bluuebunny/crossref_metadata_2025_split huggingface dataset:
DOP55 High diagnostic accuracy of 4-probe methylation biomarkers in paediatric Inflammatory Bowel Disease by epigenome-wide analysis
Authors: Noble A , Adams A T , Nayak K M , Kalla R , Ventham N , Cheng G , Ashton J J , Hansen R , Croft N , Wilson D C , Ennis S , Zilbauer M , Uhlig H H , Satsangi J 
Published: January 2024

Abstract

Background
The application of -omics technology offers important opportunities for biomarker discovery to personalise management of patients with inflammatory bowel disease (IBD). In previous work, we reported a characteristic profile of genome-wide DNA methylation in peripheral blood leucocytes from children with paediatric IBD (pIBD) at diagnosis defining the IBD methylome. This characteristic pattern of genome-wide alterations was replicated in inception cohorts of adult patients in UK and Scandinavia.

Methods
Whole blood DNA methylation profiling was performed using the Illumina EPIC array on 86 pIBD patients and 30 non-IBD controls. Patients had a median age of 12 y and were prospectively recruited from gastroenterology clinics in Oxford and Cambridge, UK. In modelling, we utilised the paediatric BISCUIT and PICTS study cohorts from Scotland as our training data. Publicly available data from the RISK paediatric CD cohort from North America was accessed (GSE11261) to further assess accuracy of the model. The model was then subjected to further testing in an Oxford-based paediatric coeliac cohort and adult cohorts with IBD, and rheumatoid arthritis (RA)(Table 1).

Results
Genome-wide methylation changes in the Oxford/Cambridge cohort were highly consistent with the index BISCUIT and PICTS cohorts. Four single methylation sites were selected to make up a diagnostic model involving the genes, RPS6KA2, VMP1, CF1 and ARHGEF3 (Figure 1) in the index cohort and validated in the Oxford/Cambridge cohort. Following receiver operating characteristic (ROC) area under the curve (AUC) analyses the model demonstrated an AUC of 0.912 (95% CI: 0.86-0.96) (Table 1). To further validate our model, we compared pIBD who were CRP-positive (&gt;5 mg/l) and CRP-negative (&lt;5 mg/l) at presentation against non-IBD children. In the CRP-positive group, we found an AUC of 0.99 (95% CI: 0.99-1). Within the CRP-negative group an AUC of 0.90 was observed against controls (95% CI: 0.83-0.96). The model was further validated using methylation data at the baseline timepoint in the RISK cohort, with an AUC of 0.93 (95% CI: 0.90-0.96). In further analyses, we demonstrated specificity for IBD compared with paediatric coeliac disease; and accuracy higher in childhood-onset AUC than adult-onset disease AUC; the model is not accurate in diagnosis of RA.

Conclusion
We confirm a characteristic pattern of DNA methylation changes in childhood-onset IBD; and derive and validate a 4-probe model for diagnosis with high accuracy in both Europe and North America. The model is specific for pIBD compared with symptomatic children with no demonstrable pathology; and children with coeliac disease; and may provide an alternative to current markers in blood and stool, including calprotectin.
DOP62 A core molecular resistome characterised by extracellular matrix dysregulation and neutrophil degranulation predicts resistance to multiple therapies in Ulcerative Colitis
Authors: Saifuddin A , Cozzetto D , Pavlidis P , Hart A , Powell N 
Published: January 2024

Abstract

Background
Several transcript signatures have been published that predict anti-TNF resistance in ulcerative colitis (UC). However, clinical translation is hampered by suboptimal validation performance, partly because of insufficient emphasis on potential pathophysiology. Moreover, predictors of ustekinumab response are lacking. We interrogated transcriptomic datasets and anti-TNF predictive signatures to investigate resistance mechanisms and applicability to ustekinumab.

Methods
Transcript profiles of colonic biopsies taken before treatment for moderate-to-severe UC were analysed from Gene Expression Omnibus. Patients were then treated with infliximab (n = 24 and n = 22), golimumab (n = 84) or ustekinumab (n = 358). Molecular pathways from the Reactome database that were differentially enriched between mucosal healing responders and non-responders post-induction were found, along with shared pathways between cohorts. The largest cohort was used to calculate enrichment of published resistance signatures in actively inflamed mucosa using Spearman correlation. Hierarchical clustering was undertaken for non-responders. Analyses were performed using R 4.2.2 (Vienna, Austria).

Results
Many biological pathways were associated with resistance across the cohorts (1079 pathways, adjusted p value&lt;0.01). Sixty pathways were shared between 3 or 4 datasets. Those relating to extracellular matrix (ECM) activity, neutrophil degranulation and interleukin signalling were most prominent. Network analyses revealed that proteins forming these pathways interact in vivo, forming a discrete functional module. The enrichment of published transcript signatures correlated highly. For instance, correlation between enrichment of UC1 (Czarnewski et al, 2019) and M5 (Friedrich et al 2021) was 0.96. Over-representation analyses showed that these signatures represent processes such as interleukin signalling and neutrophil activity. Hierarchical clustering of non-responders revealed two distinct groups, including those with high (NRhi) and low (NRlo) signature enrichment (Figure 1). Cellular deconvolution analyses showed that neutrophils are more prevalent in NRhi compared to NRlo and that eosinophils may be more frequent in NRlo than NRhi.

Conclusion
ECM dysregulation and neutrophil activity may underlie a core ‘resistome’ that heralds poor treatment outcomes to anti-TNFs and ustekinumab. Supporting this, the enrichment scores of published transcript signatures in inflamed tissue correlate highly, suggesting that many of these represent shared, or similar, molecular mechanisms of resistance. However, there is a separate resistant cohort with different molecular drivers. These findings have implications for precision medicine approaches in UC.
DOP77 Mesenchymal stem cell therapy for refractory Crohn’s perianal fistulas: a case series
Authors: Pronk A , Beek K J , Wildenberg M E , Bemelman W A , Stoker J , Buskens C J 
Published: January 2024

Abstract

Background
Nearly one-third of patients with Crohn’s disease will develop one or multiple perianal fistulas within the first two decades after diagnosis, with the majority being complex. Treatment options are limited with high recurrence rates after both medical and surgical approaches. It has been demonstrated that a completely fibrotic tract on Magnetic Resonance Imaging (MRI) with a MAGNIFI-CD &lt;6 is the best predictor for long-term clinical closure. Mesenchymal stem cell treatment (MST) has emerged as a new therapeutic strategy for these fistulas. The aim of the current study was to analyse the effectiveness of MST for complex Crohn’s perianal fistulas based on MRI.

Methods
Consecutive patients with complex Crohn’s perianal fistulas, previously failing both anti-TNF treatment and surgical closure, that underwent surgical closure of the internal opening with MST between December 2019 and March 2023 were included. All patients underwent a MRI preoperatively and between three to six months after MST. For the current study, MRI’s were read by an senior radiologist. The primary endpoint was radiological remission of the fistula(s) defined as a MAGNIFI-CD &lt;6 on MRI. Secondary endpoints were clinical closure (defined as closure of the external opening(s)), recurrence rate, change of MAGNIFI-CD over time, quality of life based on the perianal disease activity index (PDAI), and serious adverse events (SAE).

Results
In total, 30 patients (16 females) with 48 fistula tracts were included with a median clinical follow-up of 20 months. Radiological remission was achieved in thirteen patients (43.3%) after a median follow-up of 5.0 months (IQR 3.0-6.0). The median MAGNIFI-CD at baseline was 15.0 (IQR 7.0-20.0) which decreased significantly to 8.0 (IQR 3.0-15.0) after treatment (p= 0.001). Clinical closure of the fistula(s) was achieved in 21 patients (70.0%). Three patients (14.3%) developed a recurrence during long-term FU. All three patients had clinically closed fistula(s), but no radiological remission. The median PDAI decreased significantly from 10.5 (IQR 7.0-14.0) to 4.0 (IQR 0.0-7.3) (p= 0.001). Overall, in this patient group one SAE occurred requiring multiple reinterventions and temporary stoma.

Conclusion
Closure of the internal fistula opening with MST is a promising treatment strategy for therapy refractory Crohn’s perianal fistulas, resulting in &gt;40% radiological remission and clinical closure in 70%. No recurrences were seen in patients with radiological remission. MST was also associated with a significant increase in quality of life. Further research is needed to gain insight in which patients MST is most likely to induce radiological remission.
P012 Intestinal epithelial cells as targets of thiopurines in Inflammatory Bowel Disease paediatric patients-derived organoids
Authors: Muzzo A , Lucafò M , Bramuzzo M , Zudeh G , Franzin M , Hofmann U , Lazzari E , Meroni G , Schwab M , Decorti G , Stocco G 
Published: January 2024

Abstract

Background
Thiopurines, azathioprine (AZA) and mercaptopurine (MP), are immunomodulators used to maintain remission in patients with inflammatory bowel disease (IBD). Intestinal epithelial cells play a key role in the pathogenesis of IBD, however, their role as target cells for thiopurines has not been elucidated. The aim of the project is to establish an in vitro IBD model using organoids starting from intestinal biopsies of IBD paediatric patients to investigate thiopurine effects on intestinal epithelium.

Methods
Forty-four IBD patients (mean age 14.4 years, 21 males) were enrolled. Intestinal biopsies were used to isolate crypts and generate adult stem cell-derived organoids. The cytotoxicity of thiopurines (0.2-200 µM for 72 hours) was evaluated by CellTiter-Glo 3D assay. Gene expression was evaluated using TaqMan assays. Thiopurine metabolites were quantified by LC-MS/MS analysis. Proteomic profiles were analyzed by Q-Exactive Plus mass spectrometer, western blot and REACTOME pathway software. Statistical analysis was performed using GraphPad and R software.

Results
IBD patients-derived organoids were sensitive to thiopurine treatment in a dose-dependent manner with a high interpatient variability (Two-way ANOVA p-value&lt;0.0001; IC50 2.24 µM for AZA (95% CI: 1.28–5.53 µM) and 2.65 µM for MP (95% CI: 1.43–8.27 µM)).A significant negative correlation was observed between the viability after thiopurine exposure and mRNA expression of candidate genes involved in thiopurine pharmacokinetics (ITPA, TPMT, PACSIN2 and NUDT15). The most abundant thiopurine metabolite measured on organoids treated with IC50 of MP was MeTIMP, with a concentration correlated with cell viability after MP exposure (p=0.0056, ρ=-0.99). Proteomic analysis on organoids treated with IC20 of thiopurines (0.2 μM) showed 45 differentially expressed proteins in common between AZA and MP (p&lt;0.05). REACTOME enrichment analysis revealed as the most altered pathways: vesicular traffic, autophagy, protein ubiquitination and interferon signalling, while no proteins related to apoptosis were induced. Among these proteins, TRIM32, a ubiquitin ligase involved in STING activity, a key regulator of both interferon I expression and NF-kB activation, resulted downregulated by the treatment (t-test AZA: p=0.0006; MP: p=0.014) and its basal levels negatively correlated with viability rates after thiopurine exposure (AZA: p=0.006, ρ=-0.85; MP: p=0.002, ρ=-0.88). Organoid exposure to thiopurines also decreased p-STING/STING ratio, particularly after MP treatment (t-test p=0.025), and increased autophagy, assessed as LC3-II levels (t-test AZA: p=0.0006; MP: p=0.014).

Conclusion
Organoid cultures provide new insights on the mechanism of action of thiopurines in the intestinal epithelium.
P014 First-time-in-human visualisation of CCR9 expression in the gut by positron emission tomography
Authors: MacKay J , Jucaite A , Cselényi Z , Johnström P , Dahl K , Schou M , Stenkrona P , Sousa V , Cortes Gonzalez M , Giffen P , Bouma G , Nys J , Brülls M , Duncan M , Larsson J , Lundahl A , Les K , Bergare J , Elmore C , Kiraga J , Blé F X , Fowler A , Belvisi M , Marks D 
Published: January 2024

Abstract

Background
CCR9/CCL25 is the key axis for recruiting lymphocytes to the small intestine and represents a potential treatment target in Crohn’s disease (CD). AZD7798 is a first-in-class monoclonal antibody for CD that depletes CCR9+ lymphocytes. Monitoring CCR9+ cell depletion and repletion in the blood is straightforward, but difficult in the primary tissues of interest (small bowel, mesenteric lymph nodes) which require invasive procedures to access. For AZD7798, an understanding of pharmacodynamics in gut is crucial to guide further clinical development. To address this need, we have developed a CCR9-specific PET tracer based on the small molecule CCR9 antagonist vercirnon.

Methods
Specific binding of [3H]vercirnon was assessed in-vitro using autoradiography. Specific binding and dosimetry of [11C]vercirnon was assessed in-vivo in cynomolgus monkeys.
Subsequently, a first-time-in-human study (NCT05818514) was performed in healthy volunteers to assess the biodistribution, binding to CCR9 in abdominal regions of interest, and safety of [11C]vercirnon, administered intravenously at microdoses (&lt;10μg). Two PET-CT examinations were performed 14 days apart to assess reproducibility.

Results
In-vitro specific binding of [3H]vercirnon to CCR9 was confirmed in human thymus sections and CCR9-expressing cell pellets.
In-vivo specific binding of [11C]vercirnon to CCR9 was putatively demonstrated in the gut of non-human primates (NHPs) in pre-treatment/displacement studies with excess unlabelled vercirnon (20mg/kg).
8 healthy volunteers (5 male) were enrolled in the FTIH study. Detectable intestinal uptake of [11C]vercirnon was demonstrated in all participants (Figure 1) with mean±SD standardised uptake value ratio (SUVR) 1.4±0.6 (reference abdominal aorta), and a proximal to distal gradient (SUVR jejunum 1.8±0.7/ileum 0.9±0.4/colon 0.6±0.2) consistent with CCR9 gene expression data. Test-retest variability of intestinal uptake was 9.6% (intraclass correlation coefficient 0.96), in line with values for other clinically useful PET tracers.
Mean±SD injected radioactivity per PET examination was 351±105 MBq, corresponding to a radiation exposure of 1.4 mSv. This low radiation exposure (c.f. conventional FDG PET ~9 mSv) allows multiple PET examinations per participant. Administration of [11C]vercirnon was well-tolerated with no serious adverse events.

Conclusion
We have developed a CCR9-specific PET tracer which potentially enables repeatable quantification of CCR9 expression in the gut. Ongoing studies will assess the effects of AZD7798 on CCR9+ cell depletion/repletion in the gut, providing critical data to support the clinical development programme.
P027 Involvement of PD-L1 in myenteric plexitis associated with postoperative recurrence of Crohn's disease
Authors: Le Berre C , Durand T , Clavreul M , Neunlist M , Bourreille A , Neveu I , Naveilhan P 
Published: January 2024

Abstract

Background
Despite a growing therapeutic armamentarium, post-operative recurrence (POR) in Crohn’s disease (CD) remains frequent. The presence of myenteric plexitis at the proximal margin of ileocolonic resection has been recognized as a risk factor for early POR. They are defined by the accumulation of immune cells in contact with the ganglia of the myenteric plexus.
In preliminary work, we have shown an increase in the number of T lymphocytes (TL) in contact with enteric glial cells (EGC) in patients with CD depending on ICAM-1/LFA-1 pathway. As CD is classically associated with a Th1 immune response, the aim of this work was to study the impact of EGC/CD4+ TL contacts on Th1-type cytokine production (IFNγ and its transcription factor Tbet) and to identify molecules that may be involved in these regulations.

Methods
In vitro in rats, the impact of EGC/CD4+ TL co-culture on IFNγ production was studied by ELISA and qPCR of the transcription factor Tbet. The expression and regulation of PD-L1, a candidate molecule, was then analyzed in murine and human EGC by qPCR and immunohisto/cytochemistry.

Results
While EGC alone produced only a small amount of IFNγ, there was a significant reduction in IFNγ production in EGC/CD4+ TL co-cultures compared with CD4+ TL alone. Yet the proportion of CD4+ TL cells capable of producing IFNγ was significantly increased in contact with EGC, as was their survival. In parallel, Tbet was downregulated in CD4+ TL in contact with EGC. These results suggested the involvement of PD-L1, a molecule known to negatively regulate cytokine production (notably IFNγ) and Tbet expression.
Expression of PD-L1 mRNA and its protein was demonstrated in murine and human EGC and was upregulated under pro-inflammatory conditions. Analysis of transmural sections at the proximal margin of ileocolonic resection from patients (n=15 controls, n=14 CD without POR, n=15 CD with POR) showed PD-L1 expression in myenteric ganglia that was restricted to EGC (no neuronal expression) (Figure 1A). PD-L1 fluorescence intensity in myenteric ganglia at the proximal resection margin was significantly lower in CD patients with POR than in controls (∆13.5%; p=0.012) (Figure 1B).

Conclusion
Lower IFNγ production in EGC/CD4+ TL co-cultures associated with reduced Tbet expression by TL in contact with EGC suggest a role of PD-L1 in myenteric plexitis. PD-L1 expression has been demonstrated in murine and human EGC, with significantly lower expression in myenteric ganglia located at the proximal resection margin of CD patients with POR compared with controls. Blocking the ICAM-1/LFA-1 pathway and/or restoring the PD-1/PD-L1 loop could be of interest for developing strategies to prevent and/or treat POR in CD patients.
P035 3D bioprinting of gelatin derivatives: towards novel small intestinal in vitro models
Authors: Maes L , Szabó A , Van Haevermaete J , Geurs I , Dewettinck K , Van Vlierberghe S , Laukens D 
Published: January 2024

Abstract

Background
Despite significant progress achieved in the field of inflammatory bowel disease research, the precise cause of the disease has yet to be elucidated. Intestinal in vitro models can provide a fast and inexpensive alternative for in vivo studies. Nonetheless, many in vitro models lack the crypt or crypt-villus architecture as seen in the large and small intestine respectively and therefore fail to mimic the complexity of the gut. In this study, we aim to improve the physiology of small intestinal in vitro models by the development of hydrogels that mirror the villi and crypts of the digestive tract.

Methods
This study focused on the development of gelatin-methacryloyl-aminoethyl-methacrylate (gel-MA-AEMA)-, and gelatin-methacryloyl-norbornene (gel-MA-NB)-based biomaterial inks to fabricate 3D constructs, mimicking villi or a combination of villi and crypts, with digital light processing. The constructs were mechanically and morphologically evaluated with parallel plate rheology and cryo-scanning electron microscopy respectively. To assess the biocompatibility of the 3D constructs, a Caco-2/HT29-MTX co-culture in a 9:1 ratio was maintained for 21 days and confluency was confirmed with immunofluorescence. Gene expression, transepithelial electrical resistance (TEER) and paracellular permeability of the cells cultured on the constructs were compared to cells cultured on flat gel-MA-AEMA and gel-MA-NB hydrogels, a collagen type I coating or uncoated tissue culture plastic.

Results
Both gel-MA-AEMA and gel-MA-NB hydrogels exhibited physiologically relevant stiffness (1.9 ± 0.63 kPa and 1.64 ± 0.63 kPa respectively), but only the gel-MA-AEMA based biomaterial ink could be successfully utilized for printing constructs with villi and crypts. On all construct designs, confluency was reached and paracellular permeability of small sized marker molecules in combination with TEER measurements suggested the formation of a functional barrier over time, which was further confirmed by immunofluorescence and increased gene expression of tight junction proteins, occludin and ZO-1. The gene expression of enterocyte differentiation markers, villin-1, sucrase isomaltase and alkaline phosphatase, suggested the superior differentiation of Caco-2 cells on the ‘only villi’ and ‘villi and crypts’ constructs compared to flat hydrogels, collagen type I coating or uncoated tissue culture plastic.

Conclusion
Although both hydrogels promoted functional barrier formation and enterocyte differentiation, gel-MA-AEMA was more suited for DLP than gel-MA-NB. In addition, culturing intestinal epithelial cells on the 3D constructs ameliorated cell differentiation compared to conventional 2D setups.
P051 Diets enriched in fat and sugar and sex-dependant response to colitis in a mouse model
Authors: Touly N , Arnone D , Caron B , Kökten T , Umoret R , Peyrin-Biroulet L , Boudaud M 
Published: January 2024

Abstract

Background
Inflammatory bowel diseases (IBD) are chronic diseases characterized by inflammation of the mucosa linked to an alteration of the intestinal microbiota and overreaction of the immune system. The prevalence of IBD is increasing in industrialized countries where we find a “Western Diet” rich in refined sugars and saturated fats but low in fiber and essential unsaturated fatty acids1. Thus, nutrition seems to be an important environmental factor involved in the occurrence of IBD, however, underlying biological mechanisms are still poorly understood2. Additionally, most studies on colitis focused on males, yet diets effects are often very different according to sex. The aim of this study was therefore to study the sex-dependent effect of a diet enriched in sugar (sucrose) and a diet rich in fat in a mouse model of colitis.

Methods
Male and female, 9-weeks old C57BL/6 mice were fed 3 diets for 8 weeks: control diet (C), high-fat diet (HF), and high-fat/high-sucrose diet (HFHS). Then, colitis was induced using 3% Dextran sodium sulfate (DSS) for 5 days and kept 5 more days without DSS. The animals were monitored and weighed daily to establish a disease activity index (DAI). Blood and stools were collected monthly from the start of feeding. At the end of the protocol, a colonoscopy was carried out for each group before sacrifice and intestinal tissues were collected.

Results
DSS-induced colitis was earlier and more severe in both males and females fed high-caloric diets (both HF and HFHS). However, the HFHS diet accelerated DSS-induced colitis in males compared to the HF diet, while the HF diet led to more severe DSS-induced colitis than the HFHS diet in females. Males and females had similar weight gain following the high-caloric diets.

Conclusion
This difference in response to DSS after consumption of high-caloric diets between males and females indicates that the inflammatory mechanisms seem to differ depending on sex. Recent work reports the same sex-dependent effect in the Il10-/- mouse model3, suggesting that the sex has an impact on the pathophysiology of colitis and is essential to consider when studying IBD.

References
1. Arnone D, et al. Sugars and Gastrointestinal Health. Clin Gastroenterol Hepatol. 2022 Sep;20(9):1912-1924.e7.
2. Arnone D, et al. Long-Term Overconsumption of Fat and Sugar Causes a Partially Reversible Pre-inflammatory Bowel Disease State. Front Nutr. 2021 Nov 18;8:758518.
3. Maite CB, et al. The Effect of Sex-Specific Differences on IL-10-/- Mouse Colitis Phenotype and Microbiota. Int J Mol Sci. 2023 Jun 20;24(12):10364.