Project description:Crohn's Disease (CD) is a chronic inflammatory disease of the intestinal tract. We performed a whole-genome transcriptional analysis using colonic biopsies from CD patients before and after anti-TNF-α therapy. Involved colonic samples from Crohn's disease patients and healthy colonic samples from non-inflammatory controls were collected for RNA extraction and hybridization on Affymetrix microarrays. Inclusion criteria for CD patients were: age between 18 and 70, diagnosis of CD established at least 4 months before inclusion and exclusion of concomitant infection. Active disease was defined by endoscopic and clinical score: endoscopic active disease was defined as a CD endoscopic index of severity (CDEIS) of 5 or more and the presence of large ulcers (> 0.5 cm diameter) in at least one of the explored segments. Clinical activity was defined as a CD activity index (CDAI) above 150. Finally, a total of 39 biopsies were analyzed: 17 healthy controls, 10 active CD without anti-TNF therapy, 5 active CD with anti-TNF therapy (non-responders) and 7 inactive CD with anti-TNF therapy (responders).
Project description:Crohn's Disease (CD) is a chronic inflammatory disease of the intestinal tract. We performed a whole-genome transcriptional analysis using colonic biopsies from CD patients before and after anti-TNF-α therapy.
Project description:Crohn’s disease (CD) is a chronic inflammatory intestinal disease, often characterized by aberrant healing and stricturing complications. Mechanisms underlying NOD2-pathogenicity and salvage pathways in anti-TNF and refractory patients remain largely uncharacterized. Here we show that loss of NOD2 function leads to aberrant activated fibroblast and macrophage homeostasis through the upregulation of a pathogenic signature, and propose new precision therapeutic approaches involving gp130 blockade for select CD patients, to potentially supplement anti-TNF therapy
Project description:The expression of Triggering Receptor Expressed on Myeloid cells (TREM)-1 has been described as a predictive marker for anti-Tumor Necrosis Factor (TNF)-α monoclonal antibody (mAb) therapy responsiveness in patients with inflammatory bowel disease (IBD). Here we investigated expression of TREM-1 specifically in CD14+ monocytes in relation to anti-TNF response. The pretreatment TREM-1 expression levels of CD14+ monocytes of Crohn’s disease (CD) patients were predictive of outcome to anti-TNF mAb therapy, with low TREM-1 expression associated with response to anti-TNF. FACSorting of CD14+ monocytes with different TREM-1 levels showed that differentiation towards regulatory CD206+ M2 type macrophages by anti-TNF was suppressed in CD14+ monocytes with high TREM-1 expression. Activity of the Fcγ-Receptor and autophagy pathway, both necessary for M2 type differentiation and the response to anti-TNF, were decreased in CD14+ monocytes with high expression of TREM-1. We confirmed that the activity of the Fcγ-Receptor pathway was decreased in the CD patients that did not respond to anti-TNF therapy and that it was negatively correlated with TREM-1 expression levels in the CD patient cohort. In conclusion, our results indicate that TREM-1 expression levels in CD14+ monocytes associate with decreased autophagy and FcγR activity resulting in decreased differentiation to M2 type regulatory macrophages upon anti-TNF mAb treatment, which may explain anti-TNF non-response in IBD patients with high expression levels of TREM-1.
Project description:Differential gene expression analysis between CD patients and controls to identify the transcriptional signature that defines the inflamed intestinal mucosa in CD.
Project description:This study employs spectroscopy-based metabolic profiling of fecal extracts from healthy subjects and patients with active or inactive ulcerative colitis (UC) and Crohn's disease (CD) to substantiate the potential use of spectroscopy as a non-invasive diagnostic tool and to characterize the fecal metabolome in inflammatory bowel disease (IBD). Stool samples from 113 individuals (UC 48, CD 44, controls 21) were analyzed by 1H nuclear magnetic resonance (NMR) spectroscopy (Bruker 600 MHz, Bruker BioSpin, Rheinstetten, Germany). Data were analyzed with principal component analysis and orthogonal-projection to latent structure-discriminant analysis using SIMCA-P plus 12 and MATLAB. Significant differences were found in the metabolic profiles making it possible to differentiate between active IBD and controls and between UC and CD. The metabolites holding differential power primarily belonged to a range of amino acids, microbiota-related short chain fatty acids, and lactate suggestive of an inflammation-driven malabsorption and dysbiosis of the normal bacterial ecology. However, removal of patients with intestinal surgery and anti-TNF-alpha antibody treatment eliminated the discriminative power regarding UC versus CD. This study consequently demonstrates that 1H NMR spectroscopy of fecal extracts is a potential non-invasive diagnostic tool and able to characterize the inflammation-driven changes in the metabolic profiles related to malabsorption and dysbiosis. Intestinal surgery and medication are to be accounted for in future studies, as it seems to be factors of importance in the discriminative process.
Project description:Crohn's disease (CD) is a chronic condition characterized by recurrent flares of inflammation of the gastrointestinal tract. Despite decades of research, the etiology of CD is poorly understood and is characterized by dysregulated immune activation that progressively destroys the gastrointestinal tissue. At the center of the pathology is the intestinal mucosa, where the epithelium and the lamina propria are main cellular compartments. While the epithelium contains predominantly epithelial cells, the lamina propria is enriched in immune cells and contains supporting stroma. Thus, the cells constituting these compartments can be classified as epithelial cells, immune cells and stromal cells. Gaining insight into how these cell types interact with each other is important to further our understanding of CD pathogenesis. Here, using isobaric labeling-based quantitative proteomics, we perform an exploratory study to analyze in-depth proteome changes in epithelial cells, immune cells and stromal cells purified by cell sorting from the intestinal mucosa of CD patients and controls. Our workflow preserving the link between the different cell types in individuals opens the possibility to explore cellular crosstalk and to further refine data on cell-cell interactions in the development of CD.
Project description:Whole blood samples were collected from patients at baseline, 6 weeks and 8 weeks after induction (Ustekinumab or placebo) therapies for RNA extraction and microarray analysis from patients with moderate-to-severe CD who participated in stelara CD phase 3 studies (UNITI-2). These patients failed conventional therapies previously and largely naive to anti-TNF therapy. We used microarrays to detail the transcriptional programme underlying placebo and stelara treatment in the periphery at WK0, WK6, WK8.
Project description:An integrated discovery to targeted proteomics approach was used to investigate the protein profiles of good and non–responders to anti-TNF-alpha and T-cell inhibitor treatments in PsA patients. Reverse phase liquid chromatography coupled to tandem mass spectrometry was used to generate protein profiles of synovial tissue obtained at baseline from 10 PsA patients who then commenced anti-TNF-alpha therapy (adalimumab). Targeted proteomics using multiple reaction monitoring was used to confirm and pre-validate a potential protein biomarker panel in 18 and 7 PsA patient samples respectively.
Project description:Mouse intestinal innate immune cells have been identified as inducing Th17 cell differentiation. However, few studies have investigated innate immune cells that are involved in CD (Crohn's disiease) pathogenesis in humans. The present study aimed to identify a human intestinal lamina propria cell subset that induces Th17 cells, and to clarify its role in CD pathogenesis. Intestinal mucosa samples were obtained from patients who underwent resection of colorectal cancer or CD. Lamina propria cells (LPCs) were obtained by enzymatic digestion, and were analyzed for expression of HLA-DR, lineage markers (Lin), CD14, and CD163 using flow cytometry. Among HLA-DRhigh Lin- cells, we identified a CD14+ CD163low cell subset that was selectively present in intestinal LPCs. We identified CD14+ CD163low cells as having a potent capacity to induce Th17 cell differentiation in human intestinal lamina propria. The Th17-inducing activity of these cells was enhanced in CD patients. The Cy3-labeled cDNAs were hybridized on Agilent-039494 SurePrint G3 Human GE v2 8x60K Microarray (G4845A) in one color experiments, resulting in four individual microarrays.