Project description:In 15 patients with rheumatoid arthritis the baseline expression (t=0) of type I IFN-regulated genes was determined in peripheral blood cells (PAXgene) using cDNA-microarrays and compared to levels one month after (t=1) anti-TNF treatment (infliximab). Therefore, we used 43K cDNA microarrays from the Stanford Functional Genomics Facility (http://microarray.org/sfgf/) printed on aminosilane-coated slides containing ~20.000 unique genes. Only one batch of arrays was used for all experiments. First DNA spots were UV-cross linked to the slide using 150-300 mJoules. Sample preparation and microarray hybridization were performed as described previously (1,2). Data storage and filtering was performed using the Stanford Microarray Database (http://genome-www5.stanford.edu//) as described previously. 1. van der Pouw Kraan TC, Wijbrandts CA, van Baarsen LG, Voskuyl AE, Rustenburg F, Baggen JM et al.: Rheumatoid arthritis subtypes identified by genomic profiling of peripheral blood cells: assignment of a type I interferon signature in a subpopulation of patients. Ann Rheum Dis 2007, 66: 1008-1014. 2. van der Pouw Kraan TC, van Baarsen LG, Rustenburg F, Baltus B, Fero M, Verweij CL: Gene expression profiling in rheumatology. Methods Mol Med 2007, 136: 305-327. Abbreviations: - Bnrs are the different patients analyzed - PAX or Paxgene indicates the total RNA was isolated from peripheral blood obtained in PAXgene tubes - CRS: common reference sample: We made a common reference that consisted of a mixture of mRNAs isolated from 11 different cell lines (Perou CM, Jeffrey SS, van de RM et al. Distinctive gene expression patterns in human mammary epithelial cells and breast cancers. Proc Natl Acad Sci U S A. 1999; 96:9212-9217) supplemented with RNA from rheumatoid synovial tissue, fibroblasts, and activated peripheral blood mononuclear cells (PBMCs). - PMT: photomultiplier tube: the sensitivity/intensity of the scanner can be adjusted to prevent spot saturation. Compound Based Treatment: anti-TNF (infliximab)

Project description:BackgroundThe pathogenesis of rheumatoid arthritis (RA) is complex. This study aimed to identify diagnostic biomarkers and transcriptional regulators that underlie RA based on bioinformatics analysis and experimental verification.Material and methodsWe applied weighted gene co-expression network analysis (WGCNA) to analyze dataset GSE55457 and obtained the key module most relevant to the RA phenotype. We then conducted gene function annotation, gene set enrichment analysis (GSEA) and immunocytes quantitative analysis (CIBERSORT). Moreover, the intersection of differentially expressed genes (DEGs) and genes within the key module were entered into the STRING database to construct an interaction network and to mine hub genes. We predicted microRNA (miRNA) using a web-based tool (miRDB). Finally, hub genes and vital miRNAs were validated with independent GEO datasets, RT-qPCR and Western blot.ResultsA total of 367 DEGs were characterized by differential expression analysis. The WGCNA method divided genes into 14 modules, and we focused on the turquoise module containing 845 genes. Gene function annotation and GSEA suggested that immune response and inflammatory signaling pathways are the molecular mechanisms behind RA. Nine hub genes were screened from the network and seven vital regulators were obtained using miRNA prediction. CIBERSORT analysis identified five cell types enriched in RA samples, which were closely related to the expression of hub genes. Through ROC curve and RT-qPCR validation, we confirmed five genes that were specific for RA, including CCL25, CXCL9, CXCL10, CXCL11, and CXCL13. Moreover, we selected a representative gene (CXCL10) for Western blot validation. Vital miRNAs verification showed that only the differences in has-miR-573 and has-miR-34a were statistically significant.ConclusionOur study reveals diagnostic genes and vital microRNAs highly related to RA, which could help improve our understanding of the molecular mechanisms underlying the disorder and provide theoretical support for the future exploration of innovative therapeutic approaches.

Project description:In 15 patients with rheumatoid arthritis the baseline expression (t=0) of type I IFN-regulated genes was determined in peripheral blood cells (PAXgene) using cDNA-microarrays and compared to levels one month after (t=1) anti-TNF treatment (infliximab). Therefore, we used 43K cDNA microarrays from the Stanford Functional Genomics Facility (http://microarray.org/sfgf/) printed on aminosilane-coated slides containing ~20.000 unique genes. Only one batch of arrays was used for all experiments. First DNA spots were UV-cross linked to the slide using 150-300 mJoules. Sample preparation and microarray hybridization were performed as described previously (1,2). Data storage and filtering was performed using the Stanford Microarray Database (http://genome-www5.stanford.edu//) as described previously. 1. van der Pouw Kraan TC, Wijbrandts CA, van Baarsen LG, Voskuyl AE, Rustenburg F, Baggen JM et al.: Rheumatoid arthritis subtypes identified by genomic profiling of peripheral blood cells: assignment of a type I interferon signature in a subpopulation of patients. Ann Rheum Dis 2007, 66: 1008-1014. 2. van der Pouw Kraan TC, van Baarsen LG, Rustenburg F, Baltus B, Fero M, Verweij CL: Gene expression profiling in rheumatology. Methods Mol Med 2007, 136: 305-327. Abbreviations: - Bnrs are the different patients analyzed - PAX or Paxgene indicates the total RNA was isolated from peripheral blood obtained in PAXgene tubes - CRS: common reference sample: We made a common reference that consisted of a mixture of mRNAs isolated from 11 different cell lines (Perou CM, Jeffrey SS, van de RM et al. Distinctive gene expression patterns in human mammary epithelial cells and breast cancers. Proc Natl Acad Sci U S A. 1999; 96:9212-9217) supplemented with RNA from rheumatoid synovial tissue, fibroblasts, and activated peripheral blood mononuclear cells (PBMCs). - PMT: photomultiplier tube: the sensitivity/intensity of the scanner can be adjusted to prevent spot saturation. Compound Based Treatment: anti-TNF (infliximab) reference_design

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Repurposing haloperidol for the treatment of rheumatoid arthritis: an integrative approach using data mining techniques.

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