Transcription profiling by array of influenza PR8 infected mouse lung tissue
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ABSTRACT: C57BL/6J (WT) mice were infected with influenza A virus PR/8/34 or mock infected with phosphate buffered saline (PBS). 18 hours later lung tissue was harvested for transcriptional profiling.
Project description:Antiviral responses must be regulated to rapidly defend against infection while minimizing inflammatory damage, but the mechanisms for establishing the magnitude of response within an infected cell are not well understood. miRNAs are small non-coding RNAs that negatively regulate protein levels by binding target sequences on their cognate mRNA. We profiled microRNA expression in the lungs of mice infected for 24 h with Influenza A/PR/8/34 to identify microRNAs that may regulate host response to influenza infection. Mice were infected intranasally with 10e5 pfu Influenza A/PR/8/34. Lungs were isolated after 24 hours and total RNA extracted using Trizol. Equivalent quantities of RNA from 3 infected mice were pooled.
Project description:Analysis of whole lung RNA expression 21 days following influenza infection in wild-type and IL-22 -/- mice. Total lung RNA was isolated 21 days post-infection for microarray.
Project description:We report the whole genome sequencing of human bronchial cells infected with influenza A (PR/8/34) for 8 or 24 h. Examination influenza virus-specific human gene profiling at early or late stage of viral infection.
Project description:We defined the major transcriptional responses in primary human bronchial epithelial cells (HBECs) after either infection with influenza or treatment with relevant ligands. We used four different strategies, each highlighting distinct aspects of the response. (1) cells were infected with the wild-type PR8 influenza virus that can mount a complete replicative cycle. (2) cells were transfected with viral RNA (âvRNAâ) isolated from influenza particles. This does not result in the production of viral proteins or particles and identifies the effect of RNA-sensing pathways (e.g., RIG-I.). (3) Cells were treated with interferon beta (IFNb), to distinguish the portion of the response which is mediated through Type I IFNs. (4) Cells were infected with a PR8 virus lacking the NS1 gene (âDNS1â). The NS1 protein normally inhibits vRNA- or IFNb-induced pathways, and its deletion can reveal an expanded response to infection. HBECs were stimulated with a 15 minute pulse of 1000U/ml IFNÃ (PBL, Piscataway, NJ), 100ng/ml vRNA (purified directly from PR8 virus) with LTX transfection reagent (Invitrogen; Carlsbad, California), wild type H1N1 influenza (A/PR/8/34) or ?NS1 virus (PR8 with a deleted NS1 gene, gift from Dr. Garcia-Sastre). Viruses were used at a multiplicity of infection (moi) of 5. Control samples were incubated with media or LTX under the same conditions. Cells were washed, supplemented with warm media and harvested at 11 timepoints (0, .25, .5, 1, 1.5, 2, 4, 6, 8, 12, and 18 hours post-treatment). HBECs were seeded in 6 well plates at a concentration of 250,000/well 18 hours prior to stimultaion. Cells were stimulated with a 15 minute pulse of IFNb, vRNA, infected with PR8 influenza or NS1 deleted influenza, or mock treated
Project description:Phospho Proteomics Study from Pan-Tyrosine antibody enrichment and iMac-Fe3+ enrichment for Mouse MLE15 Cell Infected by Influenza Virus
Project description:CpG 1826 binds to Toll-like receptor (TLR)9, whereas influenza virus PR8 activates pDC via TLR7. Differential stimulation of pDCs is expected to result in unique activation mechanism(s) leading to a different phenotypically and functionally matured pDC; We used microarrays to detail the global programme of gene expression underlying the maturation process of pDC activated with CpG 1826 and influenza virus PR8. We identified a distinct expression profile of upregulated immunomediators. Experiment Overall Design: Sorted pDCs were cultured for 1h and 4hs in medium control or with 5 µg/ml CpG 1826 or 300 HAU/ml purified influenza A/PR/8 virus. The first experiment (e1) included pDC in media and stimulated with CpG for 4h. In two other independent experimental batches (e2 and e3), we obtained samples of sorted pDC cultured in medium alone (med), and with CpG or PR8 (flu) for 1h and 4h. RNA extraction was performed using the RNeasy Kit (Qiagen) and hybridization on Affymetrix microarrays was performed using standard protocols. We sought to obtain homogeneous populations of pDCs at different time points under defined activation conditions in order to decipher the temporal resolution of expression profiles during the process of their maturation.
Project description:Mice were infected intranasally with 1.5x10E5 PFU and total RNA were extracted from mice lungs at day 3. RNA samples were extracted from mice lung infected or not by influenza virus.
Project description:We used microarrays to analyse the global program of gene expression in response to Influenza A (X31) infection in lungs from C57BL/6 wt, 129S7 wt and IFNAR-/- (129) mice. Mock- or 5 day influenza A (X31)-infected total lungs from mice with the indicated genotypes were collected and processed for expression profiling.
Project description:Few studies have examined the local surface remodeling of primary human nasal epithelial cells (HNECs) during viral infection. Hence, the full range of upregulated adhesion molecules during respiratory viral infections that facilitate bacterial attachment and entry remain unknown. Accordingly, the current study provides a comprehensive view of HNEC responses to influenza virus pH1N1 infection, via global proteome profiling of uninfected (Mock, n = 4) and influenza-infected (Virus-only, n = 4) HNECs with isobaric tags using relative and absolute quantitation (iTRAQ) technique. A total of 3583 proteins were detected, 89 of which were significantly increased (52 proteins) or decreased (37 proteins) in the virus-infected HNECs compared to mock-infected controls (p < 0.05).
Project description:House dust mite (HDM) extract (Greer Laboratories, Lenoir, NC) was resuspended in sterile phosphate-buffered saline (PBS) at a concentration of 0.5 mg (protein)/ml and 10 ul (5 ug dose) was administered to isofluorane-anesthetized mice intranasally. Groups of mice were infected either with influenza A or exposed to PBS. Seven days later, separate groups of mice were either exposed to saline or HDM for 3 or 6 days and lungs harvested 24 h after the last exposure, at day 4 (early phase; EP) or day 7 (late phase; LP).