bluuebunny/crossref_metadata_2025_split huggingface dataset:
P004 Untargeted serum metabolome shows Crohn Disease (CD) associated signature, signals that correlate with disease activity and CRP, and signals that may precede disease flare
Authors: Levhar N , Hadar R , Braun T , Efroni G , Abramovich I , Gottlieb E , Granot M , Sandra N , Selinger L , Picard O , Yavzori M , Lahat A , Eliakim R , Weiss B , Kopylov U , Ben-Horin S , Amir A , Haberman Ziv Y 
Published: January 2023

Abstract

Background
The serum metabolome contains potential disease biomarkers, some of which are produced endogenously, derived from the diet, or formed by the gut microbiome. We aimed to characterize metabolomics profiles linked with Crohn Disease (CD), with active CD, and with remitting pre-flare CD.

Methods
Serum untargeted metabolomics was performed using liquid chromatography Mass Spectrometer (LC/MS). PERMANOVA on beta diversity was used to define factors affecting serum metabolome. Multivariate analyses in MaAsLin2 were applied to identify differentially abundant metabolites between CD and controls, between active CD and those in remission, and between those that subsequently flared and those that remained in remission within 24 months of follow-up.

Results
The cohort included 202 samples from 53 CD patients and 31 healthy controls (156 samples from CD and 46 from controls; Table 1). 545 metabolites were included in our predefined library and were subsequently analyzed. PERMANOVA on the serum metabolomic data indicated that fasting, age, gender, disease, and subject are significant factors contributing to metabolomics variations. Using multivariate analyses and controlling for fasting, age, gender, and subject, we identified 34 specific metabolites that were significantly higher in CD and 26 that were significantly higher in Controls (FDR&lt;0.25; Fig. 1). CD samples were significantly enriched for purine derivatives, simple sugars, and histidine, compared to healthy controls. In contrast, amino-acids, caffeine, and TCA cycle related metabolites were consistently downregulated in CD. 37 metabolites were higher in active CD and 24 in quiescent CD. Saturated fatty acids and acetylated amino-acids were positively correlated with active CD, while histidine and other standard amino-acids were downregulated in active samples compared to remission. Interestingly, acetylated amino acids, asparagine and phenylalanine, were also significantly correlated with CRP. A sub-cohort of 25 CD subjects was enrolled during remission and monitored every 3 months over 2 years. During that prospective follow-up, 11 flared. Focusing on the pre-flare samples, patients who eventually flared showed a significant reduction in N-acetyl glycine and reduction in two ketogenic derivates, acetoacetate and 3-hydroxybutyric acid, in comparison to patients who remained in remission throughout (FDR&lt;0.15).

Conclusion
CD, CD activity, and a subsequent flare in CD patients with longitudinal data were associated with distinct serum metabolic signature after controlling for confounders like age, gender, and fasting. Additional validation is needed to test if these can be used as predictive biomarkers.
P005 0190Molecular-based classification of ulcerative colitis and its dynamic
Authors: Seyed Tabib N S , Verstockt S , Verstockt B , Ferrante M , Sabino J , Vermeire S 
Published: January 2023

Abstract

Background
A better characterization of inflammatory bowel disease is needed. We previously developed a molecular classification tool for Crohn’s disease, based on a combination of metagenomics (dysbiosis score - MDI), transcriptomics (barrier integrity score - BIS, autophagy score – ATS, unfolded protein response score - UPRS), serum proteomics (inflammation score - IPS) and genetics (polygenic risk score - GRS)1. We here investigated if this score could also apply for ulcerative colitis (UC) and how the individual components correlate with clinical outcomes.

Methods
For a total of 173 UC patients and 181 controls (CO), we collected faecal and blood samples, as well as colon biopsies (Table). Faecal microbiota 16S-sequencing, intestinal RNA-sequencing, and serum proteomics (OLINK inflammatory panel) were performed. Using proteomics and RNA expression data, IPS, ATS, UPRS and BIS were calculated. MDI was calculated as previously described1. Based on these findings, patients were categorized into quartiles, ranging from Q1 (the least dysfunctional state) to Q4 (the most dysfunctional state). The individual scores were correlated with time to first biological or biological switch, as well as correlations with C-reactive protein (CRP), faecal calprotectin (FC), and the Montreal classification. The therapeutic value of MDI score was studied in a small dietary intervention pilot study (n=8)2.

Results
None of the individual components of the score were associated with disease extent. Patients diagnosed at a later age had increased ATS and BIS levels. BIS and UPRS showed a positive correlation with disease duration (r=0.36, r=0.54 respectively, p&lt;0.001) (Figure2). MDI (spearman r=0.27, r=0.44, r=0.46 respectively, all p&lt;0.0001) and IPS (r=0.5, r=0.7, r=0.6 respectively, all p&lt;0.0001) correlated with CRP and FC levels, and time to first biologic. UPRS, ATS, MDI, BIS, and IPS were considerably higher in patients who required a switch in biologic therapy (all p&lt;0.001). Finally, we observed a decreasing trend in MDI in response to dietary modification (Figure3).

Conclusion
In UC patients, we created a multifaceted scoring system to molecularly describe disease by the degree of dysbiosis, dysregulation of the immune proteome, and intestinal barrier integrity. We also demonstrated the dynamic of MDI of this score in relation to dietary intervention. Therefore, molecular profiling of patients may represent a new, individualized strategy to the management of UC.
References:
1. OP30 ECCO 2020, JCC 14(Supplement_1):S028-S0302.
2. P767 ECCO 2017, JCC 11(suppl_1):S473-S473.
P008 Fibrostricturing Crohn’s disease is characterised by an imbalance in active eosinophils, Th1, Th2 and regulatory T cells
Authors: Jacobs I , Ke B J , Cremer J , D’Hoore A , Bislenghi G , Matteoli G , De Hertogh G , Sabino J , Ferrante M , Vermeire S , Breynaert C , Vanuytsel T , Verstockt B 
Published: January 2023

Abstract

Background
About one third of patients with Crohn’s disease (CD) develop strictures during their disease course requiring surgical resection. The immune landscape involved in this process is poorly understood. Therefore, we aimed to characterise the fibroblast phenotype, immune cells and their mediators involved in intestinal strictures.

Methods
We included 25 CD patients with stricturing disease in the terminal ileum (TI) and 10 controls with colorectal cancer (CRC), all undergoing an ileocolonic resection. Transmural samples from the resection specimen of the TI were obtained. Macroscopically, CD tissue was divided into unaffected, fibrostenotic and inflamed regions by an experienced histopathologist. Next, mucosa was separated from deeper layers, after which single cells were isolated and fluorescently stained for flow cytometry. Protein levels were determined via the MesoScale Discovery (MSD) platform in the corresponding samples. Comparisons between CRC controls and CD patients were performed via an unpaired t-test or Mann-Whitney analysis and corrected for multiple testing.

Results
An increase in active fibroblasts and decrease in inactive fibroblasts in the fibrotic and inflamed mucosa (p=0.0002 and p&lt;0.0001) and deeper layers (p=0.003 and p=0.02) when compared to the CRC controls was observed, confirming ongoing tissue remodelling. An enrichment of active eosinophils was only seen in the fibrotic deeper layers (p=0.02), although an increase in T helper 2 (Th2) cells was observed in both the fibrotic and inflamed deeper layers (p=0.02 and p=0.04). In contrast, T helper 1 (Th1) cells were decreased in both fibrotic and inflamed mucosa (p=0.03 and p=0.02) and deeper layers (p=0.01 for both). Regulatory T cells were significantly enriched in both fibrotic and inflamed mucosa (p&lt;0.0001 and p=0.0005) and deeper layers (p=0.01 and p=0.006) (figure 1). Protein quantification confirmed a significant increase in transforming growth factor-β3 (TGF-β3) in the fibrotic (p=0.007) and inflamed (p=0.0002) layers, but not in the more superficial mucosa. Comparably, IL-1β was increased in the fibrotic (p=0.05) and inflamed (p=0.05) deeper layers. A similar observation was made for basic fibroblast growth factor (bFGF) (p=0.004), although only a trend could be seen in the fibrotic deeper layers (p=0.08) (figure 2).

Conclusion
The fibrotic and inflamed tissue of CD patients is characterized by increased activated eosinophils, Th2 and regulatory T cells and decreased Th1 cells, as well as many of their mediator cytokines. The current immunological characterisation can help to prioritise potential anti-fibrotic targets for stricturing CD.
P011 Patient-derived organoids as an epithelial model for preclinical IBD research and drug development.
Authors: Frias J , Derksen M , Verissimo C S , van Dooremalen W T M , Oldenburg B , Vries R G J , Boj S F , Pourfarzad F , Pott J 
Published: January 2023

Abstract

Background
The study of the intestinal epithelial barrier is hampered by the lack of a suitable in vitro model system. Patient-derived organoids (PDOs) recapitulate the cellular diversity and patient heterogeneity and provide a relevant platform to study the intestinal epithelium and its response to microbial or inflammatory challenges.
Here, we present the characterization of an inflammatory bowel disease (IBD) organoid biobank derived from Crohn’s disease and Ulcerative Colitis patients and its application for IBD drug development efforts.

Methods
PDOs from different regions of the gastrointestinal tract of 14 IBD patients were established. To recapitulate the full cellular diversity, specialized medium formulations were defined that support the differentiation of distinct epithelial cell lineages as confirmed by transcriptional profiles and histology.
To study the inflammation driven loss of barrier integrity observed in the mucosa of IBD patients, we employed organoid derived monolayers and mimicked the inflammatory environment by cytokine challenge (TNF, IFNγ).

Results
In response to differentiation media, intestinal organoids and monolayers displayed gene expression patterns consistent with maturation of epithelial cell types found in the human gut.
The cytokine challenge robustly induces loss of barrier integrity across seven patient-derived monolayers. The barrier breakdown was ameliorated in all models by treatment with the IBD therapy Tofacitinib.

Conclusion
Intestinal PDOs were generated from IBD patients and were capable to differentiate into mature epithelial cell types. PDO-derived monolayers built trans-epithelial electrical resistance (TEER) and are a relevant model to study epithelial barrier function and inflammation mediated barrier breakdown. Proof-of-concept studies with Tofacitinib demonstrated the suitability of the assay for IBD drug development efforts.
P012 The administration of mm-miR-378a-3p exacerbates chronic inflammation and fibrosis in a murine intestinal model
Authors: Seco-Cervera M , Argilés-Arberola D , Bauset C , Lis-López L , Coll S , Cosín-Roger J , Ortiz-Masià D , Calatayud S , Barrachina M D 
Published: January 2023

Abstract

Background
Fibrosis constitute an important complication of CD. MicroRNAs (miRNAs), which are small RNA molecules that regulate gene expression, have been shown to participate in the molecular interactions of both inflammation and fibrosis. Lower levels of miR-378a-3p have been reported associated to murine liver fibrosis (Hyun et al. DOI: 10.1038/ncomms10993) and we analyze here the relevance of miR-378a-3p on intestinal fibrosis.

Methods
B57BL/6 mice were intravenously injected with 2,5mg/kg of negative control (NC) or mm-miR-378a-3p mimic, twice a week and received vehicle or Dextran Sulfate Sodium (DSS) for 2 cycles (7 days drinking DSS 2% in water solution followed by 10 days drinking water). Body weight and DAI score was obtained every day and the colon was collected after sacrifice. Sirius and hematoxylin-eosin dyes were employed to determine the fibrosis and structural state in 5µm slides of intestinal tissue. Gene expression and miRNA profiles were analyzed by RT-qPCR. Human small intestinal fibroblasts (HSIF; P10760, Innoprot, Spain) were transfected with 20nM of NC or hsa-miR-378a-3p mimic during 24h.

Results
No significant changes in body weight, DAI score, and colon length were detected between mice receiving NC and those receiving mimic, all along the two DSS cycles. Colon of mice treated with DSS, exhibited a significant diminution in the mRNA expression of mm-miR-378a-3p compared with naïve samples. In DSS-treated mice, the iv administration of the mimic compared with the NC: a) significantly increased levels of miR-378a-3p in the colon; b) increased the number of neutrophils, and heightened changes in the glandular epithelia and architectural distortion (figure 1a) C) decreased the number of lymphoid follicles (figure 1a); d) slightly increased collagen deposition, as analyzed by sirius red (figure 1b) and e) significantly increased the mRNA expression of Tgfb1, Il1b, and Mmp2. Treatment of human intestinal fibroblasts with hsa-miR-378a-3p mimic did not significant modify the mRNA expression of markers of fibrosis but it significantly increased the mRNA expression of two antiapoptotic molecules BCL2 and MCL1.

Conclusion
Levels of mm-miR-378a-3p are diminished in a murine model of intestinal fibrosis; The exogenous administration of miR-378a-3p, increased the acute inflammatory response and the expression of fibrosis markers in murine fibrotic colon and the expression of antiapoptotic molecules in human intestinal fibroblasts.
P017 Fecal wash host transcriptomics identifies inflammation throughout the colon and terminal ileum
Authors: Dan S , Ungar B , Ben-Moshe S , Bahar Halpern K , Yavzori M , Fudim E , Picard O , Abitbol C M , Harnik S , Kopylov U , Ben-Horin S , Itzkovitz S 
Published: January 2023

Abstract

Background
Non-invasive modalities for assessing active endoscopic and histological inflammation in Crohn’s disease and Ulcerative Colitis patients are critically needed. Fecal wash host transcriptomics has been shown to be a robust classifier of endoscopic and histological inflammation in inflammatory bowel disease (IBD) patients with distal colitis. Whether such fecal washes can inform on inflammatory processes occurring in more proximal intestinal segments is currently unknown.

Methods
Crohn's disease and Ulcerative Colitis patients as well as healthy controls were prospectively enrolled. Host transcriptomes of biopsies and fecal washes obtained during colonoscopy at predefined locations throughout the colon and terminal ileum were analyzed and associated with same time-point clinical, endoscopic and histological parameters.

Results
59 IBD patients and 50 controls were prospectively enrolled. Biospies and fecal washes from the distal, proximal colon and terminal ileum were compared. We find that host transcriptomics of distal fecal washes robustly classify histological inflammation in ileal and proximal colonic Crohn’s disease, even without distal colonic involvement. Receiver operating characteristic (ROC) curve analysis demonstrated significantly higher classification power based on the fecal wash transcriptomics compared to biopsies: Area Under the Curve, AUC=0.94+-0.07 for UC distal fecal washes and AUC=0.94+-0.09 for distal fecal washes of CD patients without distal colonic involvement, as compared to all distal biopsies AUC= 0.82+-0.20. We further demonstrate that fecal washes consist of modules of co-expressed immune, stromal and epithelial genes. Fecal wash expression of gene modules previously associated with lack of response to biological therapies correlates with disease severity.

Figure 1 - Host transcriptomics of fecal washes from different intestinal segments captures information that is distinct from same – segment biopsy transcriptomics. A) Experimental layout. B) Clustergram of transcriptomes of biopsies (purple) and fecal washes (yellow). C) Principal component analysis (PCA) of biopsies (purple) and fecal washes (yellow). Black circles highlight inflamed samples. D) Spearman correlations between the transcriptomes of pairs of either biopsies or fecal washes that are both annotated as inflamed and mixed pairs (one annotated as inflamed and the other not). (A) Created with BioRender.com. TI – terminal ileum, P – proximal colon, D – distal colon.

Conclusion
Our study establishes the power of colonic fecal washes for identifying pathological processes throughout the ileal-colonic axis.
P018 Expression of annexin A1 and NLRP3 inflamassome in Chron’s disease
Authors: Koga Ito H T , Lice I , Sasso G R S , Artigiani R , Gil C D 
Published: January 2023

Abstract

Background
Despite annexin A1 (ANXA1), an anti-inflammatory and pro-resolving protein, regulates the NLRP3 inflammasome activation in inflammatory cells, this relationship has not been explored in inflammatory bowel disease. Then, this study evaluated the expression patterns of ANXA1 and NLRP3 inflammasome in Crohn’s disease (CD).

Methods
Paraffin-embedded human intestine biopsies, obtained from CD (n=20) and control (n=8) patients were processed for histological and immunohistochemical analyses. The study was approved by the UNIFESP Ethics Committee (CEP 4.491.836/2020). GSE179285 study, which features ileum (Control - n=8, CD non-inflamed - n=48, CD inflamed - n=33) and colon (Control - n=12, CD non-inflamed – n=74, CD inflamed - n=14) transcriptome, was selected from the Gene Expression Omnibus repository. Datasets were analyzed using the GEO2R tool to detect ANXA1, CASP1, NLRP3, and PYCARD expressions.

Results
Bioinformatics analysis of GSE179285 samples showed a marked increase in the ANXA1 expression in the CD inflamed ileum and colon compared to CD non-inflamed and controls samples (Fig.1A). Regarding the genes associated with the NLRP3 inflammasome activation, transcriptome analysis also revealed a significant increase in NLRP3 and CASP1 in the CD inflamed intestine compared to the other ones (Fig.1B,C). PYCARD expression did not show alterations between the different conditions (Fig.1D). Immunohistochemical analyzes showed intense immunoreactivity for ANXA1 and NLRP3 proteins in the CD ileum and colon, especially in inflammatory infiltrate and epithelium of the mucosal layer (Fig.2C,D,G,H), compared to controls (Fig. 2A,B,E,F). These findings corroborate the transcriptome analyses.

Conclusion
ANXA1 and NLRP3 expression is increased in CD-inflamed intestines. High levels of ANXA1 may contribute to controlling inflammation and repairing intestinal injuries. High levels of NLRP3 evidence dysfunction in the pro-inflammatory activity of DC, contributing to the release of IL-1β and IL-18. Support: FAPESP (grants 20/03565-2, 21/01541-1 and 21/06059-3).
Figure 1. ANXA1, NLRP3, CASP1 and PYCARD expressions. Analysis performed in the healthy or CD terminal ileum and ascending/descending colon of human biopsies from GSE179285 study. Data represent mean ± S.E.M. of the gene expression values. **p&lt;0.01; ***p&lt;0.001; ****p&lt;0.0001 (Kruskal-Wallis, Dunn’s post-test).
Figure 2. ANXA1 and NLRP3 expression in human intestine. Representative images of ileum (A, C, E, G) and colon (B, D, F, H) of control (A,B,E,F) and CD patients (C,D,G,H). ANXA1 and NLRP3 expression detected in intestinal epithelium (arrows) and inflammatory infiltrate (asterisks). Counterstaining: Carazzi’s Hematoxylin. Bars: 50 µm.
P019 Host-Enterobacteriaceae interactions and mitochondrial dysfunction in Crohn’s colitis
Authors: Holani R , Bar-Yoseph H , Krekhno Z , Moon K M , Stacey R G , Serapio-Palacios A , Bressler B , Donald K , Foster L J , Finlay B B 
Published: January 2023

Abstract

Background
Crohn’s disease (CD) is characterized by chronic gastrointestinal inflammation and unfavourable changes in the composition of intestinal microbiota (‘dysbiosis’). Members of Enterobacteriaceae such as K. pneumoniae and E. coli are often over-represented in colonic mucosa of CD patients. These bacteria contribute to vicious circle of dysbiosis and inflammation by expressing multiple virulence factors. Further, mitochondrial dysfunction in Paneth cells has recently been reported in ileal CD. In agreement, high-fat diet and repeated use of antibiotics increases susceptibility to colitis by damaging mitochondria. However, it remains unknown whether impaired mitochondrial bioenergetics could enhance mucosal colonization by Enterobacteriaceae and thus, contribute to the pathophysiology of colonic CD. Furthermore, the molecular mechanism leading to mitochondrial damage in colonic CD are incompletely understood.

Methods
Right colonic biopsies were collected from 74 adult CD patients and 28 healthy participants at St. Paul’s Hospital (Vancouver, BC, Canada). Microbes in colonic biopsies were characterized by 16S rDNA sequencing using Illumina MiSeq platform, followed by analysis of differentially abundant families via ANCOM-BC methodology. Further, high-throughput data-independent acquisition (DIA) proteomics was performed in colonic biopsies (Bruker Daltonics TimsTof Pro2 LC-MS/MS), and later analyzed via DIA-neural network software. Spearman's rank correlation approach was used for host proteomics- Enterobacteriaceae correlational analysis. Microbiome and proteomic data were corrected for false discovery rate (FDR; &lt; 0.5 for statistical significance). For validation studies, human colonic TC7 epithelial cells and patient-derived organoid cultures were infected with CD-patient derived E. coli ± C75, a chemical inhibitor of fatty acid synthase (FASN; de novo fatty acid synthesis enzyme).

Results
We found an increased abundance of Enterobacteriaceae members in colonic mucosa of CD patients compared to healthy controls (Fig A). Next, our proteomics data revealed a significant downregulation in a group of proteins which were functionally related to mitochondrial health and activity, in CD patients compared to controls (Fig B). Further, Spearman’s analysis showed a positive correlation between Enterobacteriaceae and host FASN enzyme (Fig C), a protein directly related to mitochondrial metabolism. Interestingly, we found that blocking FASN with chemical inhibitor (C75) significantly enhanced colonization by E. coli in TC7 cells (Fig D).

Conclusion
Mechanistically, our study reports that aberrant activity of host FASN enzyme may cause mitochondrial dysfunction and thus, facilitates intestinal colonization by Enterobacteriaceae in CD patients.