Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

Expression Array of Lymphoblastoid Cell lines Under Inflammatory Conditions

ABSTRACT: Comparison of gene expression across 85 cell lines after stimulation with a cocktail of inflammatory cytokines Total RNA extracted from Genotyped Lymphoblastoid cell lines under inflammatory and baseline conditions. 3 biological replicates of stimulated and 2 biological replicates of baseline samples were hybridized to the Illumina Human WG-6 v.3 gene expression chip.

ORGANISM(S): Homo sapiens

SUBMITTER: Simone Thair

PROVIDER: E-GEOD-25543 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

A single nucleotide polymorphism in NF-κB inducing kinase is associated with mortality in septic shock.

Thair Simone A SA Walley Keith R KR Nakada Taka-aki TA McConechy Melissa K MK Boyd John H JH Wellman Hugh H Russell James A JA

Journal of immunology (Baltimore, Md. : 1950) 20110121 4

We tested the hypothesis that single nucleotide polymorphisms (SNPs) within genes of the NF-κB pathway are associated with altered clinical outcome of septic shock patients. We genotyped 59 SNPs in the NF-κB pathway in a discovery cohort of septic shock patients (St. Paul's Hospital [SPH], N = 589), which identified the C allele of rs7222094 T/C within MAP3K14 (NF-κB inducing kinase; NIK) associated with increased 28-d mortality (uncorrected p = 0.00024, Bonferroni corrected p = 0.014). This res ...[more]

PMID: 21257964

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Project description:Epstein-Barr virus (EBV) transformed lymphoblastoid cell lines (LCLs) are a widely used renewable resource for functional genomic studies in humans. The ability to accumulate multidimensional data pertaining to the same individual cell lines, from complete genomic sequences to detailed gene regulatory profiles, further enhances the utility of LCLs as a model system. However, the extent to which LCLs are a faithful model system is relatively unknown. We have previously shown that gene expression profiles of newly established LCLs maintain a strong individual component. Here, we extend our study to investigate the effect of freeze-thaw cycles on gene expression patterns in mature LCLs, especially in the context of inter-individual variation in gene regulation. We found a profound difference in the gene expression profiles of newly established and mature LCLs. Once newly established LCLs undergo a freeze-thaw cycle, the individual specific gene expression signatures become much less pronounced as the gene regulatory programs in LCLs from different individuals converge to a more uniform profile, which reflects a mature transformed B cell phenotype. As expected, previously identified eQTLs are enriched among the relatively few genes whose regulations in mature LCLs maintain marked individual signatures. We thus conclude that findings and insight drawn from gene regulatory studies in mature LCLs are generally not affected by artificial nature of the LCL model system and are likely to faithfully reflect regulatory interactions in primary tissues. However, our data indicate that many aspects of primary B cell biology cannot be observed and studied in mature LCL cultures. We obtained unpurified buffy coats (of a unit of blood) from six unrelated healthy individuals of Caucasian ethnicity (age range: 20-45). We isolated CD20+ B cells and established six independent cultures of LCLs between October 2009 and January 2010. From each of these samples, we obtained genome wide gene expression data using Illumina HumanHT-12 v3 Expression BeadChip arrays. We refer to data from these experiments as M-bM-^@M-^\cycle 0M-bM-^@M-^] to acknowledge the fact that the LCLs from this cycle were newly established and therefore not frozen/thawed. Between February 2011 and October 2012, we thawed each of these LCL cultures every 3 months and cultured them until obtaining ~10 million cells (February 2011=cycle 1, June 2011=cycle 2, October 2011=cycle 3, February 2012=cycle 4, June 2012=cycle 5, and October 2012=cycle 6). We used Illumina HumanHT-12 v4 Expression BeadChip arrays and obtained genome wide gene expression data on cycle 2, cycle 4 and cycle 6 LCLs. At each hybridization batch we included a subset of RNA samples that were also hybridized in a previous batch. This allowed us to effectively consider the batch effect on gene expression profiles. Our final dataset included genome wide gene expression data from 187 samples collected at 4 different time points.

Project description:We used microarrays to identify the variation of basal gene expression level among 287 lymphoblastoid cell lines. Radiation therapy is used to treat half of all cancer patients. Response to radiation therapy varies widely among patients. Therefore, we performed a genome-wide association study (GWAS) to identify biomarkers to help predict radiation response using 277 ethnically defined human lymphoblastoid cell lines (LCLs). Basal gene expression levels and 1.3 million genome-wide SNP markers from both Affymetrix and Illumina platforms were assayed for all 277 human LCLs. MTS (3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium) assays for radiation cytotoxicity were also performed to obtain area under the curve (AUC) as a radiation response phenotype for use in the association studies. Functional validation of candidate genes, selected from an integrated analysis that used SNP, expression and AUC data, was performed with multiple cancer cell lines using specific siRNA knockdown, followed by MTS and colony-forming assays. 27 loci, each containing at least 2 SNPs within 50kb with p-values <10-4, were associated with radiation AUC. 270 expression probe sets were associated with radiation AUC with p<10-3. The integrated analysis identified 50 SNPs in 14 of the 27 loci that were associated with both AUC and the expression of 39 genes that were also associated with radiation AUC (p<10-3). Functional validation using siRNA knockdown in multiple tumor cell lines showed that C13orf34, MAD2L1, PLK4, TPD52 and DEPDC1B each significantly altered radiation sensitivity in at least 2 cancer cell lines. Studies performed with LCLs can help to identify novel biomarkers that might contribute to variation in response to radiation therapy and enhance our understanding of mechanisms underlying that variation. EBV-transformed LCLs from 95 African-American (AA), 96 Caucasian-American (CA), and 96 Han Chinese-American (HCA) unrelated healthy subjects (sample sets HD100AA, HD100CAU, HD100CHI) were purchased from the Coriell Cell Repository (Camden, NJ).

Dataset Information

Expression Array of Lymphoblastoid Cell lines Under Inflammatory Conditions

Publications

A single nucleotide polymorphism in NF-κB inducing kinase is associated with mortality in septic shock.

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