bluuebunny/crossref_metadata_2025 huggingface dataset:
P0062 Circulating immune cell populations as predictive markers of vedolizumab response in Crohn disease
Authors: Soleto I , González-de Castro C , Arribas E , Ramirez C , Baldan-Martin M , Riestra S , Lucendo A J , García M J , Martín-Arranz E , De Prado Á , Casanova M J , Chaparro M , Bernardo D , Gisbert J P ,   *Authors share co-senior authorship. $ These authors contributed equally to this work
Published: January 2025

Background

Crohn's disease (CD) is a chronic inflammatory condition of the gastrointestinal tract, with a globally rising incidence. It may result from a complex interaction among genetic predisposition, environmental influences, and alterations in gut microbiota, leading to dysregulation of both innate and adaptive immune responses. Despite advances in understanding CD, the precise immunological mechanisms underlying this disease remain unclear. Currently, biological therapies, such as vedolizumab, are among the most effective options for inducing clinical remission and controlling inflammation in CD. However, approximately two-thirds of patients do not respond adequately to these treatments. Consequently, it is essential to identify immune cell populations that can predict clinical responses to vedolizumab, facilitating more personalized treatment selection for each patient.

Methods

We characterized peripheral blood mononuclear cells (PBMCs) using spectral flow cytometry with a panel of 40 markers in patients with CD, both before and after 14 weeks of treatment with vedolizumab. Patients were categorized as responders or non-responders based on their SES-CD.

Results

At baseline Hierarchical phenotypic analysis revealed a total of 39 unique subsets of immune cells within PBMCs (Figure 1). The main populations included T cells (66%), monocytes (0.25%), B cells (12%), and natural killer cells (11%). Within the T cell group, CD4+ cells were twice as prevalent as CD8+ cells. In our evaluation of conventional dendritic cells percentages, we noted an increase in this subset within the responder group, contrasting with the trend observed in macrophages. Furthermore, an unsupervised analysis applying dimensionality reduction algorithms identified 59 clusters (Figure 2), with 37 of them showing differences when comparing responder and non-responder patients. Interestingly, no differences were observed between pre-treatment and post-treatment samples.

Conclusion

This study presents the first immunophenotyping analysis in CD patients, comparing responders and non-responders to vedolizumab based on SES-CD and highlighting differences between these groups. For the first time, these results reveal circulating immune populations as potential predictors of response to vedolizumab. Although these findings should be validated in a larger cohort to assess their clinical significance in identifying patients most likely to benefit from each therapeutic approach, they open the possibility for changes in clinical practice.
P0066 Enhanced transcription of human endogenous retroviruses and downregulation of TRIM28 in patients with Crohn’s disease and ulcerative colitis
Authors: Ribaldone D G , Bergallo M , Galliano I , Veglio V , Pitoni D , Caviglia G P , Frara S , Anxhela P , Tribocco E , Tovo P A
Published: January 2025

Background

Human endogenous retroviruses (HERVs) represent 8% of the human genome. They originate from ancestral infections of germinal cells. HERVs are no longer infectious, but some retroviral sequences can be activated, and their aberrant expressions have been implicated in a number of diseases, including chronic inflammatory and autoimmune disorders. HERV transcription is regulated by TRIM28 and SETDB1, which are directly involved in epigenetic processes and in the modulation of the immune response. HERVs and TRIM28/SETDB1 expressions have not been investigated in patients affected by inflammatory bowel disease (IBD).

Methods

We assessed, through a PCR real-time Taqman amplification assay, the transcription levels of pol genes of HERV-H, -K, and -W families, of env genes of syncytin 1 (SYN1), SYN2, and HERV-W, as well as of TRIM28 and SETDB1 in whole blood of 48 patients with Crohn’s disease (CD), 20 with ulcerative colitis (UC) and in age-matched 104 healthy controls (HC).

Results

The transcriptional levels of HERV-H-pol (p=0.0003) and HERV-K-pol (p=0.001) were significantly higher in IBD patients as compared with HC, while no significant differences between the two groups were found for the remaining HERVs (Figure 1).

The median mRNA levels of HERV-H-pol in CD was 1.53 (1.22–2.02), in UC was 1.56 (0.96–1.99), in HC was 1.24 (0.63–1.63); median HERV-K-pol in CD was 1.27 (1.00–1.76), in UC was 1.41 (0.99–1.85), in HC was 0.97 (0.71–1.34).

The mRNA levels of TRIM28 were significantly downregulated in IBD patients (median mRNA levels of TRIM28 in CD = 0.73, 0.56–0.92, in UC = 0.76, 0.60–0.92, in HC = 1.01, 0.79–1.25, p < 0.001), while the SETDB1 levels were preserved (p = 0.95) (Figure 2).

Conclusion

Over-expressions of HERVs and impaired transcription of TRM28 in patients affected by CD or UC suggest that they might be main actors in the pathophysiology of IBD and open the way to innovative targeted interventions.
P0068 Distribution of α4β7 integrin and MAdCAM-1 expression according to the location of intestinal inflammation in a murine model of spontaneous chronic enterocolitis
Authors: Martinez Lozano H , Puerto Cantero M , Ruiz-Martín C , Miranda-Bautista J , Marín-Jiménez I , Menchén Viso L
Published: January 2025

Background

The selective blockade of α4β7 integrin is a treatment for inflammatory bowel disease (IBD), with different efficacy according to the location of inflammation along the gastrointestinal tract. A more pronounced effect has been reported in patients with predominant colonic inflammation. It may be attributed to differences in the regional expression of adhesion molecules. This study aimed to investigate the expression of α4β7 integrin and mucosal addressin cell adhesion molecule-1 (MAdCAM-1) in colon, ileum, and mesentery, using IL-10 knockout (KO) mice as a model of spontaneous chronic enterocolitis.

Methods

At 20 weeks, the colon, ileum, and mesentery were extracted from IL-10KO and C57BL/6 wild-type (WT) mice. Protein levels of α4β7 integrin and MAdCAM-1 were determined by Western blot in the different tissues. Additionally, pro-inflammatory cytokines (TNF-α and IFN-γ) and epithelial adhesion molecules (Zonula Occludens-1, ZO-1; E-cadherin and Mucin 2, Muc2) expression in the ileum, as well as adiponectin expression in the mesentery, were analyzed using real-time PCR.

Results

Analysis demonstrated a significant increase in gene expression of the pro-inflammatory cytokines TNF-α and IFN-γ in the ileum of IL-10 KO mice compared with WT mice (p<0.01). In addition, IL-10KO mice exhibited reduced expression of ZO-1, E-cadherin, and Muc2 in the ileum relative to WT mice (p<0.001). IL-10 deficiency also led to a marked decrease in adiponectin expression in the mesentery (p<0.05). We then analyzed the protein level of MAdCAM-1 and α4β7 integrin according to anatomic region. Remarkably, the protein analysis revealed significantly higher levels of MAdCAM-1 (p<0.01) and α4β7 integrin (p<0.05) in the colon of IL-10 KO mice compared to WT. In the ileum, while MAdCAM-1 levels remained unchanged, α4β7 integrin expression was increased in IL-10KO mice (p<0.01). No significant differences in MAdCAM-1 or α4β7 integrin levels were observed in the mesentery between IL-10KO and WT mice

Conclusion

IL-10-deficient mice develop inflammation in the ileum, accompanied by a reduction in adiponectin production in the mesentery. In this setting, both α4β7 integrin and MAdCAM-1 showed increased expression in the colon, which was not observed in the ileum or mesentery, despite the inflammation. This differential regional expression pattern of adhesion molecules may explain the variation in the efficacy of anti-integrin therapies. Characterizing these patterns in patients with IBD could help identify those who would benefit most from these treatments.
P0072 Unveiling mesenteric and intestinal changes in Crohn’s disease: insights into fibrosis from surgical specimens
Authors: Joo Y H , Park Y , Song E M , Choe A R , Byeon J R , Noh G T , Kim H S , Choi E , Byun J , Park J , Ahn Y H , Jung S A
Published: January 2025

Background

Recent studies have emphasized the role of mesenteric changes, including creeping fat, alongside intestinal alterations in the progression of Crohn's disease (CD). This study aims to investigate intestinal fibrosis and mesenteric changes using tissues obtained from CD patients who underwent surgery for complications such as strictures and perforations.

Methods

From September 2023 to September 2024, intestinal (CD-Inflamed-Bowel/CD-Uninflamed-Bowel) and mesenteric tissues (CD-Inflamed-Mes/CD-Uninflamed-Mes/ CD-Uninvolved-Mes) were collected from nine CD patients who underwent surgery. Additionally, intestinal (Normal-Bowel) and mesenteric tissues (Normal-Mes) were obtained from one patient who underwent anterior resection for adenoma. Specimens were obtained from the small intestine (4 patients) and the colon (5 patients). Samples were analyzed for morphological changes (H&E staining), collagen deposition (Masson’s trichrome), and fibrosis marker expression (qPCR).

Results

Preoperative CT or MR enterography revealed the presence of creeping fat in all nine CD patients, with five showing strictures accompanied by upstream bowel dilatation. In intestinal tissues, increased collagen fiber deposition was observed in CD-Inflamed-Bowel compared to CD-Uninflamed-Bowel, whereas mesenteric tissues showed no significant differences in collagen deposition among Uninvolved, Uninflamed, and Inflamed groups. qPCR analysis of intestinal tissues revealed significantly higher expression of the fibrosis marker COL1A1 in CD-Inflamed-Bowel compared to Normal-Bowel and CD-Uninflamed-Bowel (119.9 ± 260 vs. 1.0 ± 0.0 vs. 1.0 ± 0.7, P = 0.041). In mesenteric tissues, COL1A1 expression was significantly elevated in CD patients compared to controls (28.0 ± 7.7 vs. 1.0 ± 0.0, P = 0.002), though no significant differences in COL1A1 expression were found among Uninvolved, Uninflamed, and Inflamed mesenteric tissues within the same patient (17.7 ± 3.7, 25.1 ± 9.2, 41.0 ± 21.2, P = 0.9372). Other fibrosis markers (Vimentin, TGF-β, MMP-1, CHI3L1, CXCL1, and PDPN) showed similar trends.

Conclusion

This study highlights inflammatory and fibrotic changes in the intestine and mesentery during CD progression. Notably, mesenteric fibrosis was evident even in uninvolved intestinal segments without active inflammation, suggesting mesenteric inflammation and fibrosis may precede intestinal involvement. These findings support the mesentery as a novel potential target for early treatment for disease progression of CD.
P0074 Colonic goblet cell dysfunction and gastric intestinal metaplasia in patients with anterior gradient 2 defects and infantile Inflammatory bowel disease
Authors: Azabdaftari A , Al-Shaibi A , Thomas T , Jainarayanan A , Capitani M , Verleur N , Agarwal D , Dooley K , Rich-Griffin C , Provine N , Klenerman P , Elawad M , Travis S , Buckley C , Dendrou C , Lo B , Uhlig H
Published: January 2025

Background

Biallelic anterior gradient 2 (AGR2) loss-of-function variants cause infantile-onset inflammatory bowel disease, alongside histological changes of the gastrointestinal epithelium. The underlying mechanisms on epithelial development and function are not understood.

Methods

We conducted single-cell RNA-sequencing of gastrointestinal biopsies and organoids from AGR2-deficient patients. Organoids were used for functional investigation of the epithelium and mucus layer by flow cytometry, confocal microscopy, and using a microfluidic organ-chip. Multiplexed imaging was used to characterise intestinal metaplasia in the antrum.

Results

AGR2-deficiency caused region-specific changes in epithelial cell populations. In the gastric antrum, AGR2-deficiency led to defective epithelial cell differentiation into premalignant metaplastic enterocytes associated with an up-regulation of gastric cancer markers. In the colon, despite a histological depletion of goblet cells, single-cell analysis demonstrated an over-abundance of goblet cells in AGR2-deficiency, exhibiting high ER-stress. We identified defective processing of MUC2 and reduced MUC2 storage in goblet cells without signs of crinophagy, suggesting a stress-induced secretion. Similar to AGR2-deficiency, in ulcerative colitis, goblet cells revealed high ER-stress and no reduction in cell numbers.

Conclusion

Our data highlight AGR2-deficiency as a genetic model of precancerous intestinal metaplasia in the gastric antrum. Apparent colonic goblet cell depletion in AGR2-deficiency and in ulcerative colitis is a histological artefact, representing a reduction of stored mucins in goblet cells via stress-induced mucus secretion.
P0079 Serum calprotectin [cpa9-hne], a biomarker of neutrophil activity, is associated with histological inflammation and fibrosis burden
Authors: Sorokina Alexdóttir M , Poulsen A , Tsapanou Katranara T , Buhl Riis L , Pehrsson M , Rasmussen J , Gögeneur I , Burisch J , Karsdal M A , Bay-Jensen A C , Seidelin J B , Mortensen J H
Published: January 2025

Background

Infiltration of neutrophils into the intestinal tissue is a crucial component of the inflammatory response in Crohn‘s disease (CD). CPa9-HNE, a calprotectin fragment released by human neutrophil elastase during NETosis, has been shown to strongly correlate with endoscopic disease activity in inflammatory bowel disease (IBD) and serves as an indicator for pharmacodynamic response. Furthermore, emerging data suggests that neutrophil extracellular traps (NETs) significantly contribute to the pathogenesis of fibrosis. For the first time, we investigated the association of CPa9-HNE with histological grading of both inflammation and fibrosis.

Methods

The cohort included 62 patients with stenotic Crohn‘s disease. Inclusion criterion was the presence of stenotic disease requiring bowel resection, as confirmed by radiographic assessment. Upon surgery, the resected specimens were collected and histologically graded according to transmural fibrosis burden, and inflammation (D'Haens score). Serum was drawn at baseline (before resection). CPa9-HNE was measured, reflecting a human neutrophil elastase-derived fragment of calprotectin. Spearman’s rank correlation was applied.

Results

CPa9-HNE showed an overall positive correlation to the D'Haens score (ρ=0.279, p<0.005). Significant positive correlation was observed between CPa9-HNE and the presence of neutrophils, both in the epithelium and the lamina propria (ρ=0.309, ρ=0.349, p<0.005 both), as well as with epithelial damage (ρ=0.262, p<0.005). We also observed a positive correlation between CPa9-HNE and transmural fibrosis burden, specifically fibrosis in the submucosa (ρ=0.294, p<0.005)

Conclusion

This study showed that CPa9-HNE, a biomarker of neutrophil activity, correlated with the presence of neutrophils in the intestinal tissue, as well as epithelial damage. Interestingly, CPa9-HNE also showed association to submucosal fibrosis, emphasizing the importance of considering neutrophil activity when targeting the fibro-inflammatory axis. These findings highlight the potential use of CPa9-HNE as a complementary tool for histological grading of inflammation and fibrosis burden, and could be used to further explore the role of neutrophil extracellular traps in intestinal fibrosis.
P0081 Novel in vitro protocol to isolate and expand murine polyclonal Treg cells for efficacy testing in murine models of IBD
Authors: Bretin A , González-King Garibotti H , Benonisson H , Preti M , Peris E , Burgueno J , Vaithilingam V , Häll M , Brännud I , Kurhe Y , Aldridge J , Jennbacken K , Nilsson G , Piras A , Dazzi F , Kozlowska A , Borde A , Birrell M , Preite S
Published: January 2025

Background

Regulatory T cells (Treg) are a subset of CD4+ T cells that play a crucial role in maintaining immune tolerance and preventing autoimmunity. In the context of Inflammatory Bowel Disease (IBD), dysregulation of Treg function has been implicated in the pathogenesis of the disease. Studies have shown that polyclonal Treg have the potential to suppress inflammatory responses and ameliorate colitis in preclinical models. This has led to a growing interest in harnessing the therapeutic potential of polyclonal Treg cells as a potential treatment for IBD. To support this new research and effectively produce Treg, we prompted the in-house development of a new platform to isolate, in vitro expand, and evaluate murine regulatory T cells in mouse models of IBD.

Methods

We used a two-step purification method using a pre-enrichment of CD4/CD25/CD62L-postive cells followed by FACS sorting and alternative cell culture methods to establish a protocol for the scalable isolation and expansion of naïve murine polyclonal Treg (muTreg). We then assessed the efficacy of the muTreg in the CD45RBhigh naïve CD4+ T cell transfer model of colitis.

Results

Quality check of the isolated muTreg showed a high purity that was sustained over a 7-day in vitro expansion period (over 96% FOXP3+) with an over 10-fold expansion. muTreg also conserved their functional activity and showed a 70% suppression of naïve T cells in vitro. Upon administration into the T cell transfer model, muTreg led to a dose-dependent improvement of colitis, as demonstrated by a decrease of inflammation markers such as colon weight, pro-inflammatory cytokine/chemokines, and colon histopathological score. muTreg also showed a strong, dose dependent suppression of effector CD4+ T cells in vivo despite a significant loss of FOXP3 expression under inflammatory conditions (over 60% loss of FOXP3 expression).

Conclusion

In summary, we successfully developed a novel platform for isolating and expanding muTreg and demonstrated their ability to alleviate T cell-induced colitis in a dose-dependent manner. Thus, our preclinical data lay the foundations for the investigation of the effect of Treg therapies in pre-clinical models and pave the way to study antigen specific CAR Treg cells or CRISPR-edited Treg in murine in vivo IBD models.
P0082 Proteomic profiling of colonic mucosa in patients with ulcerative colitis: detection of molecular patterns of inflammation.
Authors: Iakovlev A A , Nukhova S , Tarasova G , Volkov A , Zubova A , Bashtovaya O , Piskovets V
Published: January 2025

Background

The mechanisms of recurrence and progression of ulcerative colitis (UC) require detailed study. Modern advances in proteomic analysis methods are ideal for studying the molecular characteristics of inflammation in the colonic mucosa (CM) of UC patients.

Methods

The study was comparative cohort with parallel design and included 88 patients (range from 22 to 35 years, 37 men and 51 women): 53 (60.2%) pancolitis and 35 (39.8%) left-sided UC, mild and moderate activity. The duration of UC was 5.3 years. The control group included 30 healthy individuals. Biosamples of CM in patients with UC in the active stage and in healthy persons were received by ileocolonoscopy with CM biopsy. The separation of individual proteins of CM was based on technologies of IEF, SDS-PAGE, 2DPAGE, by standard sets (MB-HIC C8 Kit, MB-IMAC Cu, MB-Wax Kit, «Bruker», USA). Automated mass spectrometry imaging was performed by MALDI-TOF-MS/MS, Ultraflex II, «Bruker», USA). The partially identified sequences were then submitted to “BLAST protein-protein” and screened against the Homo sapiens Swissprot database to check if this identification matched the MASCOT-identification (Matrix Science, UK). The data of the molecular interactions and functional features of proteins were received with STRING 10 database.

Results

Based on the data of standard methods of identification of UC and molecular phenotyping of colon mucosa we conducted new prognostic markers, molecular pathways of UC and diagnostic tests in patients with UC. We identified functional groups of peptides and proteins in molecular patterns of CM (fig. 1).

SMAD family member 2 (SMAD2) activates the transcription of TFG1β, that leads the development of fibrosis in colon submucosa in UC patients; the stimulation of the expression of apoС-III in affected CM in UC is associated with the activation of the FOXO1 signaling pathway that supports inflammatory processes; caspase 8 protects colonocytes from TNFα-induced cell death through a necroptosis mechanism via the blockade of the RIP3 expression. Significant decrease of the expression of PPARγ promotes the activation of STAT and AP-1 signaling pathways, that promotes the increase of the synthesis of IL-2, 6, 8, 12,TNFα, matrix metalloproteinases, the activity of immune and inflammation processes in colon mucosa. The reduction of expression of β-defensin-1 in cells of colon mucosa are accompanied with increased expression of CCR6, that promotes the formation of inflammatory infiltrates in colonic submucosa in UC.The molecular interactions of β-defensin-1 are presented on figure 2.

Conclusion

Bioinformatics analysis revealed the presence of the new potentially markers of the progression UC, that’s may provide for the development of novel diagnostic tests for UC.
P0083 Decisional needs among patients and physicians in Crohn's disease: A qualitative analysis
Authors: Cloots M , Hendrix E M B , Okegunna Y , Mingels S M B , van de Koppel I , Mujagic Z , Jeuring S , Dijkstra G , Duijvestein M , Römkens T E H , Dekker A L A J , Pierik M J , Fijten R R R
Published: January 2025

Background

Shared decision-making (SDM) is a collaborative process between patients and healthcare professionals (HCPs) that is essential to align treatment choices with patient values. The increasing variety of treatment options for Crohn’s Disease (CD) challenges the assessment and communication of risks and benefits for both patients and HCPs. Patient decision aids (PDAs) have been shown to be effective in enhancing patient-centered care across various decision-making contexts. This study aims to identify the informational needs of both patients and HCPs to develop an effective PDA for CD.

Methods

Qualitative semi-structured interviews were conducted with 7 CD patient representatives and 5 HCPs. Purposive sampling was used for recruitment of respondents. An empirical phenomenological approach was taken in the interviews to describe similarities and differences of experiences, needs, and preferences of CD patients and HCPs. Interviews were conducted face-to-face or digitally, audio recorded, and transcribed verbatim. Transcripts were thematically analysed through consecutive open, axial, and selective coding.

Results

Preliminary results showed most patients were not actively involved but rather advised in their treatment decisions. Physicians typically preferred a particular treatment but regularly offered patients choices between similar medication options if they wanted more involvement. The primary concerns for patients were the reduction of medication related physical, mental, and social complaints that they often experienced from side effects, form of administration, and fear of surgery. This was reflected in the patients’ need for general disease information, treatment pros and cons, and patient experience stories. Physicians acknowledge that patients wish to be well-informed and believe they provide adequate information. However, many patients express feeling under-informed and frequently look for additional resources. Both groups find the idea of a PDA offering general and personalised information at varying stages in the treatment journey appealing, though physicians caution that personalised content may not be suitable for all patients.

Conclusion

This study highlights a barrier between the approach taken by physicians and patients’ desire for information and involvement in treatment. Providing a reliable and accessible source of information at key decision moments throughout the patient journey could help resolve this barrier. A well-designed PDA for CD patients with reliable information could empower them with confidence and support SDM. However, careful implementation is necessary to accommodate diverse patient needs and preferences.
P0084 Secreted inflammatory protein profiles of human inflammatory bowel disease explants following treatment ex-vivo with licenced biologic therapies and patients’ in-vivo response
Authors: Corcoran R , O'Connell F , Schellenberg N , O'Sullivan J , Kevans D
Published: January 2025

Background

Patient-derived inflammatory bowel disease (IBD) ex-plants have potential for biomarker and therapy discovery. Infliximab (IFX), ustekinumab (UST) and vedolizumab (VDZ) are biologic therapies licenced for the induction and maintenance of remission in ulcerative colitis (UC) and Crohn’s disease (CD). All have demonstrated efficacy in IBD despite differing mechanisms of action. IBD explants have the potential to be used as a precision medicine tool, to gain further insights into disease biology and therapeutic mechanisms of action.

We aimed to evaluate the effect of IFX, UST and VDZ on inflammatory protein secretion profiles in ex-vivo human IBD ex-plants, whilst comparing these explant secretome variations with in-vivo patient data of patients who commenced these therapies in clinic.

Methods

Patients with IBD due to commence in-vivo biologic therapy, undergoing endoscopy, were prospectively recruited. Endoscopic biopsies were collected from the sigmoid colon and IBD ex-plants generated as per previously described methods. IBD explants were then cultured for 24 hours with an IgG control vehicle, IFX, UST and VDZ. After 24 hours, tissue conditioned media (TCM) from IBD explants was collected. TCM secreted inflammatory protein profiles were quantified using 54 V-plex ELISA (Meso Scale Diagnostics, USA). Secreted inflammatory protein profiles were compared between IgG vehicle (control) and UST, IFX, VDZ treated ex-plants. Principal component analysis (PCA) was carried out to characterise the influence of biologics on ex-vivo explant secreted inflammatory profiles. Clinical responses of patients treated with biologics in-vivo were also compared with explants inflammatory profiles.

Results

37 patients with were included (51% CD, 49% UC); age (median, [IQR]) 41[33-53] years, 49% male; disease duration (median, [IQR]) 9 [5-14] years. 23 patients were subsequently commenced on either IFX, UST or VDZ in-vivo and followed over 2 years to determine therapy response. PCA identified significant variation in the CD and UC secretome. Biologic therapy ex-vivo resulted in significant alterations and clustering in the UC secretome compared to control.

Conclusion

The IBD explant model recapitulates expected IBD pathology. These data demonstrate that significant differences between the CD and UC secretome. Treatment with licenced biologic therapies ex-vivo significantly alters multiple pro-inflammatory cytokines, chemokines and secreted proteins in IBD explants. Further study is required to completely understand the effects of licenced biologic therapies on the IBD tissue microenvironment.
P0085 Faecalibacterium prausnitzii induces an anti-inflammatory response and a metabolic reprogramming in human monocytes
Authors: Rolhion N , Danne C , Creusot L , Formiga R , Marquet F , Sedda D , Hua L , Ruffié P , Pham H P , Alonso Salgueiro I , Brot L , Michel M L , Lefevre J , Sokol H
Published: January 2025

Background

Faecalibacterium prausnitzii, a highly abundant bacterium in the human gut microbiota, has been linked to overall health and is notably decreased in pathological conditions, such as Inflammatory Bowel Disease (IBD). F. prausnitzii has shown anti-inflammatory properties in human and mouse models, notably through the induction of IL-10 signaling. Here, we investigated which cell types from human blood and intestinal tissue are responsible for producing IL-10 induced by F. prausnitzii, and providing the first mechanistic insights.

Methods

Immune cells isolated from human blood and intestinal lamina propria of patients with IBD and non-inflamed controls, were stimulated with F. prausnitzii EXL01 strain or Escherichia coli lipopolysaccharide (LPS) and analysed by Legendplex, ELISA, flow cytometry, RNA-sequencing (RNAseq), and Seahorse analyser.

Results

F. prausnitzii EXL01 strain induced the direct and dose-dependent production of IL-10 in CD14+ monocytes from the systemic circulation and intestinal tissue of IBD patients and non-inflamed controls, without inducing a pro-inflammatory response as compared to LPS stimulation. RNAseq analysis corroborated these results and revealed that F. prausnitzii EXL01 strain differentially affects cell energy metabolism compared to LPS. The anti-inflammatory response induced by F. prausnitzii in monocytes was dependent on mitochondrial respiration.

Conclusion

F. prausnitzii induces an anti-inflammatory response and rewires energy metabolism in human monocytes, which might explain its beneficial impact on intestinal inflammation and human health in general. These results provide new insight into the mechanisms underlying the anti-inflammatory effects of F. prausnitzii and are crucial for a better understanding of its potential use in the treatment of IBD. The well-characterised F. prausnitzii EXL01 is already in clinical development in Crohn's disease (NCT05542355), and the results of the first phase 1/2A study involving 8 patients will be available in early 2025.
P0086 The role of enteric glial cells in patients with ulcerative colitis.
Authors: Miyazaki H , Ogino T , Sekido Y , Takeda M , Hata T , Miyoshi N , Uemura M , Mizushima T , Doki Y , Eguchi H
Published: January 2025

Background

Enteric glial cells (EGCs) have traditionally been regarded as supportive cells within the enteric nervous system. However, recent studies indicate that, beyond their involvement in neural circuit formation, EGCs possess antigen-presenting capabilities and actively contribute to mucosal defense. In the mouse myenteric plexus, inflammation-inducing stimuli have been shown to trigger the expression of MHC class II on glial cells, which subsequently influences the differentiation of helper T-cell and regulatory T-cell subtypes, thereby maintaining immune homeostasis under inflammatory conditions. Nevertheless, research on EGCs in humans remains limited. Thus, this study aimed to elucidate the function and involvement of EGCs in the pathogenesis of ulcerative colitis (UC).

Methods

Normal colonic mucosa was obtained from histologically and macroscopically intact areas of patients with colorectal cancer (n=6). Colonic mucosa from UC patients, diagnosed based on established clinical, bacteriological, radiologic, and endoscopic criteria, was also obtained from surgically resected specimens (n=7). Lamina propria mononuclear cells were isolated through enzymatic digestion, and mesenchymal stromal fractions (CD31-CD45-EPCAM-CD90+), containing glial cells, were further isolated using flow cytometry for single-cell RNA sequencing (scRNA-seq) analysis and T cell differentiation assay.

Results

We identified a specific EGC population (Cluster 3: PDPN+ HLA-DR high) with high antigen-presenting capacity, which was selectively elevated at inflamed sites in UC. scRNA-seq analysis, transcription factor activity analysis, and gene-enriched pathway analysis indicated that IFNγ-STAT1 signaling activation within Cluster 3 enhances MHC class II expression. To assess T-cell effects, naive CD4+ T-cells were co-cultured with PDPN+ HLA-DR high cells isolated from UC samples. This co-culture induced differentiation into IL-10+ IL-17A+ CD4+ T-cells, with a positive correlation observed between the proportions of IL-10+ IL-17A+ CD4+ T-cells and PDPN+ HLA-DR high cells. Bulk RNA sequencing and qPCR analysis further demonstrated that PDPN+ HLA-DR high cells highly express the IL-27 subunit EBI3 and the TGF-β activator integrin αvβ8. To evaluate clinical relevance, we also examined the correlation between PDPN+ HLA-DR high cell numbers and clinical factors in 45 UC patients. Immunofluorescence staining for the glial marker CDH2 and HLA-DR revealed a negative correlation between CDH2+ HLA-DR+ cell numbers and preoperative Mayo endoscopic scores.

Conclusion

PDPN+ HLA-DR high glial cells are increased in inflamed areas of UC patients, suggesting a role in anti-inflammatory processes and the maintenance of immune homeostasis.
P0087 Calprotectin contributes to refractory Crohn’s disease fistula through dysregulation of epithelial wound healing responses
Authors: Becker M , Koelink P , Ouahoud S , Meisner S , Muncan V , Frei A , Nabozny G , li F , Saris J , Bemelman W , D'Haens G , Buskens C , Wildenberg PhD M
Published: January 2025

Background

Refractory perianal fistula in Crohn’s disease (CD) still present an unmet clinical need with a high disease burden. Despite increasing interest, pathophysiology leading to the refractory phenotype remains unclear, with only epithelial-to-mesenchymal-transitioning (EMT) commonly mentioned as a contributing factor. Using a systematic approach comparing therapy refractory CD fistula, therapy responsive non-IBD fistula and normal rectum, we aimed to differentiate between general healing responses and dysregulation thereof in refractory CD fistula.

Methods

Total RNASeq (n=51), single cell RNASeq (n=31) and spatial transcriptomic (n=8) analysis were performed on the internal opening and tract of CD fistula and idiopathic fistula. Results were validated using immunohistochemistry. Functional studies were performed in human adult organoid cultures and intestinal cell lines.

Results

In the fistula tracts we identified development of a squamous epithelium characterized by expression of WFDC2, keratin 5 and keratin 13. Although some EMT markers were expressed in the squamous epithelium or its intermediates, no activation of a complete EMT program could be detected in any of the tissue types, regardless of the underlying diagnosis. Upstream analysis indicated TNFα, IL6 and TGFβ as inducers of WFDC2+ epithelium, and human adult colon organoids stimulated with these cytokines (but not TGFβ alone) indeed showed a transciptional profile similar to the fistula epithelium, confirming redifferentiation potential. While WFDC2+ epithelium did occur in CD fistula, abundance was strongly decreased compared to non-IBD fistula. In addition, those WFDC2+ cells present had a more pro-inflammatory profile and were morphologically disorganized. Surprisingly, pseudotime analysis indicated epithelial calprotectin (S100A8/9) as a key factor in the aberrant development of WFDC2+ tissue in CD. Normal intestinal epithelium does not express calprotectin, but a mix of cytokines present in fistula induced clear intracellular expression of S100A8/9 in both epithelial cell lines and primary colon organoids. In CD fistula, calprotectin expression was particularly intranuclear, suggesting a transcriptional role. CHIP analysis indeed showed binding of calprotectin to various inflammatory genes and knockdown of calprotectin abrogated expression of these genes supporting a crucial regulatory role.

Conclusion

These data identify redifferentiation of rectal mucosa to a squamous phenotype as a normal damage response during fistula formation. However, in CD patients, transcriptional effects of calprotectin derail this process, resulting in disorganized inflammatory tissue not amenable for surgical closure and contributing to refractory disease.
P0088 3D tissue-level analysis of intestinal T cell dynamics reveals dual effects of etrasimod on recruitment and tissue exit in experimental colitis
Authors: Fontes V , Müller T , Derdau J , Schett G , Neurath M F , Zundler S , Uderhardt S
Published: January 2025

Background

Sphingosin-1-phosphate (S1P) receptor (S1PR) agonists are a novel class of anti-trafficking agents approved for the treatment of ulcerative colitis. These compounds induce T lymphocyte sequestration in secondary lymphoid organs, reducing their availability in peripheral blood and presumably in inflamed intestinal tissue. However, direct evidence for their effects on T cell dynamics within intestinal tissue is lacking. Moreover, while S1PR-dependent mechanisms regulate T cell egress from the gut, it remains unclear whether S1PR agonists modulate this process. This limited understanding of tissue-specific effects presents a crucial barrier to comprehending their full therapeutic mechanism.

Methods

We developed a comprehensive volumetric imaging platform to investigate tissue-level mechanisms of colitis within the complex three-dimensional architecture of the intestine. Using the T cell transfer colitis model, mice were treated with etrasimod or vehicle control. Full-thickness colon samples underwent tissue clearing and whole-mount immunofluorescence for T cell markers. Advanced light-sheet and volumetric confocal microscopy, combined with machine learning-based image analysis, enabled three-dimensional reconstruction of T cell distributions and lymphatic networks, allowing for quantitative histocytometry and spatial mapping of immune cell dynamics.

Results

T cell transfer colitis induced substantial lymphocyte infiltration in the colon, with T cells forming both distinct clusters and diffuse distributions throughout the tissue. The presence of numerous T cells within lymphatic vessels indicated active tissue egress, suggesting that T cell accumulation reflects a dynamic balance between recruitment and exit (Figure 1). Etrasimod treatment not only reduced overall T cell infiltration but also markedly altered their spatial distribution pattern. Notably, we observed a decreased proportion of T cells within lymphatic vessels, coupled with increased accumulation of T cells in proximity to lymphatic structures, indicating a previously unrecognized effect on tissue exit dynamics.

Conclusion

Our advanced volumetric imaging approach revealed novel insights into a potential dual mechanism of etrasimod in inflammatory bowel disease. Beyond reducing lymphocyte recruitment, etrasimod appears to create a tissue exit block, effectively trapping T cells within the intestinal compartment. These findings raise important questions about how enforced tissue residency influences local inflammatory circuits and may guide optimization of therapeutic strategies.
P0094 The Role Of Sdf2l1 On Intestinal Physiology And Inflammatory Bowel Disease
Authors: Thies M , Schuran F A , Mishra N , Pimenta Bernardes J , Hinrichsen F , Rosenstiel P
Published: January 2025

Background

The intestinal tract, as the body's largest mucosal surface, is exposed to various factors, requiring specialized epithelial and immune cell interactions to maintain homeostasis. These interactions, along with a dynamic microenvironment, induce endoplasmic reticulum (ER) stress. While the ER employs coping mechanisms to preserve cellular integrity, unresolved ER stress is a key driver in the development and progression of chronic inflammatory bowel disease (IBD).1 An important player in the ER-resident unfolded protein response is stromal derived growth factor 2-like 1 (Sdf2l1), a chaperone protein that ensures proper protein folding and prevents the accumulation of misfolded proteins.2,3 Furthermore, Sdf2l1 is highly expressed in the intestine, thus we hypothesize that Sdf2l1 plays a pivotal role in the pathogenesis of IBD.

Methods

To investigate the role of Sdf2l1 in the gastrointestinal system, we used wildtype or conditional Sdf2l1 knockout mice lacking Sdf2l1 in intestinal epithelial cells (Sdf2l1ΔIE) and ex vivo organoid systems.

Results

No baseline differences were found in the intestinal mucosa of Sdf2l1-deficient mice, but molecular differences in ER stress responses were evident in small intestinal organoids. When ER stress was induced with IL-22, Sdf2l1ΔIE organoids showed significantly higher NFκB expression, suggesting enhanced UPR activation or compromised stress coping. Under tunicamycin treatment, Sdf2l1-deficient organoids had increased Hspa5/Grp78 expression, indicating elevated ER stress. Sdf2l1 is needed for proper UPR initiation, as it forms a complex with GRP78. In these deficient organoids, IRE1 signaling was impaired, as shown by reduced Xbp1s expression.

Conclusion

Insufficient activation of the UPR due to Sdf2l1 deficiency and the associated ER stress potentially leads to inflammatory reactions in the cells. Impaired cell integrity disrupts intestinal homeostasis and can lead to an imbalance of pro- and anti-inflammatory processes, thus favoring the pathophysiology of IBD.

References

1. Eugene SP, Reddy VS, Trinath J. Endoplasmic Reticulum Stress and Intestinal Inflammation: A Perilous Union. Front Immunol. 2020 Nov 25;11:543022. doi: 10.3389/fimmu.2020.543022. PMID: 33324392; PMCID: PMC7723926.

2. Kim J-S, Mochida K, Shinozaki K. ER Stress and the Unfolded Protein Response: Homeostatic Regulation Coordinate Plant Survival and Growth. Plants. 2022; 11(23):3197. https://doi.org/10.3390/plants11233197

3. Sasako, T., Ohsugi, M., Kubota, N. et al. Hepatic Sdf2l1 controls feeding-induced ER stress and regulates metabolism. Nat Commun 10, 947 (2019). https://doi.org/10.1038/s41467-019-08591-6
P0095 Human multi-lineage gut-on-chip to study near-physiological epithelial barrier function and directed secretion
Authors: Hensel I V , Steinhauer M , Éliás S , Neurath M F , Günther C , Resnik-Docampo M
Published: January 2025

Background

The human intestinal epithelium is composed of diverse cell types, enabling it to fulfil essential digestive functions such as absorption and secretion, while simultaneously acting as an immune interface. This epithelial barrier safeguards underlying immune cells in the lamina propria from luminal antigens by maintaining a mucus layer rich in antimicrobial peptides, preventing unwanted immune responses and allowing regulated cytokine and chemokine gradients. Despite advances in single-cell transcriptomics, understanding specific epithelial cell secretions remains limited.

Methods

To address this, we developed a human intestinal organ-on-chip (OoC) model, derived from human colon organoids, capable of forming a heterogenous, functional epithelial barrier without permeable supports. This OoC model provides concurrent access to luminal and basal compartments, allowing for mid- to high-throughput screenings, overcoming limitations of conventional transwell or 3D organoid cultures.

Results

In our study, we analysed barrier function by measuring transepithelial electrical resistance (TEER) and permeability as well as confirming tight junction assemble marked by ZO-1 and occludin. Cell type composition was modulated using distinct culture media, resulting in a heterogeneous cell composition including goblet cells, colonocytes, and enteroendocrine cells along with the proliferative compartment, evidenced by transcriptomic and protein analyses. Cytokine and chemokine secretions, notably CCL2, CCL20, IL-8, IL-18, and CXCL10, demonstrated cell type-dependent and spatially directed patterns, suggesting location-specific secretion along the crypt-surface axis in vivo. Under basal and stimulated conditions, our model retained responsiveness to bacterial stimuli, gluten-derived antigens, and pro-inflammatory conditions, confirming the important role of epithelial cells in immune response regulation.

Conclusion

Collectively, this OoC model offers a physiologically relevant platform for exploring human intestinal barrier function and epithelial-immune communication. This scalable, multi-lineage in vitro model advances our capacity to study directed secretion, barrier integrity, and epithelial cell type dependent immune response, supporting applications in disease modelling and personalized medicine development.
P0097 STING pathway drives cellular tryptophan degradation and pro-inflammatory responses in intestinal fibroblasts
Authors: Wu Q , Yang G , Xiang H , Shima K , Springer E , Bornhäuser J , Welz L , Aden K ,   AG Aden
Published: January 2025

Background

Tryptophan metabolism is severely disrupted in IBD. Pro-inflammatory cytokines activate the first rate-limiting enzyme IDO1 leading to tryptophan degradation through the kynurenine pathway. Although Trp degradation typically support de novo NAD synthesis via KP activation, evidence suggests a metabolic block at the downstream enzyme QPRT in IBD. STING has been described as inducing the secretion of type I interferons and other inflammatory cytokines, suggesting its important role in regulating intestinal homeostasis. Several studies have reported an association between STING-mediated IDO1 activity and its role in promoting tumour immune evasion. However, the relationship between STING pathway and tryptophan metabolism in intestine has not be understood. This study aims to investigate the effect of STING signaling on tryptophan degradation and explore how the metabolic blockage in tryptophan-NAD axis may influence STING-mediated immune response.

Methods

To investigate the STING pathway's role in tryptophan metabolism in intestine, we utilized human colonic fibroblast cell line CCD-18co. Quantitative RT-PCR and Western blot were employed to measure mRNA and protein expression. Tryptophan metabolism was quantified via LC-MS. siRNA was performed to mediate QPRT knockdown. Intracellular NAD level were assessed by NAD-glo assay. Seahorse assay was conducted to evaluate mitochondrial function.

Results

We screened a range of established IDO1-inducing cytokines (IFNγ, TNFα, etc.) and compared their effects on IDO1 expression with STING agonists diABZI, dsDNA and 2’3’cGAMP. STING agonists induced significantly higher IDO1 expression than any of the cytokines tested. While cultivation of in presence of Anifrolumab (type I IFN receptor antagonist) and Tofacitinib (pan-JAK inhibitor) strongly inhibited STING-induced IDO1 upregulation. LC-MS showed upregulation of IDO1 drive tryptophan into kynurenine pathway, which was again inhibited by Anifrolumab and Tofacitinib. To mimic the reduced QPRT expression in IBD, we performed QPRT knockdown. QPRT deficiency led to STING pathway overactivation upon diABZI treatment, resulting in elevated expression of pro-inflammatory cytokines and phosphorylation of STING and TBK1. QPRT-deficient cells exhibited reduced intracellular NAD level and impaired mitochondrial function. Supplementation with the nicotinamide riboside restored NAD levels, which effectively reduced the elevated cytokine expression caused by QPRT knockdown.

Conclusion

Our data provide a novel link of how STING activation in intestinal fibroblasts drives tryptophan degradation via KP pathway. Additionally, QPRT-related metabolic blockage enhance STING-mediated pro-inflammatory response, which can be mitigated by NAD precursor supplementation