Gene expression patterns in IgG4-RD before and after steroid treatment and healthy control groups
ABSTRACT: Transcriptome analysis in patients with IgG4-RD and healthy controls IgG4-related disease (IgG4-RD) is a new emerging disease entity characterized by elevated serum IgG4 concentrations and tissue tumefaction or infiltration by IgG4-positive plasma cells. At present, however, it is not clear whether IgG4-RD is caused by abnormalities in acquired immunity, like autoimmune diseases, or whether the excess production of IgG4 is a true cause of IgG4-RD or an epiphenomenon associated with inflammatory and/or allergic reactions. We therefore attempted to identify genes related to the pathogenesis or clinicopathology of IgG4-RD. Peripheral blood mononuclear cells (PBMCs) were obtained from patients with IgG4-RD before and after steroid therapy and from healthy controls. Total RNA was extracted and DNA microarray analysis was performed to screen for genes showing changes in expression. To exclude any gender-related differences in gene expression, DNA microarray analysis was performed only on samples obtained from male patients and controls. Total RNA was reverse transcribed to cDNA using Ambion® WT Expression kits (Applied Biosystems, Foster City, CA), labeled with GeneChip® WT Terminal Labeling and Controls kits (Affymetrix, Santa Clara, CA), and hybridized to GeneChip® Human Gene 1.0 ST Arrays (Affymetrix), which include 28869 probes. Digitalized image data were processed using GeneChip® Operating Software (Affymetrix).
Project description:RD induced DNA damage and suppressed DNA repair in PC-3 cells. Expression profiling of PC-3 cells treated with 10 umol/L RD for 24 hours. Total RNA was extracted from PC-3 cells exposed to RD for 24 h using an RNAiso plus kit (TaKaRa). Microarray analysis was performed on separate samples on the Affymetrix HG-U133 plus 2 chip.
Project description:RD induced DNA damage and suppressed DNA repair in PC-3 cells. Expression profiling of PC-3 cells treated with 10 umol/L RD for 24 hours. Overall design: Total RNA was extracted from PC-3 cells exposed to RD for 24 h using an RNAiso plus kit (TaKaRa). Microarray analysis was performed on separate samples on the Affymetrix HG-U133 plus 2 chip.
Project description:Bacterial outer membrane vesicles (OMVs) have important biological roles in pathogenesis and intercellular interactions, but a general mechanism of OMV formation is lacking. Here we show that the VacJ/Yrb ABC transport system, a proposed phospholipid (PL) transporter, is involved in OMV formation. Deletion or repression of VacJ/Yrb increases OMV production in two distantly related Gram-negative bacteria, Haemophilus influenzae and Vibrio cholerae. Within our studies we also analyzed the proteome of the OMV and outer membrane (OM) and found no massive alteration in Haemophilus influenzae Rd KW20, Rd ∆yrbE and Rd ∆vacJ. Lipidome analyses demonstrate that OMVs from VacJ/Yrb-defective mutants in H. influenzae are enriched in PLs and certain fatty acids. Furthermore, we demonstrate that OMV production and regulation of the VacJ/Yrb ABC transport system respond to iron starvation. Our results suggest a new general mechanism of OMV biogenesis based on PL accumulation in the outer leaflet of the outer membrane. This mechanism is highly conserved among Gram-negative bacteria, provides a means for regulation, can account for OMV formation under all growth conditions, and might have important pathophysiological roles in vivo.
Project description:Comparing the gene expression profiling of HDGF-silenced RD-ES cells and control RD-ES cells to identify genes regulated by HDGF in RD-ES cells. Keywords: expression analysis Control RD-ES cells and HDGF-silenced RD-ES cells were profiled on 22K Human Genome Array
Project description:To analyze gene expression of RD cells after treatment with an anti-picornavirus compound MDL-860, we have employed whole genome microarray expression profiling as a discovery platform. Human RD cells were treated with mock or MDL-860 (final concentration of 20 uM) for 24 hours. A cluster of genes regulated Nrf2-Keap1 antioxidant pathway were identified by gene ontology analysis. Overall design: RD cells (human rhabdomyosarcoma cell line) were treated with mock or MDL-860 (final concentration of 20 uM) for 24 hours. Total RNA purified from each treatment was used for microarray analysis.
Project description:Objectives: The collagen VI related muscular dystrophies (COL6-RD), Ullrich congenital muscular dystrophy (UCMD) and Bethlem myopathy (BM) are among the most common congenital muscular dystrophies, but the pathogenesis, including the role of mutant collagen VI in the matrix is poorly understood. To better define the pathways disrupted by mutations in collagen VI, we have used a transcriptional profiling approach with RNA-Seq to identify differentially expressed genes in COL6-RD patients from controls. Methods: We have used RNA-Seq to identify differentially expressed genes in cultured dermal fibroblasts from 13 COL6-RD patients (8 dominant negative and 5 null) and 6 controls. Sequence reads were analyzed using the TopHat/Cufflinks pipeline. Results: Differentially expressed transcripts between COL6-RD patient and control fibroblasts include upregulation of ECM components and downregulation of factors controlling matrix remodeling and repair. DN and null samples are differentiated by downregulation of genes involved with DNA replication and repair in null samples Overall design: Expression profiles of dermal fibroblasts from 13 COL6-RD patients with dominant negative (8) or null (5) mutations compared to 6 control fibroblasts.
Project description:<Objective> To compare gene expression in labial salivary glands (LSG) of IgG4-related disease (IgG4-RD) with Sjögren’s syndrome (SS). <Methods> Gene expression was analyzed by DNA microarray in LSG of IgG4-RD (n=5), SS (n=5), and healthy controls (n=3). Differentially expressed genes (DEGs) in IgG4-RD and SS were identified, and gene-annotation enrichment analysis of these DEGs was performed using Gene Ontology (GO) annotation. Validation of the result was performed by quantitative PCR using LSG from IgG4-RD (n=9), SS (n=10), and controls (n=4). <Results> Gene expression patterns in IgG4-RD, SS, and healthy controls were quite different with each other in hierarchical clustering as well as principal component analysis. In IgG4-RD, 1771 up-regulated probe sets (corresponding to 1321 up-regulated genes) and 1785 down-regulated probe sets (corresponding to 1320 down-regulated genes) were identified as DEGs (false discovery rate <0.05). GO term analysis indicated that the up-regulated set of DEGs in IgG4-RD encoded proteins that function in cell proliferation, extracellular matrix organization, and organ development. PCR validated significantly higher expression of lactotransferrin (LTF) in IgG4-RD than SS (P<0.05), and chemokine (C-C motif) ligand 18 (CCL18) in IgG4-RD than SS and controls (P<0.05). <Conclusion> The results clearly showed that the gene expression pattern in LSG of IgG4-RD is different from that of SS. LSG samples were collected from 5 Japanese patients with IgG4-RD, as well as from 5 Japanese patients with SS who had been followed up at University of Tsukuba Hospital (Ibaraki, Japan). All patients with IgG4-RD satisfied the comprehensive diagnostic criteria for IgG4-related disease (IgG4-RD) 2011 proposed by the All Japan IgG4 team . The diagnosis of IgG4-RD was based on the presence of all three items; 1) clinical examination showing characteristic diffuse/localized swelling or masses in single or multiple organs, 2) Hematological examination shows elevated serum IgG4 concentrations (135 mg/dl), 3) Histopathologic examination shows: (1) Marked lymphocyte and plasmacyte infiltration and fibrosis. (2) Infiltration of IgG4+ plasma cells: ratio of IgG4+/IgG+ cells > 40% and >10 IgG4+ plasma cells/HPF. All patients with SS satisfied the Japanese Ministry of Health criteria for the diagnosis of SS (1999) . These criteria included four clinicopathological findings: lymphocytic infiltration of the salivary or lacrimal glands, dysfunction of salivary secretion, keratoconjunctivitis sicca, and presence of anti-SS-A or SS-B antibodies. The diagnosis of SS was based on the presence of two or more of the above four items. Approval for this study was obtained from the local ethics committee and a signed informed consent was obtained from each subject.
Project description:We have used surgical specimens to perform a differential analysis of the transcriptome of human retinal tissues following detachment. Data analysis reveals major involvement of the immune response in the disease and interindividual variation was monitored to unravel a second crucial aspect of the pathological process, the death of photoreceptor cells. In total 38 surgical specimens (samples) were analyzed, 19 represent biopsies from patients with retinal detachment (RD) and 19 represent contral samples without retinal detachment.
Project description:We have used surgical specimens to perform a differential analysis of the transcriptome of human retinal tissues following detachment. Data analysis reveals major involvement of the immune response in the disease and interindividual variation was monitored to unravel a second crucial aspect of the pathological process, the death of photoreceptor cells. Overall design: In total 38 surgical specimens (samples) were analyzed, 19 represent biopsies from patients with retinal detachment (RD) and 19 represent contral samples without retinal detachment.