innate and adaptive immune response to VACV and highlight the importance of NK cells in T cell-independent control of VACV in the respiratory tract. Herein, we provide the first systematic evaluation of natural killer (NK) cell function in the lung after infection with vaccinia virus, a member of the Poxviridae family. The respiratory tract is an important mucosal site for entry of many human pathogens, including poxviruses, but precisely how our immune system defends the lung against these invaders remains unclear. Natural killer cells are a type of cytotoxic lymphocyte and part of our innate immune system. In recent years, NK cells have received increasing levels of attention following the discovery that different tissues contain specific subsets of NK cells with distinctive phenotypes and function. They are abundant in the lung, but their role in defense against respiratory viruses is poorly understood. What this study demonstrates is that NK cells are recruited, activated, and contribute to protection of the lung during a severe respiratory infection with vaccinia virus.

Activation of the STING Adaptor Attenuates Experimental Autoimmune Encephalitis

Cytosolic DNA sensing activates the stimulator of IFN genes (STING) adaptor to induce IFN type I (IFN-αβ) production. Constitutive DNA sensing to induce sustained STING activation incites tolerance breakdown, leading to autoimmunity. In this study, we show that systemic treatments with DNA nanoparticles (DNPs) induced potent immune regulatory responses via STING signaling that suppressed experimental autoimmune encephalitis (EAE) when administered to mice after immunization with myelin oligodendrocyte glycoprotein (MOG), at EAE onset, or at peak disease severity. DNP treatments attenuated infiltration of effector T cells into the CNS and suppressed innate and adaptive immune responses to myelin oligodendrocyte glycoprotein immunization in spleen. Therapeutic responses were not observed in mice treated with cargo DNA or cationic polymers alone, indicating that DNP uptake and cargo DNA sensing by cells with regulatory functions was essential for therapeutic responses to manifest. Intact STING and IFN-αβ receptor genes, but not IFN-γ receptor genes, were essential for therapeutic responses to DNPs to manifest. Treatments with cyclic diguanylate monophosphate to activate STING also delayed EAE onset and reduced disease severity. Therapeutic responses to DNPs were critically dependent on IDO enzyme activity in hematopoietic cells. Thus, DNPs and cyclic diguanylate monophosphate attenuate EAE by inducing dominant T cell regulatory responses via the STING/IFN-αβ/IDO pathway that suppress CNS-specific autoimmunity. These findings reveal dichotomous roles for the STING/IFN-αβ pathway in either stimulating or suppressing autoimmunity and identify STING-activating reagents as a novel class of immune modulatory drugs.