Project description:Background. Rheumatoid arthritis (RA) is a chronic inflammatory and destructive joint disease, characterized by overexpression of pro-inflammatory/-destructive genes and other activating genes (e.g., proto-oncogenes) in the synovial membrane (SM). The gene expression in disease is often characterized by significant inter-individual variances via specific synchronization/ desynchronization of gene expression. To elucidate the contribution of the variance to the pathogenesis of disease, expression variances were tested in SM samples of RA patients, osteoarthritis (OA) patients, and normal controls (NC). Results. For the comparison between RA and NC, 568 genes with significantly different variances in the 2 groups (p < 0.05; Bonferroni/Holm corrected Brown-Forsythe version of the Levene-Test) were selected. For the comparison between RA and OA, 333 genes were selected. Using the Kyoto encyclopedia of genes and genomes (KEGG), 10 pathways/complexes significantly affected by higher gene expression variances were identified in RA compared to NC, including cytokine â cytokine receptor interactions, the TGF-pathway, and anti-apoptosis. Compared to OA, 3 pathways with significantly higher variances were identified in RA (e.g., B cell receptor signaling, VEGF signaling). Functionally, the majority of the identified pathways is involved in the regulation of inflammation, proliferation, cell survival, and angiogenesis. Conclusion. In RA, a number of disease-relevant or even disease-specific pathways/complexes are characterized by broad intra-group, inter-individual expression variances. This indicates that RA pathogenesis in different individuals may depend to a lesser extent on common alterations of the expression of specific key genes, but on individual-specific alterations of different genes resulting in common disturbances of key pathways. Experiment Overall Design: Expression variances were tested in synovial membrane samples of rheumatoid arthritis patients, osteoarthritis patients, and normal controls (see publication for further details).

Project description:Rheumatoid synoviocytes, which consist of fibroblast-like synoviocytes (FLS) and synovial macrophages (SM), are crucial for the progression of rheumatoid arthritis (RA). Particularly, FLS of RA patients (RA-FLS) exhibit invasive characteristics reminiscent of cancer cells, destroying cartilage and bone, although it remains unresolved how RA-FLS exhibit invasive phenotype. RA-FLS and SM originate differently from mesenchymal and myeloid cells, respectively, but share many pathologic functions. However, the molecular signatures and biological networks representing the distinct and shared features of the two cell types are unknown. Presently, we performed global transcriptome profiling of FLS and SM obtained from RA and osteoarthritis patients. By comparing the transcriptomes, we identified distinct molecular signatures and cellular processes defining invasiveness of RA-FLS and pro-inflammatory properties of RA synovial macrophages (RA-SM), respectively. Interestingly, under interleukin1β-stimulated condition, RA-FLS newly acquired pro-inflammatory signature mimicking RA-SM without losing invasive properties. We next reconstructed a network model that delineates the shared, RA-FLS-dominant (invasive), and RA-SM-dominant (inflammatory) processes. From the network model, we selected 13 genes, including POSTN and TWIST1, as novel regulator candidates responsible for FLS invasiveness. Of note, POSTN and TWIST1 expressions were elevated in independent RA-FLS and were further instigated by interleukin1β. In vitro functional assays demonstrated the requirement of POSTN and TWIST1 for migration and invasion of RA-FLS stimulated with interleukin1β. Taken together, our systems approach to rheumatoid synovitis provides a basis for identifying novel regulators responsible for pathological features of RA-FLS and RA-SM, demonstrating how a certain type of cells acquires functional redundancy under chronic inflammatory conditions. To identify molecular signatures of FLS and MLS in RA joints, we isolated FLS from synovial tissues of RA and osteoarthritis (OA) patients, obtained synovial macrophages from synovial fluid of RA patients, and differentiated control macrophages from peripheral blood of healthy subjects. Also, we stimulated FLS with IL1β, and then analyzed gene expression profiles of both IL1β-stimulated RA-FLS and OA-FLS

Project description:Identification of molecular determinants of the inter-individual and sex variability in the development of nonalcoholic fatty liver disease (NAFLD) is of great importance. Next-generation sequencing (NGS) derived transcriptome profiling (RNA-seq) is a powerful technique to investigate gene transcripts and gene variants for the systems-based analysis of molecular and cellular pathways. The goal of this study was to determine transcriptomic profiles in the livers of male and female Collaborative Cross (CC) mice fed a high-fat and high-sucrose (HF/HS) diet or a control diet for 12 weeks and to identify inter-individual- and sex-specific gene expression differences during the development of NAFLD. Liver mRNA profiles of 10 CC mouse strains (CC003/Unc, CC011/Unc, CC013/GeniUnc, CC019/TauUnc, CC032/GeniUnc, CC040/TauUnc, CC041/TauUnc, CC042/GeniUnc, CC043/GeniUnc, and CC060/Unc) fed an HF/HS diet were generated by deep sequencing, in triplicate, using Illumina NextSeq 500 NGS system. The sequence reads that passed quality filters were analyzed at the transcript level using TopHat followed by Cufflinks. The total number of differentially expressed genes identified by RNA-seq varied between sexes and individual mouse strains and correlated with the severity of liver steatosis. Pathway analysis of differentially expressed genes showed dysregulation of genes involved in lipid metabolism, cell proliferation and death, inflammation, and fibrogenesis pathways in the livers of males and females CC mice. The RNA-seq data were validated and confirmed by qRT-PCR analysis of 27 selected genes associated with lipid metabolism and fibrogenesis using TaqMan assays. Our results show that RNA-seq based transcriptome analysis offers a comprehensive, quantitative, and qualitative evaluation of whole-genome expression that can be used for the identification of unique genes associated with the inter-individual and sex differences in susceptibility to NAFLD at the population level.

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Homo sapiens

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