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Expression profiling in RA disease pre and post anti-TNF treatment

ABSTRACT: The purpose of this study was (1) to identify novel genes involved in the pathogenesis of Rheumatoid Arthritis (RA) disease, which may provide additional targets for therapeutic intervention and (2) to examine the molecular mechanisms associated with the response to anti-TNF treatment. Microarray analysis of LPS-stimulated whole blood from RA patients pre and post anti-TNF treatment was conducted. This study identified 818 transcripts, differentially expressed in RA patients pre-treatment compared to non-RA control samples. While a number of these genes were associated with RA in previous studies, validating our data, a number of novel genes with possible functions in RA disease were also identified. The number of transcripts (1051) significantly altered post anti-TNF treatment indicates the impact of anti-TNF therapy on systemic gene expression. A number of these transcripts were confirmed to be altered in a larger patient group and may represent potential genetic markers of a patient’s clinical response to anti-TNF treatment. Keywords: Expression profiling in RA disease pre and post anti-TNF treatment

ORGANISM(S): Homo sapiens

PROVIDER: GSE7524 | GEO | 2007/07/07

SECONDARY ACCESSION(S): PRJNA99389

REPOSITORIES: GEO

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Project description:The purpose of this study was (1) to identify novel genes involved in the pathogenesis of Rheumatoid Arthritis (RA) disease, which may provide additional targets for therapeutic intervention and (2) to examine the molecular mechanisms associated with the response to anti-TNF treatment. Microarray analysis of LPS-stimulated whole blood from RA patients pre and post anti-TNF treatment was conducted. This study identified 818 transcripts, differentially expressed in RA patients pre-treatment compared to non-RA control samples. While a number of these genes were associated with RA in previous studies, validating our data, a number of novel genes with possible functions in RA disease were also identified. The number of transcripts (1051) significantly altered post anti-TNF treatment indicates the impact of anti-TNF therapy on systemic gene expression. A number of these transcripts were confirmed to be altered in a larger patient group and may represent potential genetic markers of a patientâs clinical response to anti-TNF treatment. Experiment Overall Design: Transcriptome profiling was performed on two RA patients, pre- and three months post-Enbrel therapy and two matched non-RA controls. The whole blood model described by Wang et al (2000) was used with minor modifications. Briefly, to EDTA anti-coagulated blood, LPS (30ng/ml) was added immediately after drawing, and incubated at 37oC for 4 hours, mixing at 80s. After 4 hours red blood cells were lysed and pelleted, white cells were washed once in PBS (Gibco) and frozen at â70oC for subsequent RNA extraction. Total RNA was extracted using the RNeasy kit (QIAGEN) as per manufacturerâs instructions. Samples were treated with DNA-Free (Ambion) to remove genomic DNA and checked using intron-overlapping primers prior to generating cRNA probes or cDNA (data not shown). Affymetrix array data was normalised (globally) and statistically (t-test) analysed using GeneSight software (Biodiscovery). Gene expression changes with a p value of < 0.05 and a fold change of 3 or greater were considered differentially expressed.

Project description:Objectives: Pinolenic acid (PNLA), an omega-6 polyunsaturated fatty acid from pine nuts, has anti-inflammatory and anti-atherogenic effects. We aimed to investigate the direct anti-inflammatory effect and anti-atherogenic effects of PNLA on activated purified CD14 monocytes from peripheral blood of patients with rheumatoid arthritis (RA) in vitro. Methods: Flow cytometry was used to assess the proportions of CD14 monocytes expressing TNF-α, IL-6, IL-1β, and IL-8 in purified monocytes from patients with RA after lipopolysaccharide (LPS) stimulation with/without PNLA pre-treatment. The whole genomic transcriptome (WGT) profile of PNLA-treated, and LPS-activated monocytes from patients with active RA was investigated by RNA-sequencing. Results: PNLA reduced percentage of monocytes expressing cytokines: TNF-a by 23% (p=0.048), IL-6 by 25% (p=0.011), IL-1B by 23% (p=0.050), IL-8 by 20% (p=0.066). Pathway analysis identified upstream activation of peroxisomes proliferator-activated receptors (PPARs), sirtuin3, and let7miRNA, KLF15 which are anti-inflammatory and antioxidative. In contrast, DAP3, LIF and STAT3, which are involved in TNF-a, and IL-6 signal transduction, were inhibited. Canonical Pathway analysis showed that PNLA inhibited oxidative phosphorylation (p=9.14E-09) and mitochondrial dysfunction (p=4.18E-08), while the sirtuin (SIRTs) signalling pathway was activated (p=8.89E-06) which interfere with the pathophysiologic process of atherosclerosis. Many miRNAs were modulated by PNLA suggesting potential post-transcriptional regulation of metabolic and immune response that has not been described previously. Multiple miRNAs target pyruvate dehydrogenase kinase-4 (PDK4), single-immunoglobulin interleukin-1 receptor molecule (SIGIRR), mitochondrially encoded ATP synthase membrane subunit 6 (MT-ATP6) and Acetyl-CoA Acyltranferase2 (ACAA2); genes implicated in regulation of lipid and cell metabolism, inflammation, and mitochondrial dysfunction. Conclusion: PNLA has potential anti-atherogenic and immune-metabolic effects on monocytes that are pathogenic in RA and atherosclerosis. Dietary PNLA supplementation regulates key miRNAs that are involved in metabolic, mitochondrial, and inflammatory pathways.

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Expression profiling in RA disease pre and post anti-TNF treatment

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