Introduction

Understanding the cellular and molecular mechanisms underlying response to treatment in patients with inflammatory bowel disease (IBD) is crucial to guide drug development. We have used transcriptomic approaches to assess biomarkers of response to treatment among patients with active ulcerative colitis (UC) receiving the integrin inhibitor vedolizumab (VDZ) or systemic tumour necrosis factor (TNF) inhibitor therapy.

Aims & Methods

In VARSITY, 769 patients with moderately to severely active UC were randomized to receive VDZ (n = 383) or adalimumab (ADA; n = 386) for 52 weeks; superiority was demonstrated for VDZ relative to ADA for the primary endpoint of clinical remission at Week 52 (31.3% vs 22.5%; p = 0.006).¹ An unbiased cell and gene/pathway module analysis was conducted using baseline, Week 14, and Week 52 intestinal biopsy samples from treatment responders and non-responders, with response defined according to the primary endpoint. Longitudinal analysis was conducted focusing on tyrosine kinase 2 (TYK2)-dependent signalling pathways, including type I interferon (IFN) and interleukin (IL)-23. In addition, cell–cell signalling network analysis was conducted using single-cell RNA-seq data from patients with UC.²

Results

Using single-cell RNA-seq data,² distinct but overlapping signalling networks were observed for α4β7 integrin, IL-23 and TNF. Cell and gene/pathway module analysis of VARSITY transcriptomic data identified biomarkers at baseline, Week 14, and Week 52 associated with non-response to VDZ at Week 52, including CD4+, CD8+, B-cell and fibroblast cell subsets, and elevated transcription of gene modules associated with JAK/STAT and IFN signalling. Similar regulation differences were seen for response to ADA at Weeks 14 and 52, but no predictors of ADA response were identified at baseline. Targeted longitudinal analysis demonstrated enrichment of IL-23 and type I IFN pathways among VDZ and ADA non-responders throughout the study, and incomplete resolution of IL-23 and Type I IFN pathway modulation among treatment responders.

Conclusion

Elevations in gene modules associated with TYK2-dependent signalling were identified as biomarkers of non-response to both VDZ and ADA, suggestive of mechanistic complementarity between α4β7 integrin, TNF and TYK2 inhibition. Findings support further investigation of the role of TYK2 inhibitors in IBD, irrespective of prior response to VDZ or ADA.

References

1. Sands BE, et al. N Engl J Med 2019;381:1215–26.
2. Smillie CS, et al. Cell. 2019;178:714–30 e22.

Disclosure

MP, JT, BS, AS, VA, SF, RH, JA, CA, PP, NS and IP are employees and shareholders of Takeda. AND was an employee and shareholder of Takada at the time of the analysis. SS has received consultant or speaker fees from AbbVie, Allergosan, Amgen, Arena, BMS, Celltrion, Dr. Falk Pharma, Ferring, Galapagos, Genentech (Roche), Gilead, GSK, IMAB, Janssen, Lilly, MSD, Pfizer, Shire, Takeda, and Viatris.

Introduction

Microscopic colitis (MC) is an inflammatory bowel disease (IBD) that causes chronic non-bloody watery diarrhea and abdominal pain that significantly reduces the quality of life¹. MC can be subdivided into collagenous colitis (CC) and lymphocytic colitis (LC), which are clinically indistinguishable but differ histologically². The activation of immune responses and the alteration of the intestinal epithelial barrier via water channel aquaporin downregulation and crypt hyperproliferation are characteristic features of MC³, often shared by other gastrointestinal disorders, such as “classical” IBD (Crohn’s disease and ulcerative colitis, UC), or irritable bowel syndrome (IBS). Enteric glial cells (EGC) are key regulators of gut homeostasis and epithelial permeability, that respond to inflammation by entering a reactive state, potentially driving epithelial dysfunction. Whether EGC promote resolution or perpetuate inflammation remains unclear.⁴

Aims & Methods

MC is poorly understood and EGC role in MC is still unknown. Therefore, our aim is to investigate the involvement of EGC – due to their neuroimmune functions and involvement in intestinal dysmotility – in patients with MC. In addition, mast cells will be investigated since altered interactions with EGC have been demonstrated during IBS⁵.
Paraffin-embedded colonic biopsies from patients with active CC, LC, UC, IBS-diarrhea (D) and healthy controls (HC) were collected (n=13-26 per cohort). In addition, a subset of CC patients was also sampled during successful treatment with glucocorticosteroid 9mg budesonide (n=13). The amount of EGC was quantified using glial fibrillary acidic protein (GFAP) and S100 calcium-binding protein B (S100β). Mast cells were quantified using tryptase.

Results

Immunofluorescence analysis of EGC revealed an increased amount of EGC^GFAP+ in CC compared to HC (p<0.001). Of note, CC EGC^GFAP+ levels were comparable to those shown by biopsies from patients suffering from IBS-D and UC. Among the 13 CC patients with paired samples before and under budesonide treatment, we observed a trend toward reduced EGC^GFAP+ levels during treatment (p = 0.08). In contrast, LC patients displayed similar EGC^GFAP+ levels as to HC, but lower than those observed in biopsies from patients with CC, IBS-D, and UC (p<0.001). On the other hand, EGC^S100β+ were increased only in active CC (p<0.0001). Yet, UC patients displayed the highest level of EGC^S100β+ (p<0.0001). Mast cells analysis showed an increased counts for tryptase in CC before and under treatment compared to HC (p<0.01).

Conclusion

Our findings indicate that EGC and mast cells are involved in CC pathogenesis due to an increased reactivity of EGC and mast cells in the colonic mucosa. CC patients exhibit signs of EGC^GFAP+ levels comparable to those observed in UC and IBS-D. Oppositely, LC seems to be independent of neuroinflammation as EGC and mast cells presence was comparable to HC. These preliminary findings support the theory that CC and LC are distinct disease entities. Future work will include RNA sequencing of EGC-related genes, counts of eosinophils as well as in situ analysis of mast cells and eosinophils degranulation, and assessment of budesonide effects on EGC in vitro.

References

1. Nyhlin N, Wickbom A, Montgomery SM, et al. Long-term prognosis of clinical symptoms and health-related quality of life in microscopic colitis: a case-control study. Aliment Pharmacol Ther 2014; 39: 963-972. 20140311. DOI: 10.1111/apt.12685.
2. Carrasco A, Esteve M, Salas A, et al. Immunological Differences between Lymphocytic and Collagenous Colitis. J Crohns Colitis 2016; 10: 1055-1066. 20160229. DOI: 10.1093/ecco-jcc/jjw058.
3. Escudero-Hernandez C, Munch A, Ostvik AE, et al. The Water Channel Aquaporin 8 is a Critical Regulator of Intestinal Fluid Homeostasis in Collagenous Colitis. J Crohns Colitis 2020; 14: 962-973. DOI: 10.1093/ecco-jcc/jjaa020.
4. Gonzales J and Gulbransen BD. The Physiology of Enteric Glia. Annu Rev Physiol 2024 20241115. DOI: 10.1146/annurev-physiol-022724-105016.
5. Meira de-Faria F, Casado-Bedmar M, Marten Lindqvist C, et al. Altered interaction between enteric glial cells and mast cells in the colon of women with irritable bowel syndrome. Neurogastroenterol Motil 2021; 33: e14130. 20210402. DOI: 10.1111/nmo.14130.

Introduction

Dysregulated activation of myeloid cells, including monocytes and macrophages, contributes to intestinal inflammation in Inflammatory Bowel Disease (IBD) through excessive release of pro-inflammatory cytokines. Microbial-derived short-chain fatty acids (SCFAs) modulate the inflammatory and metabolic nature of these cells through activation of a wide-range of cellular signalling pathways. Several of these pathways—including targeting Free-fatty acid receptors (FFA1, FFA2, FFA3) and other metabolic pathways (Glycolysis and Oxidative phosphorylation) have exhibited potential by influencing the polarisation from inflammatory to regulatory macrophages. In this study, we aim to unravel these mechanisms and explore the interplay between myeloid cells and SCFAs in IBD.

Aims & Methods

We first investigated the immunomodulatory effects of SCFA on THP-1 and monocyte-derived macrophage (MDM) models of inflammation. These cells were polarised toward an M1-like phenotype (LPS+ IFN-γ) in the presence of SCFAs or pharmacological agonist of FFA3 (AR420626). We analysed how these treatments impacted cytokine secretion and markers of polarisation using ELISA and RT-qPCR. To gain insight with a disease context, CD14+ monocytes from the peripheral blood of IBD patients (n=6), were exposed to M1 polarising stimuli (LPS+IFN-γ), in the presence of SCFAs or AR420626. To explore potential disease-specific responses, monocytes from actively inflamed IBD patients (n=7) and healthy controls (HC) (n=3) were cultured in presence of butyrate (1mM) in the absence of inflammatory stimulus. In both instances, RT-qPCR and ELISA were conducted to assess secretion of pro-inflammatory cytokines and activation of metabolic pathways.

Results

In THP-1 and MDM models, treatment with butyrate (4 mM) markedly suppressed IL-6 secretion, an effect that was mimicked by the FFA3 agonist AR420626 (50 µM). These responses extended to IBD patient-derived monocytes, where strong induction of IL-6 and TNF-α were repressed by both butyrate and propionate (vs M1; p< 0.001). FFA3, which plays an important role in mediating responses to SCFAs, was also upregulated in M1 polarised patient monocytes (vs Unt; p< 0.05). However, this elevated expression was abrogated in monocytes that were exposed to butyrate (1mM), giving insight toward how butyrate can regulate SCFA sensing mechanisms under inflammatory conditions. Interestingly, this pro-inflammatory monocyte phenotype was accompanied by raised expression of the glycolysis markers HK2 and GLUT1 —which, again, were repressed by butyrate (vs Unt; p< 0.05). In untreated monocytes derived from IBD patients, we observed an elevated inflammatory state relative to HCs, suggesting prior disease-associated activation. While these cells displayed heightened baseline expression of HK2 than HC monocytes, this was attenuated by exposure to butyrate (vs Unt; p< 0.05), and cells were rescued from this hyper-inflammatory phenotype.

Conclusion

Taken together, these results highlight that butyrate helps to counteract inflammation in IBD monocytes —possibly stemming from an influence on glycolysis. This work demonstrates that monocyte metabolic activity and SCFA-sensing pathways are dysregulated under inflammatory conditions. These findings provide new insight into the altered immune-metabolic state of monocytes in IBD and highlight the important role of microbial-derived SCFAs to restoring immunometabolic balance.

Introduction

There is evidence linking inflammatory bowel disease (IBD) and cardiovascular diseases. However, the roles of histologic and clinical activity of IBD in the development of arrhythmias remain unknown.

Aims & Methods

In this nationwide cohort study in Sweden, we included IBD patients with histologic data (n = 61,383; diagnosed 1969–2017; classified as histologic inflammation vs. remission based on the Swedish version of the Systematized Nomenclature of Medicine codes) and clinical activity data (n = 96,154; diagnosed 1969–2020; classified as clinically active vs. quiescent IBD based on IBD-related hospitalization, surgery, and treatment escalation) and followed them through December 31, 2021. The primary outcome was incident arrhythmias (including atrial fibrillation/flutter, bradyarrhythmias, other supraventricular arrhythmias, and ventricular arrhythmias/cardiac arrest) within 2 years after documented histologic inflammation (vs. remission) or clinically active IBD (vs. quiescent IBD). Poisson regression models estimated incidence rates (IRs) of arrhythmias, while Cox proportional hazards models estimated adjusted hazard ratios (aHRs) and 95% confidence intervals (CIs). We adjusted for age at IBD diagnosis, sex, calendar year at index date, county of residence, educational attainment, country of birth, number of health care visits, and history of CVD-related comorbidities in the model.

Results

Histologic inflammation was associated with an increased risk of developing any arrhythmias compared to histologic remission (IR: 62.0 vs. 47.2 per 10,000 person-years; aHR = 1.35, 95%CI: 1.18–1.54), corresponding to one additional event of arrhythmia per 338 IBD patients over 2 years after histologic inflammation. We found an excess risk for atrial fibrillation/flutter (aHR = 1.38, 95%CI: 1.18–1.62) and ventricular arrhythmias/cardiac arrest (aHR = 1.73, 95%CI: 1.22–2.44), but not for bradyarrhythmias or other supraventricular arrhythmias. Compared to clinically quiescent IBD, active IBD was also related to a higher risk of arrhythmias (IR: 67.4 vs. 108.4; aHR = 1.77, 95%CI: 1.66–1.88), affecting all subtypes except bradyarrhythmias. In patients with clinically quiescent IBD, histologic inflammation was also associated with an increased risk of arrhythmias (aHR = 1.16, 95%CI: 1.00–1.36, P=0.054).

Conclusion

Histologic and clinical activity were associated with a slightly increased risk of arrhythmias in patients with IBD.

Disclosure

All authors have completed the ICMJE uniform disclosure form and declare:
OO has been PI on projects at Karolinska Institutet financed by grants from Janssen, Pfizer, AbbVie, Takeda, Galapagos/Alfasigma, Ferring, and Bristol Myer Squibb. JH served as speaker or advisory board member for AbbVie, BMS, Celltrion, Eli Lilly, Ferring, Galapagos, Gilead, Index Pharma, Janssen, Medtronic, Merck, MSD, Novartis, Pfizer, Sandoz, Shire, Takeda, Thermo Fisher Scientific, Tillotts Pharma, and Vifor Pharma and received grant support from Takeda, Janssen, and MSD. FE has served as an advisory board member for Boehringer Ingelheim. JFL has coordinated a study for the Swedish IBD Quality Register (SWIBREG), which received funding from Janssen Corporation. JFL has also received financial support from MSD to develop a paper reviewing national healthcare registers in China. JFL has a research collaboration on celiac disease with Takeda and has a research collaboration on fibrosis in IBD with MSD. The other authors report no disclosures relevant to the manuscript.

Introduction

Faecal microbiota transplantation (FMT) is an emerging therapy in inflammatory bowel disease (IBD), supported by evidence for improved clinical and endoscopic outcomes. Decontamination of the recipient gastrointestinal tract using antibiotics or colonic lavage has been proposed to improve FMT efficacy by reducing microbial load and creating a niche for donor organism colonisation. The aim of this study was to assess the effects of antibiotic pre-treatment and colonic lavage on clinical and endoscopic outcomes of FMT in IBD.

Aims & Methods

A systematic review of the literature was conducted by searching PubMed, MEDLINE and Scopus from inception to July 2024. Prospective studies examining FMT in IBD with or without antibiotic pre-treatment and/or colonic lavage were included. Outcomes included clinical response, clinical remission, endoscopic response and endoscopic remission. A random-effects model was used to calculate pooled proportions with 95% confidence intervals (CI).

Results

Forty-one studies (n=1185) were included, of which 7 (17.1%) incorporated antibiotic pre-treatment and 29 (70.7%) incorporated colonic lavage prior to FMT therapy. Eight studies (19.5%) were in Crohn’s disease, 32 (78%) were in ulceratice colitis and one study incorporated any IBD. Antibiotic pre-conditioning courses varied in terms of antibiotics used, treatment duration and washout period prior to FMT. Polyethylene glycol was the most used lavage preparation (23 studies, 79.3%). Antibiotic pre-treatment was associated with numerically higher rates of clinical response (64% vs. 56%), clinical remission (47% vs. 39%), endoscopic response (59% vs. 45%) and endoscopic remission (47% vs. 21%) than no antibiotics, but these differences did not reach statistical significance. Colonic lavage was associated with significantly lower clinical response (52% vs. 71%) compared to no lavage. Clinical remission rates were numerically lower in the lavage group (37% vs. 46%), but again, not statistically significant. Endoscopic response and remission rates were similar. Risk of bias was high or moderate in most studies.

Conclusion

The results of this review suggest while antibiotics may improve FMT outcomes in IBD, the evidence remains inconclusive, and further adequately powered studies are needed before changing clinical practice. Colonic lavage appears to reduce clinical response to FMT and should be avoided where feasible. FMT delivery methods that avoid the need for bowel preparation, such as capsules, enemas or upper gastrointestinal administration should be considered preferable.

Disclosure

RV Bryant, SP Costello and E Tucker are shareholders in BiomeBank.

Introduction

IIn Crohn’s disease, chronic inflammation drives fibroblast activation, excessive collagen deposition, and creeping fat expansion, all contributing to fibrostenotic strictures. We examined serum markers related to collagen formation, macrophage and neutrophil activation, and NETosis, exploring their associations with stricturing Crohn’s disease in a treatment-naïve IBD inception cohort.

Aims & Methods

142 Serum samples from CD patients were analysed (IBSEN III population matched baseline and 1-year follow-up). The Montreal classification (luminal, fibrostenotic strictures, fistula) was applied to assess disease complications at the time of serum sampling, e.g. baseline biomarkers measurements were paired with baseline Montreal B assessment, and one-year followup biomarkers measurements were paired with one-year followup Montreal B assessment. The biomarkers C4M, C6M, C7M, PRO-C3, PRO-C4, PRO-C5, PRO-C6, PRO-C11, PRO-C16, PRO-C22, VICM, CTX-III, CPa9-HNE were measured in the serum samples at baseline (total n=142; luminal n=114; fibrostenotic n=23, fistula n=5) and 1-year follow-up (total n=142; luminal n=111; fibrostenotic n=24, fistula n=7). One-way-ANOVA with LSD correction was applied for statistical analysis.

Results

The PRO-C6 (p<0.001) and CPa9-HNE (p<0.05) biomarkers were significantly elevated in CD patients with fibrostenotic strictures compared to luminal CD patients at baseline, and PRO-C6 was continually elevated in serum after 1 year of follow-up in patients with fibrostenotic strictures (p<0.01). At baseline, C7M was demonstrated to be elevated in fistulizing CD patients compared to luminal and fibrostenotic CD patients (P<0.01). PRO-C5 (p<0.001) and PRO-C16 (p<0.01) were significantly elevated at 1-year follow-up in the CD patients with fibrostenotic strictures compared with luminal CD patients. None of the remaining biomarkers were associated with stenotic or fistulizing CD.

Conclusion

These encouraging biomarker data from the IBSEN III, especially PRO-C6, seem to be a promising biomarker for intestinal fibrosis, as PRO-C6 was demonstrated to be elevated at baseline and after 1 year follow-up. CPa9-HNE levels were highest in the fibrostenotic group, suggesting neutrophil activity and NETosis involvement in intestinal fibrogenesis. Elevated levels of C7M were associated with fistula in CD patients, suggesting that C7M is reflective of transmural inflammation. The biomarkers PRO-C5 and PRO-C16 are also relevant biomarkers of established intestinal fibrosis, as these were elevated after 1 year of follow-up. Collectively, these biomarker data highlight the importance of quantifying collagen protein fragments in serum as tools for assessing intestinal fibrosis in Crohn’s disease.

Disclosure

Full time employee at Nordic Bioscience A/S

Introduction

Our group previously demonstrated activation of mitochondrial unfolded protein response (UPR^mt) in colonic mucosa from Inflammatory Bowel Disease (IBD) patients and linked this activation to pro-inflammatory signaling¹.

Aims & Methods

In the present study, we further investigate the role of Growth Differentiation Factor 15 (GDF15), a downstream mediator of the UPR^mt.
Circulating levels of GDF15 and Lipopolysaccharide Binding Protein (LBP) were assessed in plasma from IBD patients and healthy controls (HC) using ELISA. Myocardial infarction (MI) patients were used as positive control for elevated GDF15. Biochemical parameters from blood test were determined. Colonic organoids were used to deepen into the effects of GDF15: a permeability assay using FITC-Dextran 4K was performed, and levels of Zonula Occluddens 1 (ZO-1) were assessed using immunofluorescence.

Results

A total of 23 HC, 39 IBD and 19 MI patients were recruited (table 1). We found no statistical differences in sex ratio, age or body mass index between HC and IBD patients. Levels of GDF15 in plasma from IBD patients were significantly increased compared to HC (p=0.0401). When stratified, only UC patients maintained significant higher levels (p=0.0359), while CD patients showed higher levels, but not statistically significant. We observed a correlation between circulating GDF15 levels and markers of inflammation (C-reactive protein, Spearman r=0.3290, p=0.0066) and intestinal permeability (haptoglobin, Pearson r=0.4066, p=0.0017; and LBP, Spearman r=0.2032, p=0.0423). In vitro treatment of colonic organoids with 100 and 400 ng/mL of GDF15 showed higher entrance of FITC-Dextran 4K to organoids lumen compared to untreated organoids (p=0.0106 and p=0.0195, respectively), suggesting a GDF15-induced barrier alteration. We finally observed that 100 ng/mL of GDF15 induce a significant down-regulation of ZO-1 (p=0.0129), a critical component of the tight junctions.

Conclusion

Our findings confirm previous reports of increased circulating GDF15 levels in IBD patients and the activation of UPR^mt. Moreover, we describe a novel mechanism in IBD pathophysiology, linking mitochondrial stress to the disruption of the intestinal barrier and increased intestinal permeability.

References

1. Martín-Reyes F, Bernal M, Rodríguez-Díaz C, et al. Mitochondrial Stress Links Environmental Triggers with Pro-Inflammatory Signaling in Crohn’s Disease. Antioxidants. 2023;12(12). doi:10.3390/antiox12122105

Disclosure

A.F.C. has received honoraria from AbbVie, Pfizer, Galapagos, and Janssen. F.G.M. has received honoraria from AbbVie, Pfizer, Galapagos, and Janssen. F.J.R.G. has received honoraria from AbbVie, Pfizer, Galapagos, and Janssen; G.A.M. has received honoraria from AbbVie, Pfizer, Kern Pharma, Takeda, Galapagos, Amgen, MSD, and Janssen. The remaining authors have nothing to declare.