Formalin-inactivated respiratory syncytial virus (RSV) vaccine preparations have been shown to cause enhanced disease in naive hosts following natural infection. In this study we demonstrate a similar pattern of enhanced disease severity following primary RSV infection of IFN-nonresponsive STAT1−/− mice. STAT1−/− mice showed markedly increased illness compared with wild-type BALB/c animals following RSV inoculation despite similar lung virus titers and rates of virus clearance. Histologically, STAT1−/− animals had eosinophilic and neutrophilic pulmonary infiltrates not present in wild-type or IFN-γ−/−-infected mice. In cytokine analyses of infected lung tissue, IFN-γ was induced in both STAT1−/− and wild-type mice, with preferential IL-4, IL-5, and IL-13 induction only in the STAT1−/− animals. Eotaxin was detected in the lungs of both wild-type and STAT1−/− mice following infection, with a 1.7-fold increase over wild-type in the STAT1−/− mice. Using a peptide epitope newly identified in the RSV fusion protein, we were able to demonstrate that wild-type memory CD4+ T cells stimulated by this peptide produce primarily IFN-γ, while STAT1−/−CD4+ cells produce primarily IL-13. These findings suggest that STAT1 activation by both type I (αβ) and type II (γ) IFNs plays an important role in establishing a protective, Th1 Ag-specific immune response to RSV infection.

CD1d-deficient mice have normal numbers of T lymphocytes and natural killer cells but lack V␣14؉ natural killer T cells. Respiratory syncytial virus (RSV) immunopathogenesis was evaluated in 129؋C57BL/6, C57BL/6, and BALB/c CD1d ؊/؊ mice. CD8 ؉ T lymphocytes were reduced in CD1d ؊/؊ mice of all strains, as shown by cell surface staining and major histocompatibility complex class I tetramer analysis, and resulted in strain-specific alterations in illness, viral clearance, and gamma interferon (IFN-␥) production. Transient activation of NK T cells in CD1d؉/؉ mice by ␣-GalCer resulted in reduced illness and delayed viral clearance. These data suggest that early IFN-␥ production and efficient induction of CD8؉ -T-cell responses during primary RSV infection require CD1d-dependent events. We also tested the ability of ␣-GalCer as an adjuvant to modulate the type 2 immune responses induced by RSV glycoprotein G or formalin-inactivated RSV immunization. However, immunized CD1-deficient or ␣-GalCer-treated wild-type mice did not exhibit diminished disease following RSV challenge. Rather, some disease parameters, including cytokine production, eosinophilia, and viral clearance, were increased. These findings indicate that CD1d-dependent NK T cells play a role in expansion of CD8 ؉ T cells and amplification of antiviral responses to RSV.

To address the complex chronic effector properties of interleukin (IL)-10, we generated transgenic mice in which IL-10 was overexpressed in the lung. In these mice, IL-10 inhibited endotoxin-induced tumor necrosis factor production and neutrophil accumulation. IL-10 also caused mucus metaplasia, B and T cell-rich inflammation, and subepithelial fibrosis and augmented the levels of mRNA encoding Gob-5, mucins, and IL-13. In mice bred to have null mutations of IL-13, IL-4R␣, or STAT-6, transgenic IL-10 did not induce mucus metaplasia but did induce inflammation and fibrosis. IL-10 was also a critical mucin regulator of virus-induced mucus metaplasia. Thus, IL-10, although inhibiting lipopolysaccharide-induced inflammation, also causes mucus metaplasia, tissue inflammation, and airway fibrosis. These responses are mediated by multiple mechanisms with mucus metaplasia being dependent on and the inflammation and fibrosis being independent of an IL-13/ IL-4R␣/STAT-6 activation pathway.

Johnson, T. R., and van Densen, W. L. T. 2007. The benefits and organization of cooperative research for fisheries management. – ICES Journal of Marine Science, 64: 834–840. Drawing on research in the northeastern USA and northwestern Europe, a description is given of how cooperative research is organized and a statement made of how involving fishers in research can contribute to better fisheries management. The focus is on improving stock assessments through the collection of better fishery-dependent and -independent data and through efforts to address bycatch through gear-selectivity studies. Direct benefits of cooperative research include increased quantity and quality of data, inclusion of fishers' knowledge in science and management, improved relevance of research to fisheries management, and reduced costs of science. Indirect benefits are the buy-in of science and management by industry and improved relationships and trust between fishers and scientists (and managers). These indirect benefits are best achieved under conditions of transparency and communication. In some cases, cooperative research also provides income to the industry and supports the maintenance of fishing infrastructure. Most important, cooperative research improves capacity-building and establishes intellectual property rights within the fishing industry, and it encourages innovative approaches to management, such as adaptive and ecosystem-based approaches. Finally, guidelines for making cooperative research more effective are outlined.

Although well studied in settings of helminth infection and allergen sensitization, the combined contributions of IL-4 and IL-13 and their signaling pathways in models of viral pathogenesis have not been reported. Using a murine model of respiratory syncytial virus (RSV) infection, we evaluated the contribution of IL-13, alone and in conjunction with IL-4, during immunization with recombinant vaccinia virus expressing RSV G glycoprotein (vvGs) or with formalin-inactivated RSV (FI-RSV). We showed that both IL-4 and IL-13 activity must be inhibited to modulate G-specific responses resulting in severe RSV-induced disease. Inhibition of IL-4 or IL-13 activity alone had minimal impact on disease in vvGs-immunized mice. However, treatment of IL-4-deficient mice with IL-13Ra during vvGs immunization reduced IL-5, IL-13, and eotaxin production and pulmonary eosinophilia after RSV challenge. In contrast, FI-RSV-induced immune responses were diminished when either IL-4 or IL-13 activity was blocked. After RSV challenge, these type 2 T cell responses were also diminished in vvGs-primed IL-4Rα-deficient mice. Our data suggest that secreted vvGs uses mechanisms requiring signaling through the IL-4Rα-chain by either IL-4 or IL-13 for induction of eosinophilia and is the first description of the relative contributions of IL-4, IL-13, and their receptors in viral pathogenesis.

Respiratory syncytial virus (RSV) is an important human pathogen that can cause severe and life-threatening respiratory infections in infants, the elderly, and immunocompromised adults. RSV infection of HEp-2 cells induces the activation of RhoA, a small GTPase. We therefore asked whether RhoA signaling is important for RSV replication or syncytium formation. The treatment of HEp-2 cells with Clostridium botulinum C3, an enzyme that ADP-ribosylates and specifically inactivates RhoA, inhibited RSV-induced syncytium formation and cell-to-cell fusion, although similar levels of PFU were released into the medium and viral protein expression levels were equivalent. Treatment with another inhibitor of RhoA signaling, the Rho kinase inhibitor Y-27632, yielded similar results. Scanning electron microscopy of C3-treated infected cells showed reduced numbers of single blunted filaments, in contrast to the large clumps of long filaments in untreated infected cells. These data suggest that RhoA signaling is associated with filamentous virus morphology, cell-to-cell fusion, and syncytium formation but is dispensable for the efficient infection and production of infectious virus in vitro. Next, we developed a semiquantitative method to measure spherical and filamentous virus particles by using sucrose gradient velocity sedimentation. Fluorescence and transmission electron microscopy confirmed the separation of spherical and filamentous forms of infectious virus into two identifiable peaks. The C3 treatment of RSV-infected cells resulted in a shift to relatively more spherical virions than those from untreated cells. These data suggest that viral filamentous protuberances characteristic of RSV infection are associated with RhoA signaling, are important for filamentous virion morphology, and may play a role in initiating cell-to-cell fusion.

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Respiratory syncytial virus (RSV) interaction with epithelial and dendritic cells (DCs) is known to require divalent cations, suggesting involvement of C-type lectins. RSV infection and maturation of primary human DCs are reduced in a dose-dependent manner by EDTA. Therefore, we asked whether RSV infection involves DC-SIGN (CD209) or its isoform L-SIGN (CD299) (DC-SIGN/R). Using surface plasmon resonance analysis, we demonstrated that the attachment G glycoprotein of RSV binds both DC- and L-SIGN. However, neutralization of DC- and L-SIGN on primary human DCs did not inhibit RSV infection, demonstrating that interactions between RSV G and DC- or L-SIGN are not required for productive infection. Thus, neither DC- nor L-SIGN represents a functional receptor for RSV. However, inhibition of these interactions increased DC activation, as evidenced by significantly higher levels of alpha interferon (IFN-α), MIP-1α, and MIP-1β in plasmacytoid DCs (pDCs) exposed to RSV after neutralization of DC-and L-SIGN. To understand the molecular interactions involved, intracellular signaling events triggered by purified RSV G glycoprotein were examined in DC- and L-SIGN-transfected 3T3 cells. RSV G interaction with DC- or L-SIGN was shown to stimulate ERK1 and ERK2 phosphorylation, with statistically significant increases relative to mock-infected cells. Neutralization of DC- and L-SIGN reduced ERK1/2 phosphorylation. With increased DC activation following DC- and L-SIGN neutralization and RSV exposure, these data demonstrate that the signaling events mediated by RSV G interactions with DC/L-SIGN are immunomodulatory and diminish DC activation, which may limit induction of RSV-specific immunity.