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Real time quantitative PCR analysis of murine macrophages

ABSTRACT: Murine macrophages were isolated from the lungs of mice given a pulmonary challenge with C. neoformans strain H99. Mice were either given a protective (H99γ) or a mock (HKCn) immunization prior to C. neoformans H99 challenge, and macrophages were isolated from the lungs of mice 24 hours, 3 days, or 7 days post-challenge using anti-CD11b microbeads according to the Miltenyi cell sorting system. We used SA Biosciences Toll-like Receptor PCR assay panel to quantitate gene expression of signal transduction factors in total RNA isolated from macrophages derived from immunized mice compared to non-immunized.

ORGANISM(S): Mus musculus

PROVIDER: GSE39990 | GEO | 2012/08/09

SECONDARY ACCESSION(S): PRJNA172210

REPOSITORIES: GEO

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Real time quantitative PCR analysis of murine macrophages

Project description:Murine macrophages were isolated from the lungs of mice given a pulmonary challenge with C. neoformans strain H99. Mice were either given a protective (H99γ) or a mock (HKCn) immunization prior to C. neoformans H99 challenge, and macrophages were isolated from the lungs of mice 24 hours, 3 days, or 7 days post-challenge using anti-CD11b microbeads according to the Miltenyi cell sorting system. We used SA Biosciences Toll-like Receptor PCR assay panel to quantitate gene expression of signal transduction factors in total RNA isolated from macrophages derived from immunized mice compared to non-immunized. qPCR gene expression profiling. Macrophages from 5 mice per group were pooled and assayed as indicated in the summary. Each experiment was performed 3 times and the resulting Ct values of each group from each experiment averaged prior to data analysis. TIme points were analyzed separately

2012-08-09 | E-GEOD-39990 | biostudies-arrayexpress

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Arginase 1 expression by macrophages promotes Cryptococcus neoformans proliferation and invasion into brain microvascular endothelial cells.

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Gene Expression Analysis of Host Immune Responses in CD11b positive cells isolated from lungs of the UV inactivated SARS-CoV-immunized mice following Lethal SARS-CoV Infection

Project description:To better understand the biological pathways by which UV inactivated SARS-CoV-induced pulmonary eosinophilia occurs, we examined global transcriptional changes in macrophages from the lungs of mouse. Female BALB/c mice were used at 21 weeks of age. Mice were subcutaneously immunized with UV inactivated SARS-CoV (UV-V) or UV-V and Toll like receptor (TLR) ligands at 6 and 1 weeks prior to mouse-adapted SARS-CoV (n=6 per group). Mice were intranasally challenged with 1E+6 TCID50 in 30M-NM-<L. MEM was challenged in six mice as control infection. Mice were sacrificed and collected lungs at 1 days after challenge, then CD11b positive cells were isolated from the lungs of these mice. These cells were used for the analysis of microarray.

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Expression data analysis of murine pulmonary cryptococcosis induced by Cryptococcus gattii and Cryptococcus neoformans

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Pianalto KM et al "Characterization of novel components of the environmental pH-sensing complex in Cryptococcus neoformans"

Project description:Three analyses performed by the Duke Proteomics and Metabolomics Shared Resource in support of the project entitled "". Sample ID21519 was called Rra1-GFP, a 1D coomassie-stained gel band from a GFP-Trap pulldown of Rra1-GFP from the membrane protein fraction of Cryptococcus neoformans. Sample ID21570 was called no-GFP control, and consisted a GFP-Trap pulldown from the insoluble fraction of C. neoformans H99 lysate (non GFP expressing). ID21571 was called Rra1-GFP, and consisted of a GFP-Trap pulldown from the insoluble fraction of C. neoformans H99 lysate containing Rra1-GFP.

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Immunization with Attenuated Equine Herpesvirus 1 Strain KyA Induces Innate Immune Responses that Protect Mice from Lethal Challenge

Project description:Equine herpesvirus 1 (EHV-1) is a major pathogen affecting equines worldwide and causes respiratory disease, abortion, and in some cases, neurological disease. EHV-1 strain KyA is attenuated in the mouse and equine, whereas wild-type strain RacL11 induces severe inflammatory infiltration of the lung, causing infected mice to succumb at 4 to 6 days post-infection. Our previous results showed that EHV-1 KyA immunization protected CBA mice from pathogenic RacL11 challenge at 2 and 4 weeks post-immunization, and that the infection with the attenuated KyA elicits protective humoral and cell-mediated immune responses. To investigate the protective mechanisms of EHV-1 KyA by innate immune responses, CBA mice immunized with live KyA were challenged with RacL11 at various times post-vaccination. KyA immunization effectively protected CBA mice from RacL11 challenge at 1 to 7 days post-immunization. Immunized mice lost less than 10% of their preinfection body weight and rapidly regained body weight. Lung virus titers in EHV-1 KyA-immunized CBA mice were 1,000-fold lower at 2 days post-RacL11 challenge than lungs of non-immunized mice, which was indicative of accelerated virus clearance. Affymetrix microarray analysis revealed that IFN-γ and 16 antiviral interferon-stimulated genes (ISGs) were upregulated 3.1- to 48.2-fold at 8 h post-challenge in the lungs of RacL11-challenged mice that had been immunized with KyA. Murine IFN-γinhibited EHV-1 infection of murine alveolar macrophage MH-S cells and effectively protected mice against lethal EHV-1 challenge, suggesting that IFN-γ expression may be important in mediating protection elicited by KyA immunization. These results suggest that EHV-1 KyA can be used as a live attenuated EHV-1 vaccine as well as a prophylactic agent in horses.

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