ABSTRACT: This experiment tested whether pretreatment of human microvascular endothelial cells (HMECs) with interferon-gamma (IFNγ) for 2h affected the transcriptional response to 1h exposure to interleukin-6 (IL-6). Overall design: Human microvascular endothelial cells in culture; exposure to IFNγ 1 ng/ml for 3 h alone, with IFNγ 1 ng/ml for 2 h and then with IL6 50 U/ml for 1 h, with IL6 50 U/ml alone, or with neither. 4 replicates for each condition. RNA isolation, Illumina expression arrays. The supplementary file 'GSE19082_non-normalized_data.txt' contains non-normalized data for Samples GSM472434-GSM472449.
Project description:Here, we investigated the time-course changes in the pattern of microRNA (miRNA) expression of TNFα and IFNγ-stimulated and unstimulated hCMEC/D3 cells, an immortalized human cerebral microvascular endothelial cell line. In order to investigate pro-inflammatory cytokine-induced changes in miRNA levels in hCMEC/D3 cells, we challenged brain endothelial cells with TNFα and IFNγ (100 ng/ml) for 2 h, 6 h and 24 h and determined microRNA expression in cytokine-stimulated and unstimulated cells
Project description:Human monocyte THP-1 cells obtained from ATCC were cultured in RPMI 1640 (Invitrogen, Carlsbad, CA) containing 10% FBS and supplemented with 10 mM Hepes (Gibco BRL). THP-1 was differentiated into macrophages by 24-h incubation with 160 nM phorbol 12-myristate 13-acetate (PMA; Sigma, St. Louis, MO) followed by 24-h incubation in RPMI medium. Macrophages were further polarized to M1 macrophages by incubation with 10 pg/ml of lipopolysaccharide (LPS; Sigma) and 20 ng/ml of interferon (IFN)-γ (R&D Systems, MN) and are referred to as M(LPS+IFN-γ) cells. M2 macrophages were obtained by incubation with 20 ng/ml of interleukin (IL)-4 (R&D Systems) and are referred to as M(IL4) cells. To test the represented polarization marker of PMA differentiated-THP-1 macrophages stimulated with 20 ng ml(-1) IFNγ + 10 pg ml(-1) LPS and 20 ng ml(-1) IL-4, which are known to influence macrophage polarization in vetro into the M1 and M2 state, respectively. We used microarrays to detail the gene expression pools to identify distinct M1 and M2 state during this process. Overall design: To test the polarization of macrophage. PMA differentiated-THP-1 macrophages were stimulated with IFNγ+LPS and IL-4. THP-1 derived macrophages, including M(LPS+INFγ) and M(IL4) cells, were processed in Trizol and then analyzed using the Affymetrix U133 Array platform.
Project description:While it is clear that T cell derived IFNγ has to act on tumor stroma cells for rejection of solid tumors, it is not clear which tumor stroma cells are targets. We studied how IFNγ affects gene expression in tumor blood vessels in vivo. To study the effect on endothelial cells, we either used a model of ectopic IFNγ (MCA313 tumors) or IFNγ-GFP fusion protein (J558L tumors) expression in tumors, or we used T cell derived IFNγ in large vascularized 16.113 tumours. Tumors were grown in mice that were expressing the IFNγ receptor ubiquitously (J558L tumors + IFNγ-GFP treatment and 16.113 tumors + T cell treatment) or in some experiments the IFNγ-receptor was expressed exclusively in endothelial cells (MCA313 tumor + IFNγ treatment). We compared RNA expression profiles from mouse endothelial cells exposed to IFNγ (IFNγ-GFP) or not exposed to IFNγ (IFNγ-GFP). Endothlial cells were drived from tumors (or in one experimantal situation -J558L- as a control also from kidney) Overall design: Tumors were grown in mice (either with ubiquitous IFNγ-receptor expression -WT mice- or in transgenic mice with endothelial cell specific IFNγ-receptor expression - PIG x Pdgfb-Cre x IFNγ-del mice) to ca. the size of ca. 1.0 cm in diameter. Then tumors were exposed for 24h or 48h to IFNγ (MCA313-IFNγ-IND tumors) or IFNγ-GFP (J558L-IFNγ-GFP-IND tumors) by ectopic expression from the tumor cells. As controls we used endothelial cells from tumors which were not induced to express IFNγ(0h). To confirm that T cell derived IFNγ acts similar as in our ectopic models, we used T cells for T cell therapy of 16.113 tumors that could express IFNγ (IFNγ+/+ T cells) or could not express IFNγ (IFNγ-/- T cells).
Project description:BMP9 signaling has been implicated in hereditary hemorrhagic telangiectasia and vascular remodeling, acting via the HHT target genes, endoglin and ALK1. This study sought to identify endothelial BMP9-regulated proteins that could affect the HHT phenotype. Gene ontology analysis of cDNA microarray data obtained following BMP9 treatment of primary human endothelial cells indicated regulation of chemokine, adhesion, and inflammation pathways. The sample set is comprised of three biological replicate control human dermal microvascular endothelial cells, and three treated (5 ng/ml human recombinant BMP9) biological replicate human dermal microvascular endothelial cells
Project description:Myeloid-derived suppressor cells (MDSCs) have emerged as major regulators of immune responses in cancer and other pathological conditions. Multiple factors including cytokines, transcription factors and multiple signaling pathways are involved in MDSC differentiation. Cytokines such as granulocyte-macrophage colony-stimulating factor (GM-CSF) and interleukin(IL-6) etc could in vitro mediate development of MDSCs.IL-6 with GM-CSF mediated MDSC not only had stronger suppressive function but also the dynamics of their suppressive function was different from GM-CSF alone mediated MDSCs.To found a new regulatory factor (s) in tumor and inflammatory environments, we compared GM-CSF and IL-6 mediated MDSCs with GM-CSF alone mediated MDSCs using lncRNA microarray, miRNA microarrays and protein-coding mRNA microarrays. Overall design: Inflammatory factors associated MDSCs were generated by cultivating bone marrow cells of C57BL/6 mice in petri dishes for 4 days in the presence of 5% FBS medium containing GM-CSF (40 ng/ml) only, GM-CSF (40 ng/ml) plus IL-6 (40 ng/ml) using different processing group for each experiment.
Project description:Myeloid-derived suppressor cells (MDSCs) have emerged as major regulators of immune responses in cancer and other pathological conditions. Multiple factors including cytokines, transcription factors and multiple signaling pathways are involved in MDSC differentiation. Cytokines such as granulocyte-macrophage colony-stimulating factor (GM-CSF) and interleukin(IL-6) etc could in vitro mediate development of MDSCs.IL-6 with GM-CSF mediated MDSC not only had stronger suppressive function but also the dynamics of their suppressive function was different from GM-CSF alone mediated MDSCs.To found a new regulatory factor (s) in tumor and inflammatory environments, we compared GM-CSF and IL-6 mediated MDSCs with GM-CSF alone mediated MDSCs using lncRNA microarray and protein-coding mRNA microarrays. Overall design: Inflammatory factors associated MDSCs were generated by cultivating bone marrow cells of C57BL/6 mice in petri dishes for 4 days in the presence of 5% FBS medium containing GM-CSF (40 ng/ml) only, GM-CSF (40 ng/ml) plus IL-6 (40 ng/ml) using different processing group for each experiment.
Project description:To unravel the underlying mechanisms of inflammation-related idiosyncratic drug toxicity Overall design: HepG2 cells were exposed to 3 idiosyncratic and 3 non-idiosyncratic compounds with or without the cytokine mix (TNF 100 ng/ml + IFNγ 100 ng/ml + IL-1α 20 ng/ml + IL-6 20 ng/ml + LPS 10 µg/ml)as described previously (Cosgrove et al., 2009) for 6, 12 or 24 h.
Project description:To identify differentially expressed genes in mouse IL6 and Myc double TG plasmacytoma, total RNAs from mouse plasmacytoma generated from IL6 + Myc TG mice were applied to NIAID mouse gene expression arrays. The transcriptome in IL6+ Myc TG mouse PCT was compared to transcriptome in IL6 TG mouse PCT Overall design: Eleven set of IL6+Myc TG PCTs and ten set of IL6 TG PCTs were used. Total RNAs from each mouse labelled with Cy3, and reference RNAs labeled with Cy5 were hybridzed to spotted oligomicroarray.
Project description:Episodic Ebola virus (EBOV) outbreaks, such as the current one in West Africa, emphasize the critical need for novel antivirals against this highly pathogenic virus. Here, we demonstrate that interferon gamma (IFNγ) prevents morbidity and mortality associated with EBOV infection when administered to mice either 24 hours prior to or 2 hours following EBOV infection. Microarray studies with IFNγ-stimulated human macrophages identified novel interferon-stimulated genes (ISGs) that inhibit EBOV infection upon ectopic expression. IFNγ treatment reduced viral RNA levels in macrophages to a similar degree as cells treated with the protein synthesis inhibitor, cycloheximide, suggesting that IFNγ treatment inhibits genome replication. As IFNγ treatment robustly protects mice against EBOV infection, we propose that this FDA-approved drug may serve as a useful prophylactic or therapeutic strategy during EBOV outbreaks, contributing to the currently limited arsenal of filovirus antivirals. Overall design: Total RNA was isolated from alveolar macrophages from healthy individuals. All cells were treated with 50 ng/mL of M-CSF plus or minus 20 ng/mL of IFNg for 24 hours prior to infection.