Martin L. Moore

Toward Primary Prevention of Asthma. Reviewing the Evidence for Early-Life Respiratory Viral Infections as Modifiable Risk Factors to Prevent Childhood Asthma

A first step in primary disease prevention is identifying common, modifiable risk factors that contribute to a significant proportion of disease development. Infant respiratory viral infection and childhood asthma are the most common acute and chronic diseases of childhood, respectively. Common clinical features and links between these diseases have long been recognized, with early-life respiratory syncytial virus (RSV) and rhinovirus (RV) lower respiratory tract infections (LRTIs) being strongly associated with increased asthma risk. However, there has long been debate over the role of these respiratory viruses in asthma inception. In this article, we systematically review the evidence linking early-life RSV and RV LRTIs with asthma inception and whether they could therefore be targets for primary prevention efforts.

Fatal Disseminated Mouse Adenovirus Type 1 Infection in Mice Lacking B Cells or Bruton's Tyrosine Kinase

ABSTRACT Mouse adenovirus type 1 (MAV-1) infection of B-cell-deficient and Brutons tyrosine kinase (Btk)-deficient mice resulted in fatal disseminated disease resembling human adenovirus infections in immunocompromised patients. Mice lacking B cells or Btk were highly susceptible to acute MAV-1 infection, in contrast to controls and mice lacking T cells. To our knowledge, this is the first demonstration that mice with an X-linked immunodeficiency phenotype (Btk deficient) are susceptible to virus-induced disease. Mice lacking B cells or Btk on a C57BL/6 background succumbed with encephalomyelitis, hepatitis, and lymphoid necrosis. Mice lacking B cells on a BALB/c background succumbed with enteritis and hepatitis. Survival of acute MAV-1 infection correlated with early T-cell-independent neutralizing antibody and T-cell-independent antiviral immunoglobulin M. Treatment of MAV-1-infected Btk−/− mice 4 to 9 days postinfection with antiserum harvested 6 to 9 days postinfection from MAV-1-infected Btk+/+ mice was therapeutic. Our findings implicate a critical role for B-cell function in preventing disseminated MAV-1 infection, particularly production of early T-cell-independent antiviral immunoglobulin M.

A Critical Phenylalanine Residue in the Respiratory Syncytial Virus Fusion Protein Cytoplasmic Tail Mediates Assembly of Internal Viral Proteins into Viral Filaments and Particles

ABSTRACT Respiratory syncytial virus (RSV) is a single-stranded RNA virus in the Paramyxoviridae family that assembles into filamentous structures at the apical surface of polarized epithelial cells. These filaments contain viral genomic RNA and structural proteins, including the fusion (F) protein, matrix (M) protein, nucleoprotein (N), and phosphoprotein (P), while excluding F-actin. It is known that the F protein cytoplasmic tail (FCT) is necessary for filament formation, but the mechanism by which the FCT mediates assembly into filaments is not clear. We hypothesized that the FCT is necessary for interactions with other viral proteins in order to form filaments. In order to test this idea, we expressed the F protein with cytoplasmic tail (CT) truncations or specific point mutations and determined the abilities of these variant F proteins to form filaments independent of viral infection when coexpressed with M, N, and P. Deletion of the terminal three FCT residues (amino acids Phe-Ser-Asn) or mutation of the Phe residue resulted in a loss of filament formation but did not affect F-protein expression or trafficking to the cell surface. Filament formation could be restored by addition of residues Phe-Ser-Asn to an FCT deletion mutant and was unaffected by mutations to Ser or Asn residues. Second, deletion of residues Phe-Ser-Asn or mutation of the Phe residue resulted in a loss of M, N, and P incorporation into virus-like particles. These data suggest that a C-terminal Phe residue in the FCT mediates assembly through incorporation of internal virion proteins into virus filaments at the cell surface. IMPORTANCE Respiratory syncytial virus (RSV) is a leading cause of bronchiolitis and pneumonia in infants and the elderly worldwide. There is no licensed RSV vaccine and only limited therapeutics for use in infected patients. Many aspects of the RSV life cycle have been studied, but the mechanisms that drive RSV assembly at the cell surface are not well understood. This study provides evidence that a specific residue in the RSV fusion protein cytoplasmic tail coordinates assembly into viral filaments by mediating the incorporation of internal virion proteins. Understanding the mechanisms that drive RSV assembly could lead to targeted development of novel antiviral drugs. Moreover, since RSV exits infected cells in an ESCRT (endosomal sorting complexes required for transport)-independent manner, these studies may contribute new knowledge about a general strategy by which ESCRT-independent viruses mediate outward bud formation using viral protein-mediated mechanisms during assembly and budding. Respiratory syncytial virus (RSV) is a leading cause of bronchiolitis and pneumonia in infants and the elderly worldwide. There is no licensed RSV vaccine and only limited therapeutics for use in infected patients. Many aspects of the RSV life cycle have been studied, but the mechanisms that drive RSV assembly at the cell surface are not well understood. This study provides evidence that a specific residue in the RSV fusion protein cytoplasmic tail coordinates assembly into viral filaments by mediating the incorporation of internal virion proteins. Understanding the mechanisms that drive RSV assembly could lead to targeted development of novel antiviral drugs. Moreover, since RSV exits infected cells in an ESCRT (endosomal sorting complexes required for transport)-independent manner, these studies may contribute new knowledge about a general strategy by which ESCRT-independent viruses mediate outward bud formation using viral protein-mediated mechanisms during assembly and budding.

Allergen-Induced Airway Hyperresponsiveness Mediated by Cyclooxygenase Inhibition Is Not Dependent on 5-Lipoxygenase or IL-5, but Is IL-13 Dependent

Cyclooxygenase (COX) inhibition during allergic sensitization and allergen airway challenge results in augmented allergic inflammation. We hypothesized that this increase in allergic inflammation was dependent on increased generation of leukotrienes that results from COX inhibition, as leukotrienes are important proinflammatory mediators of allergic disease. To test this hypothesis, we allergically sensitized and challenged mice deficient in 5-lipoxygenase (5-LO). We found that 5-LO knockout mice that were treated with a COX inhibitor during allergic sensitization and challenge had significantly increased airway hyperresponsiveness (AHR) (p < 0.01) and airway eosinophilia (p < 0.01) compared with 5-LO knockout mice that were treated with vehicle. The proinflammatory cytokines have also been hypothesized to be critical regulators of airway inflammation and AHR. We found that the increase in airway eosinophilia seen with COX inhibition is dependent on IL-5, whereas the increase in AHR is not dependent on this cytokine. In contrast, the COX inhibition-mediated increase in AHR is dependent on IL-13, but airway eosinophilia is not. These results elucidate the pathways by which COX inhibition exerts a critical effect of the pulmonary allergen-induced inflammatory response and confirm that COX products are important regulators of allergic inflammation.

IL-13 is associated with reduced illness and replication in primary respiratory syncytial virus infection in the mouse.

Abstract The role of IL-13 in respiratory syncytial virus (RSV) immunopathogenesis is incompletely described. To assess the effect of IL-13 on primary RSV infection, transgenic mice which either overexpress IL-13 in the lung (IL-13 OE) or non-transgenic littermates (IL-13 NT) were challenged intranasally with RSV. IL-13 OE mice had significantly decreased peak viral titers four days after infection compared to non-transgenic littermates. In addition, IL-13 OE mice had significantly lower RSV-induced weight loss and reduced lung IFN-γ protein expression compared with IL-13 NT mice. In contrast, primary RSV challenge of IL-13 deficient mice resulted in a small, but statistically significant increase in viral titers on day four after infection, no difference in RSV-induced weight loss compared to wild type mice, and augmented IFN-γ production on day 6 after infection. In STAT1-deficient (STAT1 KO) mice, where primary RSV challenge produced high levels of IL-13 production in the lungs, treatment with an IL-13 neutralizing protein resulted in greater peak viral titers both four and six days after RSV and greater RSV-induced weight loss compared to mice treated with a control protein. These results suggest that IL-13 modulates illness from RSV-infection.

Cyclooxygenase Inhibition during Allergic Sensitization Increases STAT6-Independent Primary and Memory Th2 Responses

Immune sensitization and memory generation are required for the development of allergic inflammation. Our previous studies demonstrate that the cyclooxygenase (COX) metabolic pathway is actively involved in allergic responses and COX inhibition increases allergic airway inflammation in a STAT6-independent fashion. To test the hypothesis that COX inhibition augments allergic inflammation by enhancing immune sensitization and memory, we sensitized STAT6 knockout mice with an i.p. injection of OVA with aluminum hydroxide as an adjuvant and treated the mice with the COX inhibitor indomethacin or vehicle for analyses of the primary and memory immune responses. We found that COX inhibition during immune sensitization, but not the allergic challenge phase, was necessary and sufficient to increase allergic inflammation. COX inhibition during sensitization increased the numbers of mature dendritic cells and activated CD4 T cells in the spleen and augmented OVA-specific IL-5 and IL-13 responses of the splenic CD4 T cells at day 5 after sensitization. COX inhibition during sensitization also augmented allergic Th2 response to OVA challenge 90 days after the sensitization. Therefore, COX inhibition during allergic sensitization augments allergic responses by enhancing Th2 cell activation and memory generation and the proallergic effect is STAT6-independent. These findings provide a mechanistic explanation for the increased allergic inflammation previously shown in the mice treated with COX inhibitors and in COX-deficient mice and suggest that use of COX-inhibiting drugs during initial allergen exposure may increase the risk of developing allergic responses.

Nasopharyngeal Lactobacillus is associated with a reduced risk of childhood wheezing illnesses following acute respiratory syncytial virus infection in infancy

Background: Early life acute respiratory infection (ARI) with respiratory syncytial virus (RSV) has been strongly associated with the development of childhood wheezing illnesses, but the pathways underlying this association are poorly understood. Objective: To examine the role of the nasopharyngeal microbiome in the development of childhood wheezing illnesses following RSV ARI in infancy. Methods: We conducted a nested cohort study of 118 previously healthy, term infants with confirmed RSV ARI by RT‐PCR. We used next‐generation sequencing of the V4 region of the 16S ribosomal RNA gene to characterize the nasopharyngeal microbiome during RSV ARI. Our main outcome of interest was 2‐year subsequent wheeze. Results: Of the 118 infants, 113 (95.8%) had 2‐year outcome data. Of these, 46 (40.7%) had parental report of subsequent wheeze. There was no association between the overall taxonomic composition, diversity, and richness of the nasopharyngeal microbiome during RSV ARI with the development of subsequent wheeze. However, the nasopharyngeal detection and abundance of Lactobacillus was consistently higher in infants who did not develop this outcome. Lactobacillus also ranked first among the different genera in a model distinguishing infants with and without subsequent wheeze. Conclusions: The nasopharyngeal detection and increased abundance of Lactobacillus during RSV ARI in infancy are associated with a reduced risk of childhood wheezing illnesses at age 2 years.

The intersection of respiratory syncytial virus infection, innate immunity and allergic lung disease

Abstract. Respiratory syncytial virus (RSV) infection is the leading cause of bronchiolitis, pneumonia, and respiratory failure in infants younger than one year of age and is becoming more prevalently associated with serious illness and morbidity in elderly patients. A better understanding of the innate immune response to RSV infection may lead to developing effective therapy. The main objective of this chapter is to discuss the interplay between RSV infection, the environment, and the host innate immune response. Alterations in the host innate immune response as a result of allergic inflammation, ozone exposure, and ambient particulate matter may determine disease severity from RSV infection. In turn, RSV infection may modify host immunity to environmental stimuli. The impact of RSV infection on host innate immunity with comparison between the allergic asthmatic host and non-allergic, non-asthmatic host will be highlighted. Effects of RSV on inflammatory responses in the airway will also be discussed. Defining the factors that modulate the host innate immune response to RSV infection will shed light on the pathogenesis of this virus and provide insights into the development of treatment and prevention strategies.

Chapter 2 – Mouse Adenoviruses

Publisher Summary Mouse adenoviruses are useful for study of adenovirus pathogenesis in the natural host, in which mouse adenoviruses cause acute and persistent infections. The availability of immunocompetent and immunodeficient inbred mouse strains, immunological reagents for mice, and tools for genetic mapping studies combine to make mouse adenovirus studies ideal for understanding virus-host interactions. Commercial mouse suppliers standardly monitor for mouse adenoviruses; the viruses have been eliminated in commercial mouse colonies and are rare if not absent in institutional colonies. Methods for propagating and titrating the virus are described in the chapter. The molecular biology of the human adenovirus (hAds) is extensively studied since their discovery. Adenoviruses are associated with acute pneumonia in children in developing countries, where they are a major cause of illness and death. Mouse adenovirus type 1 (MAV-1) is the first mouse adenovirus to be isolated. Mutant strains of MAV-1 have been constructed by site directed mutagenesis techniques using the standard strain as the starting virus.