Immune sensitization of equine bronchus: glutathione, IL-1β expression and tissue responsiveness

BACKGROUND: Increasing clinical epidemiological and experimental evidence indicates that excess of production of reactive oxygen free radicals (ROS) induced by an oxidative stress is involved in the pathogenesis of a number of human airway disorders, as well as equine recurrent airway obstruction. Free-radicals modulate the activation of transcription factors, such as nuclear factor-(NF)-κB and activator protein (AP)-1, in several different cells. This activation leads to expression of many pro-inflammatory cytokines, including interleukin (IL)-1β. We have hypothesized that equine airway sensitization might induce an oxidative stress and increase the ROS production, which in turn might enhance a production of IL-1β and airway hyperresponsiveness. METHODS: We have examined the effect of passive sensitization on IL-1β mRNA expression and electrical field stimulation (EFS)-induced contraction in equine isolated bronchi, and the potential interference of reduced-glutathione (GSH), an antioxidant, with these responses. Bronchi passively sensitized with serum from animals suffering from heaves and having high total level of IgE, and control tissues, either pretreated or not with GSH (100 μM), were used to quantify IL-1β mRNA. Other tissues were used to study the effect of EFS (3–10–25 Hz). RESULTS: Mean IL-1β mRNA expression was higher in passively sensitized than in control rings. GSH significantly (p < 0.05) reduced the IL-1β mRNA expression only in passively sensitized bronchi. ELF induced a frequency-dependent contraction in both non-sensitized and passively sensitized tissues, with a significantly greater response always observed in sensitized tissues. GSH did not modify the EFS-induced contraction in non-sensitized bronchi, but significantly (p < 0.05) decreased it in passively sensitized tissues. CONCLUSION: Our data indicate that the passive sensitization of equine bronchi induces inflammation and hyperresponsiveness. These effects might be due to an oxidative stress because a pretreatment with GSH decreased the increased IL-1β mRNA expression and responsiveness to EFS of passively sensitized bronchi

NGF Is an Essential Survival Factor for Bronchial Epithelial Cells during Respiratory Syncytial Virus Infection

Background: Overall expression of neurotrophins in the respiratory tract is upregulated in infants infected by the respiratory syncytial virus (RSV), but it is unclear where (structural vs. inflammatory cells, upper vs. lower airways) and why, these changes occur. We analyzed systematically the expression of neurotrophic factors and receptors following RSV infection of human nasal, tracheal, and bronchial epithelial cells, and tested the hypothesis that neurotrophins work as innate survival factors for infected respiratory epithelia. Methodology: Expression of neurotrophic factors (nerve growth factor, NGF; brain-derived neurotrophic factor, BDNF) and receptors (trkA, trkB, p75) was analyzed at the protein level by immunofluorescence and flow cytometry and at the mRNA level by real-time PCR. Targeted siRNA was utilized to blunt NGF expression, and its effect on virus-induced apoptosis/ necrosis was evaluated by flow cytometry following annexin V/7-AAD staining. Principal Findings: RSV infection was more efficient in cells from more distal (bronchial) vs. more proximal origin. In bronchial cells, RSV infection induced transcript and protein overexpression of NGF and its high-affinity receptor trkA, with concomitant downregulation of the low-affinity p75 NTR. In contrast, tracheal cells exhibited an increase in BDNF, trkA and trkB, and nasal cells increased only trkA. RSV-infected bronchial cells transfected with NGF-specific siRNA exhibited decreased trkA and increased p75 NTR expression. Furthermore, the survival of bronchial epithelial cells was dramaticall

Genome Sequence of Brucella abortus Vaccine Strain S19 Compared to Virulent Strains Yields Candidate Virulence Genes

The Brucella abortus strain S19, a spontaneously attenuated strain, has been used as a vaccine strain in vaccination of cattle against brucellosis for six decades. Despite many studies, the physiological and molecular mechanisms causing the attenuation are not known. We have applied pyrosequencing technology together with conventional sequencing to rapidly and comprehensively determine the complete genome sequence of the attenuated Brucella abortus vaccine strain S19. The main goal of this study is to identify candidate virulence genes by systematic comparative analysis of the attenuated strain with the published genome sequences of two virulent and closely related strains of B. abortus, 9–941 and 2308. The two S19 chromosomes are 2,122,487 and 1,161,449 bp in length. A total of 3062 genes were identified and annotated. Pairwise and reciprocal genome comparisons resulted in a total of 263 genes that were non-identical between the S19 genome and any of the two virulent strains. Amongst these, 45 genes were consistently different between the attenuated strain and the two virulent strains but were identical amongst the virulent strains, which included only two of the 236 genes that have been implicated as virulence factors in literature. The functional analyses of the differences have revealed a total of 24 genes that may be associated with the loss of virulence in S19. Of particular relevance are four genes with more than 60bp consistent difference in S19 compared to both the virulent strains, which, in the virulent strains, encode an outer membrane protein and three proteins involved in erythritol uptake or metabolism