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Title: A validated gene regulatory network and GWAS identifies early regulators of T-cell associated diseases (Th17 Treg)      
dateReleased:
11-12-2015
description:
In this work we present an analytical strategy to systematically identify early regulators by combining gene regulatory networks (GRN) with GWAS. We hypothesized that early regulators in T-cell associated diseases could be found by defining upstream transcription factors (TFs) in T-cell differentiation. Time series expression and DNA methylation profiling of T-cell differentiation identified several upstream TFs, of which TFs involved in Th1/2 differentiation were most enriched for disease associated SNPs identified by GWAS. Naïve CD4+ T cells were isolated from buffy coat of four healthy donors using a naïve CD4+ T cell isolation kit (Miltenyi, Bergisch-Gladbach, Germany). Naïve CD4+ T cells were stimulated with TGF-β (1 ng/mL), IL-1β (10 ng/mL), IL-6 (25 ng/mL), IL-21 (25 ng/mL) and IL-23 (25 ng/mL) for Th17, and TGF-β (10 ng/mL) and IL-2 (10 ng/mL) for Treg. Cells were cultured for six days in Iscove’s modified Dulbecco medium (IMDM) supplemented with 2 mM L-glutamine (PAA Laboratories, Linz, Austria), 10% heat-inactivated FCS (PAA Laboratories, Linz, Austria), 5 µM β–mercaptoethanol (Sigma-Aldrich, St. Louis, Missouri, USA) and 50 ug/mL gentamicin (Sigma-Aldrich, St. Louis, Missouri, USA). Cells were cultured for 6 days and then re-stimulated with plate-bound anti-CD3 and soluble anti-CD28 in the presence of corresponding polarizing cytokines and antibodies for another 2 days (Zhang et al. 2013). RNA was extracted using a miRneasy Mini Kit (Qiagen). The RNA concentrations were analysed with NanoDrop ND-1000 Spectrophotometer (NanoDrop Technologies). For the gene expression microarray analysis, The cRNA was prepared using a Low Input QuickAmp Labeling Kit. For Th17 and Treg cells the gene expression microarray analysis was performed using SurePrint G3 Human Gene Expression 8x60K v2 microarray kit, according to the manufacture’s instruction (Agilent Technologies).
privacy:
not applicable
aggregation:
instance of dataset
ID:
E-GEOD-60679
refinement:
raw
alternateIdentifiers:
60679
keywords:
functional genomics
dateModified:
11-24-2015
availability:
available
types:
gene expression
name:
Homo sapiens
ID:
A-GEOD-17077
name:
Agilent-039494 SurePrint G3 Human GE v2 8x60K Microarray 039381 (Probe Name version)
accessURL: https://www.ebi.ac.uk/arrayexpress/files/E-GEOD-60679/E-GEOD-60679.raw.1.zip
storedIn:
ArrayExpress
qualifier:
gzip compressed
format:
TXT
accessType:
download
authentication:
none
authorization:
none
accessURL: https://www.ebi.ac.uk/arrayexpress/files/E-GEOD-60679/E-GEOD-60679.processed.1.zip
storedIn:
ArrayExpress
qualifier:
gzip compressed
format:
TXT
accessType:
download
authentication:
none
authorization:
none
accessURL: https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE60679
storedIn:
Gene Expression Omnibus
qualifier:
not compressed
format:
HTML
accessType:
landing page
primary:
true
authentication:
none
authorization:
none
abbreviation:
EBI
homePage: http://www.ebi.ac.uk/
ID:
SCR:004727
name:
European Bioinformatics Institute
homePage: https://www.ebi.ac.uk/arrayexpress/
ID:
SCR:002964
name:
ArrayExpress

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