Metabolomics,Unknown,Transcriptomics,Genomics,Proteomics

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Expression data measured by microarray of CD4+ T cells from healthy individuals stimulated with anti-CD3/CD28 with or without IFNb or Th17 polarizing cytokines

ABSTRACT: Variation in individuals' adaptive immune response is believed to influence susceptibility to complex diseases in humans. The genetic basis of such variation is poorly understood. We measured gene expression from resting and activated CD4+ T cells derived from the peripheral blood of healthy individuals. We activated the primary T cells with anti-CD3/CD28 beads. We collected peripheral blood from each human donor. We isolated peripheral blood mononuclear cells by Ficoll, and negatively selected for CD4+ T cells using RosettaSep. We then either left cells unstimulated or stimulated them with beads conjugated with anti-CD3 and anti-CD28. Cells from 15 individuals were harvested at up to 3 time points (0hr, 4hr or 48hr), lysed and RNA isolated to be profiled on microarray.

ORGANISM(S): Homo sapiens

SUBMITTER: Richard Cruse

PROVIDER: E-GEOD-60235 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:Early diagnosis of transthyretin (TTR) amyloid diseases remains challenging because of variable disease penetrance. Currently, patients must have an amyloid positive tissue biopsy to be eligible for disease modifying therapies. Early diagnosis is often difficult because the patient exhibits apparent symptoms of polyneuropathy or cardiomyopathy, but has a negative amyloid biopsy. Thus, there is a pressing need for more objective, quantitative diagnostics and biomarkers of TTR-aggregation-associated polyneuropathy and cardiomyopathy. This is especially true in the context of clinical trials demonstrating significant disease modifying effects, e.g. when the TTR tetramer stabilizer tafamidis was administered to familial amyloid polyneuropathy (FAP) patients early in the disease course. When asked if the findings of the tafamidis registration trial were âsufficiently robust to provide substantial evidence of efficacy for a surrogate endpoint that is reasonably likely to predict a clinical benefitâ the advisory committee said yes, but the FDA rejected the tetramer stabilization surrogate biomarker required for orphan tafamidis approvalâhence, acceptable biomarkers are badly needed. Herein, we explored whether peripheral blood cell mRNA expression profiles could differentiate symptomatic from asymptomatic V30M FAP patients, and if such a profile would normalize upon tafamidis treatment. We demonstrate that blood cell gene expression patterns reveal sex-independent as well as male and female specific inflammatory signatures in symptomatic FAP patients, but not in asymptomatic carriers, that normalize in FAP patients 6 months after tafamidis treatment. Thus these signatures have potential both as an early diagnostic and as a surrogate biomarker for measuring response to treatment in FAP patients. Study Subjects: Tafamidis-treated and untreated V30M FAP patients, asymptomatic V30M carriers, and healthy, age- and sex-matched controls without TTR mutations had whole blood drawn. RNA was extracted from a total of 309 peripheral blood samples collected in PAXgene tubes (Qiagen). RNA Extraction and Microarray Processing: RNA was extracted with the PAXgene RNA Whole Blood Kit. Each RNA sample (100ng) was amplified and labeled with the Ambion WT Express Kit and hybridized to Affymetrix HuGene 1.1 ST Arrays using the Affymetrix GeneTitan Multi-Channel (MC) platform. Normalized signals were generated using Robust Multichip Average (RMA) in Partek Genomics Suite software version 6.6. Probesets on the DNA microarrays with low expressed signals were determined using a kernel density plot, and signals <Log2 of 6 were excluded from analysis.

Dataset Information

Expression data measured by microarray of CD4+ T cells from healthy individuals stimulated with anti-CD3/CD28 with or without IFNb or Th17 polarizing cytokines

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