Exposure to ultraviolet radiation enhances mortality and pathology associated with influenza virus infection in mice.

Ultraviolet radiation (UVR) causes systemic immune suppression, decreasing the delayed type and contact hypersensitivity responses in animals and humans and enhancing certain mycobacterial, parasitic and viral infections in mice. This study tests the hypothesis that prior exposure to UVR enhances influenza infections in mice. BALB/c female mice were exposed to 0-8.2 kJ/m2 of UVR. Exposed and unexposed mice were infected intranasally three days later with 150-300 plaque-forming units/mouse (lethal dose (LD)20-LD40) of mouse-adapted Hong Kong Influenza A/68 (H3N2) virus or sham infected with 50 microL Hanks' balanced salt solution/mouse. Mortality from viral infection ranged from 25-50%. UVR exposure increased virus-associated mortality in a dose-dependent manner (up to a two-fold increase at 8.2 kJ/m2). The increased mortality was not associated with bacterial pneumonia. The highest dose of UVR also accelerated the body weight loss and increased the severity and incidence of thymic atrophy associated with influenza infection. However, UVR treatment had little effect on the increase in lung wet weight seen with viral infection, and, to our surprise, did not cause an increase in virus titers in the lung or dissemination of virus. The mice died 5-6 days after infection, too early for adaptive immune responses to have much impact. Also, UVR did not interfere with the development of protective immunity to influenza, as measured by reinfection with a lethal challenge of virus. Also, cells adoptively transferred from UVR or untreated mice were equally protective of recipient mice challenged with a lethal dose of virus. The mice resemble mice succumbing to endotoxin, and influenza infection increased the levels of tumor necrosis factor alpha (TNF-alpha) in bronchoalveolar lavage fluid and serum cortisol levels; however, UVR preexposure did not increase either of these responses to the virus. The results show that UVR increased the morbidity, mortality and pathogenesis of influenza virus in mice without affecting protective immunity to the virus, as measured by resistance to reinfection. The mechanism of enhanced mortality is uncertain, but the data raises concerns that UVR may exacerbate early responses that contribute to the pathogenesis of a primary viral infection.

Solubilization and display of G protein-coupled receptors on beads for real-time fluorescence and flow cytometric analysis.

G protein-coupled receptors (GPCR) and cellular signaling elements are prime targets for drug discovery. Sensitive real-time methods that expand the analytical capabilities for these elements can play significant roles in basic research and drug discovery. Here, we describe novel approaches for the real-time fluorescence analysis of GPCRs. Using the G protein-coupled N-formyl peptide receptor (FPR) as a model system in concert with a fluorescent ligand, we showed the quantitative solubilization of his-tagged FPRs in 1% dodecyl maltoside. Solubilized receptors reconstitute in dodecyl maltoside with a mixture of bovine brain Gi/Go showing an apparent Kd of 100 nM. Solubilized receptors were also bound to Ni(2+)-silica particles and were detected in a flow cytometer by the binding of fluorescent ligand. The efficiency of receptor uptake by the particles was in excess of 80% with an apparent affinity for the bead in the nM range. The receptors had largely homogeneous dissociation characteristics, an appropriate Kd for the ligand in the low nM range and a high site number, with several million receptor molecules per particle. However, the G protein reconstitution was not detected on the beads, apparently for steric reasons. These approaches for displaying receptors could prove useful in drug discovery and in the analysis of the molecular assemblies in signal transduction.

Characterization of upper respiratory disease in rats following neonatal inoculation with a rat-adapted influenza virus.

Neonatal F344 rats were infected with a rat-adapted influenza virus (RAIV) to use as a potential model to study the combined effects of air pollutant exposure with early life respiratory viral infections. Initially, 6-day-old pups were intranasally inoculated with RAIV or medium alone, and nasal and lower respiratory tract (LRT) tissues were assessed histologically at 1, 3, 6, and 13 days postinoculation (DPI). Immunologic assessments included thymic lymphocyte quantification and anti-RAIV immunoglobulin production. Pups then received two inoculations (at 6 and 30 days of age), with histologic and immunologic assessment 6 and 13 days after the second inoculation and bronchoprovocation testing 5-8 weeks later. Following the single RAIV inoculation, IgM and IgG1 measurements increased at 6, 11, and 15 DPI, with IgG1 being greater at 11 and 15 DPI. Nasal lesions were evident as early as 1 DPI and primarily involved the anterior dorsal medial meatus and adjacent dorsal atrio- and nasoturbinates. Alterations included epithelial cell exfoliation and necrosis, mild erosions, suppurative and nonsuppurative inflammation, intraepithelial neutrophil accumulations, and intraluminal exudate. By 3 DPI, olfactory epithelial damage was multifocal or locally diffuse, with degeneration of sensory cells and variable inflammation. By 13 DPI, lesions were essentially repaired. Minimal changes were apparent in the LRT despite evidence of viral replication in the lungs 24 hours after inoculation (> 3 log10 plaque-forming units/lung). Pups reinoculated with RAIV at 30 days of age did not develop significant histologic lesions, nor did they exhibit increased airway responsiveness when assessed as young adults. In spite of their immature immune status at the time of initial infection, 13 days after the second RAIV inoculation, IgG1 increased substantially. Thus, neonatal RAIV infection resulted in acute nasal epithelial injury and inflammation, alterations that may allow subsequent evaluation of viral disease-air pollutant interactions.