scholarly journals P017 C86/CD16 macrophages accumulate in the mucosa of B3 patients and could mediate EMT in Crohn’s disease

2020 ◽  
Vol 14 (Supplement_1) ◽  
pp. S138-S138
Author(s):  
A Boronat-Muñoz ◽  
A Cejudo-Garces ◽  
P Lledo-Gil ◽  
J Cosín-Roger ◽  
S Coll ◽  
...  
Keyword(s):  
Crohn’S Disease ◽  
Statistical Analysis ◽  
Crohn's Disease ◽  
Mrna Expression ◽  
U937 Cells ◽  
Macrophage Phenotype ◽  
Rt Pcr ◽  
Intestinal Tissue ◽  
Healthy Donors ◽  
Emt Markers

Abstract Background Macrophages contribute to fibrosis through the release of different mediators and the pattern of secretion may vary according to their phenotype. Methods The aim of the present study is to analyse the pattern of expression of macrophages, of EMT-related genes and cytokines in surgical resections from Crohn’s disease (CD, n = 43) patients which were categorised according to Montreal classification (B2 or B3); unaffected mucosa of patients with ileocecal cancer was used as control (n = 20). mRNA was isolated from intestinal samples and the expression of macrophage, EMT markers and cytokines were analysed by RT-PCR. PBMCS were isolated from healthy donors and treated during 5 days with secretomes, from control, B2 or B3 surgical resections; the mRNA expression of macrophage markers was determined by RT-PCR. U937 cells were differentiated to macrophages and then treated with IFNγ (20 ng/ml) for 4 days, the mRNA expression of macrophages markers were determined by RT-PCR. Results are expressed as mean ± SEM (n ≥ 5). Statistical analysis was performed by ANOVA + Newman–Keuls or t-test. Correlations between data were analysed using Pearson’s correlation coefficient (*p < 0.05). Results The expression of CD16 and CD86 was significantly higher in intestinal samples from B3 CD patients (7.2 ± 1.1 and 7.7 ± 1.3, respectively) than in controls (1.4 ± 0.2 and 2.5 ± 0.4, respectively) or B2 CD patients (4.8 ± 0.9 and 4.5 ± 0.6, respectively). The mRNA expression of CD16 and CD86 were significantly higher in PBMCS treated with B3-secretomes than in those treated with B2- or control secretomes. The expression of CD16 and CD86 significantly correlated with FSP1 (r = 0.74, p = 0.002* and r = 0.66, p = 0.003*, respectively), VIMENTIN (r = 0.60, p = 0.02* and r = 0.82, p = 0.001*, respectively), SNAIL1 (r = 0.61, p < 0.01* r = 0.52, p = 0.04*, respectively), IL4 (r = 0.63, p = 0.01* and r = 0.60, p = 0.02*, respectively) and IFNγ (r = 0.56, p = 0.001* and r = 0.58, p = 0.01*, respectively) in intestinal tissue from the fistulising CD group. U937 cells treated with IFNγ increased significantly the mRNA expression of CD16 (1.94 ± 0.24* vs. vehicle) and CD86 (1.60 ± 0.17* vs. vehicle). Conclusion A macrophage phenotype expressing CD86/CD16 may act as a source of EMT mediators in intestinal tissue from CD patients with a penetrating (B3) behaviour. IFNγ could be responsible for the increase in the number of CD86/CD16 macrophages in the B3 behaviour.

scholarly journals PTPN2 is Associated with Crohn’s Disease and Its Expression Is Regulated by NKX2-3

2012 ◽  
Vol 32 (2) ◽  
pp. 83-91 ◽  
Author(s):  
Wei Yu ◽  
John P. Hegarty ◽  
Arthur Berg ◽  
Ashley A. Kelly ◽  
Yunhua Wang ◽  
...  
Keyword(s):  
Ulcerative Colitis ◽  
Crohn’S Disease ◽  
B Cells ◽  
Crohn's Disease ◽  
Mrna Expression ◽  
Genetic Variants ◽  
Microvascular Endothelial Cells ◽  
Rt Pcr ◽  
Risk Gene ◽  
Mrna Expression Levels

PTPN2 is a risk gene for Crohn's disease (CD). We investigated whether PTPN2 genetic variants (rs2542151 and rs2542152) were associated with CD in a familial IBD registry. Both rs2542151 and rs2542152 are associated with CD, but not ulcerative colitis (UC). mRNA expression levels of PTPN2 were significantly increased in intestinal tissues (p=0.0493), and nearly significantly increased in B cells (p=0.0889) from CD patients, but not significantly altered in UC. cDNA microarray results found that PTPN2 was down-regulated by NKX2-3 knockdown in human cells. We confirmed this observation by RT-PCR analyses in NKX2-3 knockdown in B cells from IBD patients and human intestinal microvascular endothelial cells (HIMEC). In addition, we found that mRNA expression of another IBD-associated gene, NKX2-3, was increased in intestinal tissues and B cells from CD patients, but not significantly increased in UC patients. A positive correlation was observed between mRNA expression of PTPN2 and NKX2-3 in B cells and in intestinal tissues from both CD and UC patients. These results suggest that PTPN2 may have an important role in CD pathogenesis and may represent a potential diagnostic and therapeutic target for IBD.

scholarly journals P089 IFNγ-macrophages could mediate EMT in Crohn’s disease

2021 ◽  
Vol 15 (Supplement_1) ◽  
pp. S189-S189
Author(s):  
J Marín-Aracil ◽  
M D Barrachina ◽  
C Bauset ◽  
J Cosin-Roger ◽  
S Coll ◽  
...  
Keyword(s):  
Protein Expression ◽  
Mrna Expression ◽  
Epithelial Mesenchymal Transition ◽  
Macrophage Phenotype ◽  
Rt Pcr ◽  
Mesenchymal Transition ◽  
Ht29 Cells ◽  
Mrna And Protein Expression ◽  
Fold Induction ◽  
Emt Markers

Abstract Background Macrophages contribute to fibrosis by releasing different mediators and the pattern of secretion may vary depending on the surrounding environment. We previously described that the mRNA expression of IFNγ was significantly higher in intestinal samples from CD patients. The aim of the present study is to analyze the role of IFNγ-treated macrophages in epithelial mesenchymal transition (EMT). Methods The mRNA and protein expression of IFN in surgical resections from Crohn′s disease. U937 were differentiated to macrophages and then treated with IFNγ (2 ng/ml) for 4 days, the mRNA expression of macrophages markers were determined by RT-PCR. IFNγ-U937 were coculture with HT29 cells for 2 days and the expression of EMT markers in HT29 cells were analyzed by RT-PCR and WB. Results are expressed as mean±SEM (n≥5). Statistical analysis was performed by ANOVA + Newman-Keuls. Results The mRNA and protein expression of IFNγ were significantly higher in intestinal samples from B2 CD patients (11.4±1.6 fold induction and 3.5±0.3 pg/mg, respectively) and B3 CD patients (14.2±1.8 fold induction and 3.1±0.1 pg/mg, respectively) than in controls (1,0±0,1 fold induction and 0.8±0,1 pg/mg, respectively). U937 cells treated with IFNγ increased significantly the mRNA expression of CD16 (1.9±0.2* vs vehicle) and CD86 (1.6±0.1* vs vehicle). IFNγ-U937 cocultured with HT29 increased significantly the mRNA and protein expression of EMT markers in HT29 cells (Vimentin: 3.0±0.5* vs vehicle-HT29; αSMA: 24.4±8.3* vs vehicle-HT29; SNAIL1: 2.7±0.5* vs vehicle-HT29) respect IFNγ-HT29 cells (Vimentin: 0.6±0.01 vs vehicle-HT29; αSMA: 1.1±0.4 vs vehicle-HT29; SNAIL1: 0.7±0.06 vs vehicle-HT29). Conclusion IFNγ could be responsible for the increase in the number of CD86/CD16 macrophages in CD patients. A macrophage phenotype expressing CD86/CD16 may act as a source of EMT mediators in intestinal tissue from CD patients.

ACTIVATED INTESTINAL MUSCLE CELLS PROMOTE PREADIPOCYTE MIGRATION: A NOVEL MECHANISM OF CREEPING FAT FORMATION IN CROHN’S DISEASE

2021 ◽  
Vol 27 (Supplement_1) ◽  
pp. S34-S34
Author(s):  
Ren Mao ◽  
Genevieve Doyon ◽  
Ilyssa Gordon ◽  
Jiannan Li ◽  
Sinan Lin ◽  
...  
Keyword(s):  
Extracellular Matrix ◽  
Crohn’S Disease ◽  
Crohn's Disease ◽  
Ex Vivo ◽  
Muscularis Propria ◽  
Dominant Factor ◽  
Stricture Formation ◽  
Intestinal Tissue ◽  
Mesenteric Fat

Abstract Background and Aims Creeping fat, the wrapping of mesenteric fat around the bowel wall, is a typical feature of Crohn’s disease, and is associated with stricture formation and bowel obstruction. How creeping fat forms is unknown, and we interrogated potential mechanisms using novel intestinal tissue and cell interaction systems. Methods Tissues from normal, ulcerative colitis, non-strictured and strictured Crohn’s disease intestinal specimens were obtained. Fresh and decellularized tissue, mesenteric fat explants, primary human adipocytes, pre-adipocytes, muscularis propria cells, and native extracellular matrix were used in multiple ex vivo and in vitro systems involving cell growth, differentiation and migration, proteomics, and integrin expression. Results Crohn’s disease muscularis propria cells produced an extracellular matrix scaffold which is in direct spatial and functional contact with the immediately overlaid creeping fat. The scaffold contained multiple proteins, but only fibronectin production was singularly upregulated by TGF-b1. The muscle cell-derived matrix triggered migration of pre-adipocytes out of mesenteric fat, fibronectin being the dominant factor responsible for their migration. Blockade of α5β1 on the pre-adipocyte surface inhibited their migration out of mesenteric fat and on 3D decellularized intestinal tissue extracellular matrix. Conclusion Crohn’s disease creeping fat appears to result from the migration of pre-adipocytes out of mesenteric fat and differentiation into adipocytes in response to an increased production of fibronectin by activated muscularis propria cells. These new mechanistic insights may lead to novel approaches for prevention of creeping fat-associated stricture formation.

Probing serum N-glycan patterns for rapid and precise detection of Crohn's disease

2021 ◽  
Author(s):  
Yonglei Wu ◽  
Yijie Chen ◽  
Haolin Chen ◽  
Chenjie Yang ◽  
Xizhong Shen ◽  
...  
Keyword(s):  
Crohn’S Disease ◽  
Statistical Analysis ◽  
Crohn's Disease ◽  
Carbon Matrix ◽  
Healthy Controls

Serum N-glycan patterns from 50 Crohn‘s disease (CD) patients and 50 healthy controls were acquired by a carbon matrix-based platform. According to statistical analysis, eight specific N-glycans revealed remarkable performance for CD diagnosis.

scholarly journals Differential between Protein and mRNA Expression of CCR7 and SSTR5 Receptors in Crohn's Disease Patients

2009 ◽  
Vol 2009 ◽  
pp. 1-9 ◽  
Author(s):  
Nathalie Taquet ◽  
Serge Dumont ◽  
Jean-Luc Vonesch ◽  
Didier Hentsch ◽  
Jean-Marie Reimund ◽  
...  
Keyword(s):  
Crohn’S Disease ◽  
Crohn's Disease ◽  
Protein Expression ◽  
Mrna Expression ◽  
Large Scale ◽  
Mononuclear Cells ◽  
Biologically Active ◽  
Chronic Inflammatory Bowel Disease ◽  
Peripheral Blood Mononuclear ◽  
Mrna And Protein Expression

Crohn's disease (CD) is a multifactorial chronic inflammatory bowel disease of unknown cause. The aim of the present study was to explore if mRNA over-expression of SSTR5 and CCR7 found in CD patients could be correlated to respective protein expression. When compared to healthy donors, SSTR5 was over-expressed 417±71 times in CD peripheral blood mononuclear cells (PBMCs). Flow cytometry experiments showed no correlation between mRNA and protein expression for SSTR5 in PBMCs. In an attempt to find a reason of such a high mRNA expression, SSTR5 present on CD PBMCs were tested and found as biologically active as on healthy cells. In biopsies of CD intestinal tissue, SSTR5 was not over-expressed but CCR7, unchanged in PBMCs, was over-expressed by 10±3 times in the lamina propria. Confocal microscopy showed a good correlation of CCR7 mRNA and protein expression in CD intestinal biopsies. Our data emphasize flow and image cytometry as impossible to circumvent in complement to molecular biology so to avoid false interpretation on receptor expressions. Once confirmed by further large-scale studies, our preliminary results suggest a role for SSTR5 and CCR7 in CD pathogenesis.

scholarly journals Application value of tissue tuberculosis antigen combined with Xpert MTB/RIF detection in differential diagnosis of intestinal tuberculosis and Crohn’s disease

2020 ◽  
Author(s):  
Baoying Fei ◽  
Lin Zhou ◽  
Yu Zhang ◽  
Linhe Luo ◽  
Yuanyuan Chen
Keyword(s):  
Differential Diagnosis ◽  
Crohn’S Disease ◽  
Crohn's Disease ◽  
Intestinal Tuberculosis ◽  
Surgical Excision ◽  
Endoscopic Biopsy ◽  
Detection Sensitivity ◽  
Intestinal Tissue ◽  
Tissue Samples ◽  
Tuberculosis Antigen

Abstract Background: The purpose of this study was to evaluate the value of Xpert MTB/RIF detection and tuberculosis antigen detection of Mycobacterium tuberculosis cluster (MTBC) in intestinal tissues for differentiating intestinal tuberculosis (ITB) from Crohn’s disease (CD). Methods: A total of 110 patients who were clinically diagnosed with CD or ITB were monitored. Several specimens of intestinal tissue from endoscopic biopsy or surgical excision were used for culture and Xpert MTB/RIF for detection of MTBC, respectively. Four antigens (38KDa, ESAT-6, MPT64, Ag85 complex) of MTBC in intestinal tissue were detected by immunohistochemistry. Results: A total of 42 cases of intestinal tuberculosis and 46 cases of CD were included in the experimental analysis. Perianal lesions and longitudinal ulcers were more common in CD patients (p < 0.05), while caseous granuloma and annular ulcers were more common in ITB patients (P < 0.05). The positive rate of MTBC detected by Xpert MTB/RIF in intestinal tissue samples of ITB patients was 33.33%, which was significantly higher compared to CD patients (p < 0.05) and compared to acid-fast staining smears (9.52%) (p < 0.05). The positive MPT64 expression rate in patients with intestinal tuberculosis was 40.48%, which was significantly higher than that observed in CD patients, which was 19.56% (p<0.05). Conclusions: The detection of Xpert MTB/RIF in intestinal tissue is a rapid and useful method for establishing an early diagnosis of intestinal tuberculosis. The detection of Xpert MTB/RIF and MPT64 antigen in intestinal tissues have definitive value in the differential diagnosis of intestinal tuberculosis and Crohn’s disease. The combination of these two methods could improve detection sensitivity.

scholarly journals Long-read sequencing to interrogate strain-level variation among adherent-invasive Escherichia coli isolated from human intestinal tissue

2020 ◽  
Author(s):  
Jeremy Wang ◽  
Rachel Bleich ◽  
Sandra Zarmer ◽  
Janelle Arthur
Keyword(s):  
Escherichia Coli ◽  
Crohn’S Disease ◽  
Crohn's Disease ◽  
Pathogenic Bacteria ◽  
Genomic Variation ◽  
Strain Level ◽  
Level Variation ◽  
Intestinal Tissue ◽  
E Coli ◽  
Long Read

AbstractAdherent-invasive Escherichia coli (AIEC) are a pathovar linked to inflammatory bowel diseases (IBD), especially Crohn’s disease, and colorectal cancer. AIEC have no known molecular or genomic markers, but instead are defined by in vitro functional attributes. Futhermore, it is unknown if strains classified as AIEC truly colonize intestinal tissues better than non-AIEC strains. To evaluate strain-level variation among tissue-associated E. coli, we must develop a sequencing approach capable of long reads and with the ability to exclude mammalian DNA. We also must evaluate genomic variation among strains that have demonstrated ability to colonize intestinal tissues. Here we have assembled complete genomes using ultra-long-read nanopore sequencing for a model AIEC strain, NC101, and seven strains isolated from the intestinal mucosa of Crohn’s disease and non-Crohn’s tissues. We show these strains can colonize the intestinal tissue in a Crohn’s disease mouse model and induce varying levels of inflammatory cytokines from cultured macrophages. We demonstrate these strains can be quantified and distinguished in the presence of 99.5% mammalian DNA and from within a fecal population. Analysis of global genomic structure and specific sequence variation within the ribosomal RNA operon provides a framework for efficiently tracking strain-level variation of closely-related E. coli and likely other commensal/pathogenic bacteria impacting intestinal inflammation in mice and IBD patients.

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