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In vitro differentiated Th0, Th17, and Tr1 cells

ABSTRACT: Gene expression profiling of in vitro differentiated murine Th cell subsets. Flow cytometrically sorted naive Th cells (CD4+ CD44- Foxp3-) were polyclonally stimulated in vitro for 3 days using 4 µg/ml plate-bound antibody to CD3 (145-2C11) and 2 µg/ml soluble antibody to CD28 (PV-1). Th0 cells were cultured in the absence of exogenous cytokines. Th17 cells were differentiated with 50 ng/ml IL-6 plus 0.5 ng/ml TGF-β. Tr-1 cells were differentiated with 100 ng/ml IL-27 plus 0.5 ng/ml TGF-β.

ORGANISM(S): Mus musculus

PROVIDER: GSE56021 | GEO | 2014/03/20

SECONDARY ACCESSION(S): PRJNA242233

REPOSITORIES: GEO

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in vitro differentiated Th0, Th17, and Tr1 cells

Project description:Gene expression profiling of in vitro differentiated murine Th cell subsets. Flow cytometrically sorted naive Th cells (CD4+ CD44- Foxp3-) were polyclonally stimulated in vitro for 3 days using 4 Âµg/ml plate-bound antibody to CD3 (145-2C11) and 2 Âµg/ml soluble antibody to CD28 (PV-1). Th0 cells were cultured in the absence of exogenous cytokines. Th17 cells were differentiated with 50 ng/ml IL-6 plus 0.5 ng/ml TGF-Î². Tr-1 cells were differentiated with 100 ng/ml IL-27 plus 0.5 ng/ml TGF-Î². Five biological replicates per Th subset (Th0, Th17, Tr1)

2014-03-20 | E-GEOD-56021 | biostudies-arrayexpress

Notch1 activation and TGF-beta stimulation in transformed human esophageal cells

Project description:Transformed human esophageal keratinocyte cell line EPC2-T (EPC2-hTERT-EGFR-cyclin D1-p53R175H) cells were stimulated with or without 2.5 ng/ml recombinant human TGF-beta1 for 10 days. The above cells were subjected to treatment for 10 days with or without 0.5 µg/ml doxycycline (DOX) to activate tetracycline-inducible (tet-on) ICN1, an active form of Notch1.

2014-12-31 | GSE37994 | GEO

Real-time quantitative PCR analysis of human induced T regulatory cells (Tregs) for metabolic enzymes

Project description:Human naïve CD4 T cells were purified from healthy volunteers' blood and were differentiated into induced Tregs in vitro by culturing for 5 days in the presence of anti-CD3 and CD28 antibodies (2 µg/ml each), IL-2 (100 units/ml) and TGF-β1 (5 ng/ml) for 5 days, then in presence of anti-CD3 and CD28 antibodies for 2 days. Human iTregs were harvested on day 7 post differentiation, then treated with either vehicle or IL-21 (100 ng/ml) in serum-free RPMI media for 18 hours.

2023-04-30 | GSE224033 | GEO

Viral Effect on Metabolism (part I)

Project description:Small airway epithelial cell (SAEC) (Lonza Walkersville, Inc., MD) were grown according to the manufacturer's instructions in growth medium (SAGM) containing 0.03 mg/ml bovine pituitary extract (BPE), 0.5 µg/ml hydrocortisone, 0.5 ng/ml hEGF, 0.5 µg/ml epinephrine, 10 µg/ml transferrin, 5 µg/ml insulin, 0.1 ng/ml retinoic acid, 6.5ng/ml triiodothyronine, 50 µg/ml gentamicin and 50ng/ml Amphotericin-B, and 0.5 mg/ml bovine serum albumin (BSA, fatty acid free). When SAE grew to around 80 to 90% confluence, the cells were put into basal medium not supplemented with growth factors 3hours. Then the cell monolayers were infected with naïve hMPV at multiplicity of infection (MOI) of 3 to certain time. The supernatant was saved and cells were harvested and stored exactly according to the protocol provided by University of Michigan Metabolomics Core Facility for metabolomics analysis. 50mM ammonium acetate in water was used for rinse buffer.

2014-10-24 | ST000278 | MetabolomicsWorkbench

Viral Effect on Metabolism (part II)

2015-02-05 | ST000279 | MetabolomicsWorkbench

Potential cyclooxygenase-2-independent mechanisms of the anti-proliferation and anti-fibrotic effects of celecoxib on the human intrahepatic biliary epithelial cell line HIBEpiC assessed by bulk-RNA sequencing

Project description:This experiment utilized next-generation sequencing (NGS) to identify potential cyclooxygenase-2-independent mechanisms of the anti-proliferation and anti-fibrotic effects of celecoxib on the human intrahepatic biliary epithelial cell line HIBEpiC. HIBEpiC cells were cultured in vitro. Treatments were vehicle, 20 ng/mL of TGF-β (T20), combination of 10μM of celecoxib plus 20 ng/mL of TGF-β (C10+T20), or combination of 10μM of 2,5-dimethyl-celecoxib plus 20 ng/mL of TGF-β (DC10+T20) before NGS. The gene expression were analyzed. Overall, the results suggested that celecoxib had a COX-2-independent inhibitory effect on the proliferation and profibrotic behavior of TGF-β-stimulated CXCL12+ HIBEpiC cells, probably because of the regulation of the cell cycle and several other signaling pathways.

2023-12-31 | GSE220643 | GEO

Extracellular CIRP Induces an Inflammatory Phenotype in Pulmonary Fibroblasts via TLR4

Project description:The wild type (WT) and TLR4 -/- pulmonary fibroblasts were treated with phosphate-buffered saline (PBS), 1 µg/mL recombinant mouse (rm) CIRP, 2 ng/mL rmTGF-ß1 (R&D Systems), or rmCIRP plus rmTGF-ß1. The cell lysates collected at 24 hours were used for high throughput mRNA sequencing.

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Role of areca nut induced TGF-β and epithelial mesenchymal interaction in pathogenesis of precancerous Oral submucous fibrosis

Project description:To understand the role of areca nut and TGF-β induced gene expression changes in fibroblasts and its contibution in the manifestation of Oral submucous fibrosis, we studied gene expression profile in primary human gingival fibroblast (hGF) cells following treatment with areca nut, TGF-β and both together. Control Vs Areca nut 5 µg/ml water extract (5H) (2), Contro Vs TGF-β (2), Control Vs Areca nut (5 µg/ml) and TGF-β (5 ng/ml) (5H+T) (2). (2)- Biological duplicates.

2015-07-06 | E-GEOD-59414 | biostudies-arrayexpress

Untargeted Metabolomics analysis of A549 treated with 0.5 mM extracellular ATP and 10 ng/ml TGF-beta

Project description:Control, 0.5 mM extracellular ATP and 10 ng/ml TGF-beta were used to treated 5 million A549 lung cancer cells in vitro for 2, 6 and 12 hours. The untargeted metabolomics analysis was performed on the cell lysates. The main objective of the study was to determine changes in metabolite abundances in lung cancer after treatment with extracellular ATP and TGF-beta (a known EMT inducer).

2021-04-04 | ST001779 | MetabolomicsWorkbench

Transcriptional profiling of Th1, Th2, Th9, and Th17 T cell clones

Project description:We next sought to identify the transcriptional program that differentiates IL-9+Th2 cells from “conventional” Th2 cells. To this end, we selected representative Th1, Th17, Th2, and IL-9+Th2 clones (Figure 5A) and determined their transcriptome in the resting state and at different time points after activation using RNAseq. Peripheral Blood Mononuclear Cells (PBMC) were isolated by Ficoll-Plaque Plus (GE Healthcare, UK) density gradient centrifugation. Human CD4+ T cells were isolated from PBMC by EasySep positive selection kit (Stemcell Technologies) according to manufacturer’s instruction. Positively selected CD4+ T cells were washed with PBS and stained for subsequent Th cell subset sorting. Memory Th cell subsets were sorted to over 90% purity according to their expression of chemokine receptors from CD45RA-CD25-CD8-CD3+ cells: Th1(CXCR3+CCR8-CCR6-CCR4-), Th2 (CXCR3-CCR8-CCR6-CCR4+), Th17 (CXCR3-CCR8-CCR6+CCR4+), Th9 (CXCR3-CCR8+CCR6-CCR4+). Single cell Th subset clones were directly sorted into 96well plate according to their expression of chemokine receptors (see above). Single cell clones were expanded and maintained by periodic restimulation with PHA (phytohaemaglutinine, 1 µg/ml, Sigma-Chemicals) and irradiated allogenic feeder cells (5x104/well) in culture medium. T cells were polyclonally activated using beads coated with antibodies against CD3, CD2, and CD28 (T cell/bead = 2:1, human T cell activation/expansion Kit, Miltenyi). Cell cultures were sampled before activation (time 0h) and 2, 4, 6, 9, 12, 24, and 48 hours after activation.

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