Alteration in gene expression in the lung following neonatal RSV infection
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ABSTRACT:
ORGANISM(S): Mus musculus
SUBMITTER: Laura Lambert
PROVIDER: E-MTAB-3654 | biostudies-arrayexpress |
REPOSITORIES: biostudies-arrayexpress
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Project description:A few transformed cell lines and two historic strains have been extensively used to study respiratory syncytial virus (RSV). We report a thorough molecular and cell biological characterization of HEp-2 and A549 cells infected with four strains of RSV representing both major subgroups as well as historic and more contemporary genotypes -- [RSV/A/Tracy (GA1), RSV/A/Ontario (ON), RSV/B/18537 (GB1), RSV/B/Buenos Aires (BA)] -- via measurements of viral replication kinetics and viral gene expression, immunofluorescence-based imaging of gross cellular morphology and cell-associated RSV, and measurements of host response including transcriptional changes and levels of secreted cytokines and growth factors. Our findings strongly suggest 1) the existence of a conserved difference in gene expression between RSV subgroups A and B; 2) the A549 cell line is a more stringent and natural host of replicating RSV than the HEp-2 cell line; and 3) consistent with previous studies, determining the full effects of viral genetic variation in RSV pathogenesis requires model systems as tractable as transformed cell lines but better representative of the human host.
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Project description:Rationale: Respiratory syncytial virus (RSV) and Streptococcus pneumoniae are major respiratory pathogens. Co-infection with RSV and S. pneumoniae is associated with severe and often fatal pneumonia but the molecular basis for this remains unclear. Objectives: To determine if interaction between RSV and pneumococci enhances pneumococcal virulence. Methods: We used confocal microscopy and western blot to identify the receptors involved in direct binding of RSV and pneumococci, the effects of which were studied in both in vivo and in vitro models of infection. Human ciliated respiratory epithelial cell cultures were infected with RSV for 72h and then challenged with pneumococci. Pneumococci were collected after 2h exposure and changes in gene expression determined using qRT-PCR. Results: Following incubation with RSV or purified G protein, pneumococci demonstrated a significant increase in the inflammatory response and bacterial adherence to human ciliated epithelial cultures and markedly increased virulence in a pneumonia model in mice. This was associated with extensive changes in the pneumococcal transcriptome and significant upregulation in the expression of key pneumococcal virulence genes, including the gene for the pneumococcal toxin, pneumolysin. We show that mechanistically this is due to RSV G glycoprotein binding penicillin binding protein 1a. Conclusion: The direct interaction between a respiratory virus protein and the pneumococcus resulting in increased bacterial virulence and worsening disease outcome is a new paradigm in respiratory infection. Comparison of the Streptococcus pneumoniae D39 RSV treated compared to BSA Treated in BEBM medium One condition design comparision of two strains including a dye swap
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