Metabolomics,Unknown,Transcriptomics,Genomics,Proteomics

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Expression data from Middle Ear Mucosal Metaplasia in Mice

ABSTRACT: Chronic Otitis Media (OM) develops after sustained inflammation and is characterized by secretory middle ear epithelial metaplasia and effusion, most frequently mucoid. Non-typeable Haemophilus influenzae (NTHi), the most common acute OM pathogen, is known to activate inflammation and mucin expression in vitro and in animal models of OM. The goals of this study were to: examine expression profiling epithelial effects of NTHi challenge in murine middle ears. We used microarrays to detail examine the global programme of gene expression underlying epithelial effects of NTHi challenge in murine middle ears during this study. Weekly transtympanic inoculation of Balb/c mice with 300 Âµg/ml of NTHi lysates vs saline was performed. Bacteria were grown on chocolate agar at 37ÂºC in 5% CO2 overnight and inoculated in brain heart infusion (BHI) broth supplemented with 3.5 mg of nicotinamide adenine dinucleotide per ml. After overnight incubation, bacteria were subcultured into 5 ml of fresh brain heart infusion (BHI) and upon reaching log phase growth, NTHi were washed and suspended in phosphate-buffered saline (PBS) followed by sonication for lysis. Three transtympanic inoculation of 6 Balb/c mice middle ears (3 animals, 6 ears) with 50 uL of 300 ug/ml of NTHi bacterial lysate and 6 Balb/c mice middle ears (3 animals, 6 ears) with 50 uL of 1X phosphate buffered saline (PBS) were carried out weekly over 4 weeks (injection on days 7, 14, and 21). On day 28, the mice were euthanized and their bullae harvested. Expression microarray analysis was performed at 1 and 7 days. Microarray findings were validated in independent animal samples and in a cultured murine middle ear epithelial cell (mMEEC) line.

ORGANISM(S): Mus musculus

SUBMITTER: Svetlana Ghimbovschi

PROVIDER: E-GEOD-40087 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:Influenza A virus (IAV) predisposes individuals to secondary infections with the bacterium Streptococcus pneumoniae (the pneumococcus). Infections may manifest as pneumonia, sepsis, meningitis or otitis media (OM). It remains controversial as to whether secondary pneumococcal disease is due to the induction of an aberrant immune response or IAV induced immunosuppression. Moreover, as the majority of studies have been performed in the context of pneumococcal pneumonia, it remains unclear how far these findings can be extrapolated to other pneumococcal disease phenotypes. Here, we demonstrate that the viral hemagglutinin (HA) mediates bacterial OM by inducing a pro-inflammatory response in the middle ear cavity in a replication-dependent manner. Importantly, our findings show that it is the inflammatory response that mediates pneumococcal replication; not viral suppression of the immune system or epithelial damage. This study provide the first evidence that HA induced inflammation drives pneumococcal replication in the middle ear cavity, which has important consequences to the treatment of pneumococcal OM. Five-day old C57BL/6 mice were colonised intranasally (i.n.) with 2M-CM-^W103 colony forming units (CFU) of S. pneumoniae EF3030Lux in 3 M-BM-5Ls of PBS. Alternatively, mice were mock-infected with an equivalent volume of phosphate buffered saline (PBS). At 14-days of age, infant mice were infected i.n. with 20 plaque forming units (PFU) (PR8/34, Cambridge/34 and WSN/33) or 102.5 PFU (all other virus strains) of egg-grown IAV in 3 M-BM-5Ls of PBS. Viral doses were selected to ensure a reproducible infection with minimum morbidity and no mortality. Six days post-IAV infection, mice were euthanised and organs were collected for analysis. Six independent biological replicates (where both ears from one mouse were pooled to create one sample) were used for each condition and analyzed using the NimbleGen platform (12M-CM-^W135K Mouse Gene Expression Arrays, Roche Nimblegen, USA). Indirect labeling, hybridization and washing was performed according to the manufacturerM-bM-^@M-^Ys instructions. Array images were acquired with a NimbleGen MS200 scanner, and images were processed with NimbleScan software using the RMA algorithm. Data was processed using Arraystar (DNASTAR, USA) with default settings as described in the manual. Differential expression tests were performed with a moderated T-test implemented in Arraystar, followed by FDR correction of the P values (Q-values) according to the method of Storey and Tibshirani

Project description:Pneumonia is a serious problem worldwide. We recently demonstrated that innate defense mechanisms of the lung are highly inducible against pneumococcal pneumonia. To determine the breadth of protection conferred by stimulation of lung mucosal innate immunity, and to identify cells and signaling pathways activated by this treatment, mice were treated with an aerosolized bacterial lysate, then challenged with lethal doses of bacterial and fungal pathogens. Mice were highly protected against a broad array of Gram-positive, Gram-negative, and Class A bioterror bacterial pathogens, and Aspergillus fumigatus. Protection was associated with rapid pathogen killing within the lungs, and this effect was recapitulated in vitro using a respiratory epithelial cell line. Gene expression analysis of lung tissue showed marked activation of NF-kappaB, Type I and II interferon, and antifungal Card9-Bcl10-Malt1 pathways. Cytokines were the most strongly induced genes, but the inflammatory cytokines TNF and IL-6 were not required for protection. Lung-expressed antimicrobial peptides were also highly upregulated. Taken together, stimulated innate resistance (StIR) appears to occur through the activation of multiple host defense signaling pathways in lung epithelial cells, inducing rapid pathogen killing, and conferring broad protection against virulent bacterial and fungal pathogens. Augmentation of innate antimicrobial defenses of the lungs might have therapeutic value for protection of patients with neutropenia or impaired adaptive immunity against opportunistic pneumonia, and for defense of immunocompetent subjects against a bioterror threat or epidemic respiratory infection. Keywords: differential gene expression; time course; innate immunity; pneumonia; immunocompromised host; lung epithelium Gene expression patterns in mouse lung homogenates were analyzed 2h after exposure to aerosolized PBS (Sham treatment), 2h after exposure to aerosolized NTHi lysate or 4h after exposure to aerosolized NTHi lysate. Each group consisted of six mice.

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Expression data from Middle Ear Mucosal Metaplasia in Mice

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