Thermal & Infrared Analyses of Calcium & Lead Hydroxylapatites & Their Solid Solutions

904-90

Preparation, Characterization & Solubility of Arsenic Hydroxylapatite

1014-101

Neutrophilia and NETopathy as Key Pathologic Drivers of Progressive Lung Impairment in Patients With COVID-19

There is an urgent need for new therapeutic strategies to contain the spread of the novel coronavirus disease 2019 (COVID-19) and to curtail its most severe complications. Severely ill patients experience pathologic manifestations of acute respiratory distress syndrome (ARDS), and clinical reports demonstrate striking neutrophilia, elevated levels of multiple cytokines, and an exaggerated inflammatory response in fatal COVID-19. Mechanical respirator devices are the most widely applied therapy for ARDS in COVID-19, yet mechanical ventilation achieves strikingly poor survival. Many patients, who recover, experience impaired cognition or physical disability. In this review, we argue the need to develop therapies aimed at inhibiting neutrophil recruitment, activation, degranulation, and neutrophil extracellular trap (NET) release. Moreover, we suggest that currently available pharmacologic approaches should be tested as treatments for ARDS in COVID-19. In our view, targeting host-mediated immunopathology holds promise to alleviate progressive pathologic complications of ARDS and reduce morbidities and mortalities in severely ill patients with COVID-19

High susceptibility to lipopolysaccharide-induced lethal shock in encephalomyocarditis virus-infected mice

Secondary bacterial infection in humans is one of the pathological conditions requiring clinical attention. In this study, we examined the effect of lipopolysaccharide (LPS) on encephalomyocarditis virus (EMCV) infected mice. All mice inoculated with EMCV at 5 days before LPS challenge died within 24 h. LPS-induced TNF-α mRNA expression was significantly increased in the brain and heart at 5 days after EMCV infection. CD11b+/TLR4+ cell population in the heart was remarkably elevated at 5 days after EMCV infection, and sorted CD11b+ cells at 5 days after EMCV infection produced a large amount of TNF-α on LPS stimulation in vivo and in vitro. In conclusion, we found that the infiltration of CD11b+ cells into infected organs is involved in the subsequent LPS-induced lethal shock in viral encephalomyocarditis. This new experimental model can help define the mechanism by which secondary bacterial infection causes a lethal shock in viral encephalomyocarditis

Matrix Metalloprotease 9 Mediates Neutrophil Migration into the Airways in Response to Influenza Virus-Induced Toll-Like Receptor Signaling

The early inflammatory response to influenza virus infection contributes to severe lung disease and continues to pose a serious threat to human health. The mechanisms by which neutrophils gain entry to the respiratory tract and their role during pathogenesis remain unclear. Here, we report that neutrophils significantly contributed to morbidity in a pathological mouse model of influenza virus infection. Using extensive immunohistochemistry, bone marrow transfers, and depletion studies, we identified neutrophils as the predominant pulmonary cellular source of the gelatinase matrix metalloprotease (MMP) 9, which is capable of digesting the extracellular matrix. Furthermore, infection of MMP9-deficient mice showed that MMP9 was functionally required for neutrophil migration and control of viral replication in the respiratory tract. Although MMP9 release was toll-like receptor (TLR) signaling-dependent, MyD88-mediated signals in non-hematopoietic cells, rather than neutrophil TLRs themselves, were important for neutrophil migration. These results were extended using multiplex analyses of inflammatory mediators to show that neutrophil chemotactic factor, CCL3, and TNFα were reduced in the Myd88−/− airways. Furthermore, TNFα induced MMP9 secretion by neutrophils and blocking TNFα in vivo reduced neutrophil recruitment after infection. Innate recognition of influenza virus therefore provides the mechanisms to induce recruitment of neutrophils through chemokines and to enable their motility within the tissue via MMP9-mediated cleavage of the basement membrane. Our results demonstrate a previously unknown contribution of MMP9 to influenza virus pathogenesis by mediating excessive neutrophil migration into the respiratory tract in response to viral replication that could be exploited for therapeutic purposes

Large Scale Comparison of Innate Responses to Viral and Bacterial Pathogens in Mouse and Macaque

Viral and bacterial infections of the lower respiratory tract are major causes of morbidity and mortality worldwide. Alveolar macrophages line the alveolar spaces and are the first cells of the immune system to respond to invading pathogens. To determine the similarities and differences between the responses of mice and macaques to invading pathogens we profiled alveolar macrophages from these species following infection with two viral (PR8 and Fuj/02 influenza A) and two bacterial (Mycobacterium tuberculosis and Francisella tularensis Schu S4) pathogens. Cells were collected at 6 time points following each infection and expression profiles were compared across and between species. Our analyses identified a core set of genes, activated in both species and across all pathogens that were predominantly part of the interferon response pathway. In addition, we identified similarities across species in the way innate immune cells respond to lethal versus non-lethal pathogens. On the other hand we also found several species and pathogen specific response patterns. These results provide new insights into mechanisms by which the innate immune system responds to, and interacts with, invading pathogens

Multifunctional Adaptive NS1 Mutations Are Selected upon Human Influenza Virus Evolution in the Mouse

The role of the NS1 protein in modulating influenza A virulence and host range was assessed by adapting A/Hong Kong/1/1968 (H3N2) (HK-wt) to increased virulence in the mouse. Sequencing the NS genome segment of mouse-adapted variants revealed 11 mutations in the NS1 gene and 4 in the overlapping NEP gene. Using the HK-wt virus and reverse genetics to incorporate mutant NS gene segments, we demonstrated that all NS1 mutations were adaptive and enhanced virus replication (up to 100 fold) in mouse cells and/or lungs. All but one NS1 mutant was associated with increased virulence measured by survival and weight loss in the mouse. Ten of twelve NS1 mutants significantly enhanced IFN-β antagonism to reduce the level of IFN β production relative to HK-wt in infected mouse lungs at 1 day post infection, where 9 mutants induced viral yields in the lung that were equivalent to or significantly greater than HK-wt (up to 16 fold increase). Eight of 12 NS1 mutants had reduced or lost the ability to bind the 30 kDa cleavage and polyadenylation specificity factor (CPSF30) thus demonstrating a lack of correlation with reduced IFN β production. Mutant NS1 genes resulted in increased viral mRNA transcription (10 of 12 mutants), and protein production (6 of 12 mutants) in mouse cells. Increased transcription activity was demonstrated in the influenza mini-genome assay for 7 of 11 NS1 mutants. Although we have shown gain-of-function properties for all mutant NS genes, the contribution of the NEP mutations to phenotypic changes remains to be assessed. This study demonstrates that NS1 is a multifunctional virulence factor subject to adaptive evolution

Adaption of Seasonal H1N1 Influenza Virus in Mice

The experimental infection of a mouse lung with influenza A virus has proven to be an invaluable model for studying the mechanisms of viral adaptation and virulence. The mouse adaption of human influenza A virus can result in mutations in the HA and other proteins, which is associated with increased virulence in mouse lungs. In this study, a mouse-adapted seasonal H1N1 virus was obtained through serial lung-to-lung passages and had significantly increased virulence and pathogenicity in mice. Genetic analysis indicated that the increased virulence of the mouse-adapted virus was attributed to incremental acquisition of three mutations in the HA protein (T89I, N125T, and D221G). However, the mouse adaption of influenza A virus did not change the specificity and affinity of receptor binding and the pH-dependent membrane fusion of HA, as well as the in vitro replication in MDCK cells. Notably, infection with the mouse adapted virus induced severe lymphopenia and modulated cytokine and chemokine responses in mice. Apparently, mouse adaption of human influenza A virus may change the ability to replicate in mouse lungs, which induces strong immune responses and inflammation in mice. Therefore, our findings may provide new insights into understanding the mechanisms underlying the mouse adaption and pathogenicity of highly virulent influenza viruses

Type I Interferon Signaling Regulates Ly6Chi Monocytes and Neutrophils during Acute Viral Pneumonia in Mice

Type I interferon (IFN-I) plays a critical role in the homeostasis of hematopoietic stem cells and influences neutrophil influx to the site of inflammation. IFN-I receptor knockout (Ifnar1−/−) mice develop significant defects in the infiltration of Ly6Chi monocytes in the lung after influenza infection (A/PR/8/34, H1N1). Ly6Chi monocytes of wild-type (WT) mice are the main producers of MCP-1 while the alternatively generated Ly6Cint monocytes of Ifnar1−/− mice mainly produce KC for neutrophil influx. As a consequence, Ifnar1−/− mice recruit more neutrophils after influenza infection than do WT mice. Treatment of IFNAR1 blocking antibody on the WT bone marrow (BM) cells in vitro failed to differentiate into Ly6Chi monocytes. By using BM chimeric mice (WT BM into Ifnar1−/− and vice versa), we confirmed that IFN-I signaling in hematopoietic cells is required for the generation of Ly6Chi monocytes. Of note, WT BM reconstituted Ifnar1−/− chimeric mice with increased numbers of Ly6Chi monocytes survived longer than influenza-infected Ifnar1−/− mice. In contrast, WT mice that received Ifnar1−/− BM cells with alternative Ly6Cint monocytes and increased numbers of neutrophils exhibited higher mortality rates than WT mice given WT BM cells. Collectively, these data suggest that IFN-I contributes to resistance of influenza infection by control of monocytes and neutrophils in the lung