Project description:We used microarrays to compare interferon-alpha (IFNa)- and interferon-gamma (IFNg)-stimulated genes under an equivalent biological input. The goal was to compare IFNa- and IFNg-stimulated genes, as well as to identify common and distinct sets of type I and II ISGs. Bone marrow macrophages derived from mouse bone marrow in M-CSF for 7 days. The cells were stimulated with 62U/mL IFNa and 1U/mL of IFNg for 2.5 hrs in culture. These concentrations induced equivalent STAT1 phosphorylation in BMMs.
Project description:To identify signaling pathways that are induced by macrophages infected with Histoplasma capsulatum, we examined the whole genome expression profile of murine bone marrow derived macrophages infected with conidia, the natural infectious particle of Histoplasma. Conidia induced the expression of signature Type I interferon response genes. The induction of a Type I interferon response was specific to conidia, yeast cells did not trigger the response. Macrophages that lack the Type I interferon receptor, IFNAR1, were infected and compared to wild-type macrophages. The expression of some Type I IFN response genes are dependent upon Keywords: Murine bone marrow derived macrophage transcriptional response to infection with Histoplasma capsulatum conidia We analyzed a series of 24 MEEBO arrays on which were hybed RNA amplified from bone marrow derived macrophages from C57BL/6 (WT) or ifnar1-/- mice either mock infected or infected with H. capsulatum conidia or yeast cells.
Project description:Identifying interactors of IRF1 and STAT1 in bone marrow-derived macrophages during type I and type II interferon treatment using the proximity-dependent labeling approach TurboID followed by Mass Spectrometry
Project description:To identify signaling pathways that are induced by macrophages infected with Histoplasma capsulatum, we examined the whole genome expression profile of murine bone marrow derived macrophages infected with conidia, the natural infectious particle of Histoplasma. Conidia induced the expression of signature Type I interferon response genes. The induction of a Type I interferon response was specific to conidia, yeast cells did not trigger the response. Macrophages that lack the Type I interferon receptor, IFNAR1, were infected and compared to wild-type macrophages. The expression of some Type I IFN response genes are dependent upon Keywords: Murine bone marrow derived macrophage transcriptional response to infection with Histoplasma capsulatum conidia
Project description:Toxoplasma strains are known to inhibit the expression of several interferon-gamma induced genes, and a type II strain was shown to dysregulate genome-wide responses to interferon-gamma in human fibroblasts (Kim et al., 2007, J Immunol.). In this study we aimed to determine the effect of infection with three clonal lineages of Toxoplasma, type I, II, and III strains on genome-wide interferon-gamma induced transcription in murine macrophages. We also assessed the effect of the two main Toxoplasma modulators of mouse macrophage transcription, ROP16 and GRA15 (Jensen et al., 2011, Cell Host Microbe). We used Affymetrix microarrays to analyze host cell transcription after Toxoplasma infection and interferon-gamma stimulation. RAW264.7 murine macrophages were left uninfected or infected with type I (RH), type I ?rop16 (RH ?rop16), type II (Pru), type II ?gra15 (Pru ?gra15), or type II (CEP) parasites at an MOI ~5 for 18 hours and subsequently stimulated with murine IFN-? for six hours. Plaque assays were done to assess parasite viability. Total RNA was isolated and hybridized to Affymetrix Mouse 430A 2.0 gene chips.
Project description:The CCAAT/enhancer-binding proteins (CEBPs) are transcription factors involved in hematopoietic cell development and induction of several inflammatory mediators. Here, we generated CEBP-beta (CEBPB) and CEBP-epsilon (CEBPE) double-knockout (bbee) mice and compared their phenotypes to those of single-deficient (bbEE and BBee) and wild-type (BBEE) mice. The bbee mice were highly susceptible to fatal infections and died within 2-3 months. Morphologically, their neutrophils were blocked at the myelocytes/metamyelocytes stage, and clonogenic assays of bone marrow cells indicated a significant decrease in the number of myeloid colonies of the bbee mice. In addition, the proportion of hematopoietic progenitor cells [Lin(-)Sca1(+)c-Kit(+)] in the bone marrow of the bbee mice was significantly increased, reflecting the defective differentiation of the myeloid compartment. Furthermore, microarray expression analysis of lipopolysaccharide (LPS)- and interferon-gamma (IFN-gamma)-activated bone marrow-derived macrophages from bbee compared to single knockout mice revealed decreased expression of essential immune response-related genes and networks, including some direct CEBP targets such as Marco and Clec4e. Overall, the phenotype of the bbee mice is distinct from either the bbEE or BBee mice, demonstrating that both transcription factors are crucial for the maturation of neutrophils and macrophages, as well as the innate immune system, and can at least in part compensate for each other in the single knockout mice. To rule out the regulatory influence of both CEBPB and CEBPE on macrophage-related genes, expression analysis of bone marrow-derived macrophages was performed. Macrophages were derived from murine bone marrow with the use of murine M-CSF. The macrophages were stimulated with both LPS (100 ng) and IFN-gamma (100 ng) for 24h, and RNA was extracted for array analysis. Overall, RNA was extracted from stimulated macrophages of one WT mouse, one CEBPB-KO mouse, one CEBPE-KO mouse and one double-KO mouse.
Project description:Primary murine bone marrow derived macrophages (BMDMs) of Setdb2GT/GT and WT mice were stimulated with polyI:C and harvested at 0, 2 and 8h. Everything with biological triplicates (=cells from 3 individual mice per condition)
Project description:Transcriptomic analysis of the temporal changes induced in mouse bone marrow derived macrophages (BMDMs) by the cytokine Interferon-beta over a timecourse of 0 to 24 hours of treatment. We set out to study the transcriptional events in mouse macrophages over time following stimulation with Interferon-beta. Mouse bone marrow derived macrophages were stimulated for 1, 2, 4, 8 and 24 hours with 10U/mL mouse interferon-beta or left untreated.