Pathogenesis Of Crohn Research Articles

A New Probiotic Formulation Promotes Resolution of Inflammation in a Crohn's Disease Mouse Model by Inducing Apoptosis in Mucosal Innate Immune Cells.

The interaction between gut-residing microorganisms plays a critical role in the pathogenesis of Crohn's disease (CD), where microbiome dysregulation can alter immune responses, leading to unresolved local inflammation. The aim of this study is to analyze the immunomodulatory properties of a recently developed probiotic + amylase blend in the SAMP1/YitFc (SAMP) mouse model of CD-like ileitis. Four groups of SAMP mice were gavaged for 56 days with the following treatments: 1) probiotic strains + amylase (0.25 mg/100 µL PBS); 2) only probiotics; 3) only amylase; PBS-treated controls. Ilea were collected for GeoMx Digital Spatial Profiler (DSP) analysis and histological evaluation. Histology assessment for inflammation indicated a significantly reduced level of ileitis in mice administered the probiotics + amylase blend. DSP analysis showed decreased abundance of neutrophils and increased abundance of dendritic cells, regulatory T cells, and macrophages, with a significant enrichment of five intracellular pathways related to apoptosis, in probiotics + amylase-treated mice. Increased apoptosis occurrence was confirmed by (TdT)- deoxyuridine triphosphate (dUTP)-biotin nick end labeling assay. Our data demonstrate a beneficial role of the probiotic and amylase blend, highlighting an increased apoptosis of innate immunity-associated cell subsets, thus promoting the resolution of inflammation. Hence, we suggest that the developed probiotic enzyme blend may be a therapeutic tool to manage CD and therefore is a candidate formulation to be tested in clinical trials.

Advances in the research of intestinal fungi in Crohn's disease

This article reviews of the original research published by Wu et al in the World Journal of Gastroenterology , delving into the pivotal role of the gut microbiota in the pathogenesis of Crohn's disease (CD). Insights were gained from fecal microbiota transplantation (FMT) in mouse models, revealing the intricate interplay between the gut microbiota, mesenteric adipose tissue (MAT), and creeping fat. The study uncovered the characteristics of inflammation and fibrosis in the MAT and intestinal tissues of patients with CD; moreover, through the FMT mouse model, it observed the impact of samples from healthy patients and those with CD on symptoms. The pathogenesis of CD is complex, and its etiology remains unclear; however, it is widely believed that gut microbiota dysbiosis plays a significant role. Recently, with the development and application of next-generation sequencing technology, research on the role of fungi in the pathogenesis and chronicity of CD has deepened. This editorial serves as a supplement to the research by Wu et al who discussed advances related to the study of fungi in CD.

Abnormal Promoter Methylation of Nucleotide-Binding Oligomerization Domain Containing 2 (NOD2) Gene in the Pathogenesis of Crohn’s Disease

Background: Changes in the expression of nucleotide-binding oligomerization domain containing 2 (NOD2) play an important role in the pathogenesis of a variety of autoimmune diseases including inflammatory bowel diseases (IBDs). Epigenetic modifications, including DNA methylation, are considered an important mechanism in the suppression of gene activity. In this study, we investigated the relationship between DNA methylation patterns of the promoter region of the NOD2 gene and the pathogenesis of Crohn’s disease (CD). Methods: Colonic mucosa samples were obtained from 15 Iranian patients with IBD and 15 age- and sexmatched healthy controls with no history of autoimmune disease. After the bisulfite conversion of genomic DNA, the DNA methylation status of three CpG sites in the promoter region of the NOD2 gene was determined by the real-time quantitative multiplex methylation-specific PCR (QM-MSP) assay. Results: Using this approach, we identified that IBD patients showed a decreased level of methylation of the NOD2 promoter in the colonic mucosa than did the healthy controls. (Unmethylated DNA in Crohn's disease vs. healthy controls; 0.128±0.093 vs. 0.025±0.016, P<0.000). Conclusion: According to our findings, promoter hypomethylation of the NOD2 gene in the colonic mucosa might contribute to the development and severity of CD. Furthermore, aberrant DNA methylation levels are expected to serve as a clinically useful risk marker.

Interferon γ Expressing Mucosal Cells in Pediatric Chronic Inflammatory Bowel Disease.

The pathogenesis of Crohn's disease (CD) and ulcerative colitis (UC) is multifactorial and includes aberrations in the composition of gastrointestinal mucosal inflammatory cells. Accurate identification of CD and UC is important as treatment and prognosis differs; however, CD and UC may be difficult to differentiate. Interferon γ (IFNγ) expression appears to be increased in ileal mucosa from CD patients, implying that IFNγ could be a diagnostically useful marker to differentiate CD from UC. This study uses automated assessment of IFNγ immunohistochemical expression in archival GI mucosal biopsies from stomach, duodenum, terminal ileum, and colon in a pediatric population to address this possibility. IFNγ positive mucosal cells are increased in the colon in both CD and UC compared to normal colon and in the ileum of CD compared to normal and UC. The abundance of IFNγ positive cells is not correlated with the presence of active inflammation, indicating that active inflammation is not responsible for the variance in abundance of IFNγ positive cells between cohorts and sites. Overlap between CD, UC, and normal suggests that IFNγ immunohistochemistry may only be clinically useful in select situations such as undetermined inflammatory bowel disease and additional study in these areas is warranted.

The effect of dietary emulsifiers and thickeners on intestinal barrier function and its response to acute stress in healthy adult humans: A randomised controlled feeding study.

Although dietary emulsifiers are implicated in the pathogenesis of Crohn's disease, their effect has not been studied in humans. To determine the effects of high- and low-emulsifier diets (HED, LED) on intestinal barrier function in healthy subjects in unstressed and acutely stressed states. We conducted a single-blinded, cross-over, controlled feeding trial in 22 healthy adults. After recording 7 days of their habitual diet, we randomised participants to HED or LED with ≥3-week washout between diets. On dietary completion, acute stress was induced via intravenous corticotrophin-releasing hormone. We assessed dietary adherence, effects on 2-h urinary lactulose: rhamnose ratio (LRR), serum concentrations of lipopolysaccharide-binding protein, soluble-CD14 and markers of epithelial injury and inflammation. Dietary adherence was excellent. In an unstressed state, median (interquartile range) LRR during HED was 0.030 (0.018-0.042); on LED, this was 0.042 (0.029-0.078; p = 0.04). LPB concentrations were lower on HED than LED (p = 0.026), but no differences were observed for epithelial injury or inflammation. Under acute stress, LRR increased by 89% (-1% to 486%) on HED (p = 0.004), differing (p = 0.001) from 39% (1%-90%) decrease on LED (p = 0.009). Soluble-CD14 also increased (p < 0.001). The LED had a prolonged carry-over effect on suppressing HED-induced changes during stress. Similar changes in LRR and soluble-CD14 were observed when HED was used as the first diet (both p < 0.01). High intake of emulsifiers improved barrier function in the unstressed state, but increased intestinal permeability to stress, without evidence of inflammation. A LED was protective of the stress effect.

Integrated analyses reveal the diagnostic and predictive values of COL5A2 and association with immune environment in Crohn’s disease

The pathogenesis of Crohn’s disease (CD) involves abnormal immune cell infiltration and dysregulated immune response. Therefore, thorough research on immune cell abnormalities in CD is crucial for improved treatment of this disease. Single-cell RNA sequencing (scRNA-seq) and bulk RNA-seq data of CD were obtained from the Gene Expression Omnibus (GEO) database. Cell-type identification by estimating relative subsets of RNA transcripts (CIBERSORT), weighted gene co-expression network analysis (WGCNA), protein–protein interaction (PPI) networks evaluated the proportion of immune infiltrating cells, constructed co-expression network and identified key genes, respectively. Based on the dataset (GSE134809), 15 cell clusters were defined and labeled as different cell types. Among the 11 modules, the yellow module had the closest relationship with plasma cells (cluster 5). Confirmed using RNA sequencing and IHC assay, the expression of COL5A2 in CD samples was higher than that in control samples. Furthermore, the COL5A2 protein expression remarkably decreased in the group of patients who responded to anti–tumor necrosis factor (TNF) treatments, compared to the non-response group. The comprehensive analyses described here provided novel insight into the landscape of CD-associated immune environment. In addition, COL5A2 were identified as potential diagnostic indicators for CD, as well as promising predictive markers for CD patients.

Targeting Inflammation and Oxidative Stress to Improve Outcomes in a TNBS Murine Crohn's Colitis Model.

Inflammation and oxidative stress are implicated in the pathogenesis of Crohn's disease. Cerium oxide nanoparticle (CNP) conjugated to microRNA 146a (miR146a) (CNP-miR146a) is a novel compound with anti-inflammatory and antioxidative properties. We hypothesized that local administration of CNP-miR146a would improve colitis in a 2,4,6-Trinitrobenzenesulfonic acid (TNBS) mouse model for Crohn's disease by decreasing colonic inflammation. Balb/c mice were instilled with TNBS enemas to induce colitis. Two days later, the mice received cellulose gel enema, cellulose gel with CNP-miR146a enema, or no treatment. Control mice received initial enemas of 50% ethanol and PBS enemas on day two. The mice were monitored daily for weight loss and clinical disease activity. The mice were euthanized on days two or five to evaluate their miR146a expression, inflammation on histology, and colonic IL-6 and TNF gene expressions and protein concentrations. CNP-miR146a enema successfully increased colonic miR146a expression at 12 h following delivery. At the end of five days from TNBS instillation, the mice treated with CNP-miR146a demonstrated reduced weight loss, improved inflammation scores on histology, and reduced gene expressions and protein concentrations of IL-6 and TNF. The local delivery of CNP-miR146a in a TNBS mouse model of acute Crohn's colitis dramatically decreased inflammatory signaling, resulting in improved clinical disease.

Complanatuside A ameliorates 2,4,6-trinitrobenzene sulfonic acid-induced colitis in mice by regulating the Th17/Treg balance via the JAK2/STAT3 signaling pathway.

Immunity imbalance of T helper 17 (Th17)/regulatory T (Treg) cells is involved in the pathogenesis of Crohn's disease (CD). Complanatuside A (CA), a flavonol glycoside, exerts anti-inflammatory activities and our study aimed to identify its effect on TNBS-induced colitis and the possible mechanisms. We found that CA alleviated the symptoms of colitis in TNBS mice, as demonstrated by prevented weight loss and colon length shortening, as well as decreased disease activity index scores, inflammatory scores, and levels of proinflammatory factors. Flow cytometry analysis showed that CA markedly reduced the percentage of Th17 cells while increasing the percentage of Treg cells in TNBS mice. Under Th17 cell polarizing conditions, CA inhibited the differentiation of Th17 cells while the Treg cell differentiation was elevated under Treg cell polarizing conditions. Furthermore, it was observed that JAK2 interacted with CA through six hydrogen bonds via molecular docking. The phosphorylation of JAK2/STAT3 was reduced by CA, which might be correlated with the protective effect of CA on colitis. In conclusion, CA reduced the imbalance of Th17/Treg cells by inhibiting the JAK2/STAT3 signaling pathway in TNBS-induced colitis, which may provide novel strategies for CD treatment.

Monoclonal Antibodies Against Mature Interleukin-18 Ameliorate Colitis and Repair Goblet Cell Function

BackgroundNumerous biological interventions and small molecules are used to treat Crohn’s disease; however, the effectiveness of these treatments varies largely. Non-responsiveness to biological therapies is associated with interleukin (IL)-18 gene polymorphisms and high IL-18 expression has been implicated in the pathogenesis of Crohn’s disease.AimsThe aim of this study was to elucidate the expression of precursor and mature IL-18 in patients with Crohn’s disease who exhibited varied responses to cytokine-targeted treatments and determine whether selective inhibition of mature IL-18 offers a novel therapeutic avenue.MethodsWe generated a monoclonal antibody that specifically recognizes the neoepitope of caspase-cleaved mature IL-18. Expression of precursor and mature IL-18 was analyzed in patients with Crohn’s disease. Anti-mature IL-18 monoclonal antibodies were intraperitoneally administered in an acute colitis mouse model, and the disease activity index, body weight loss, tissue pathology, proinflammatory cytokine expression, goblet cell function, and microbiota composition were assessed.ResultsPrecursor and mature IL-18 expression was upregulated and goblet cell function was impaired in patients with Crohn’s disease who were unresponsive to biological therapies. Administration of anti-mature IL-18 antibodies ameliorated induced colitis by repairing goblet cell function and restoring the mucus layer.ConclusionsThe newly developed monoclonal antibody holds promise as a therapeutic alternative for Crohn’s disease.

TolDC Restores the Balance of Th17/Treg via Aryl Hydrocarbon Receptor to Attenuate Colitis.

Tolerogenic dendritic cells (TolDCs) have been evidenced to trigger regulatory T cell's (Treg's) differentiation and be involved in the pathogenesis of Crohn's disease (CD). Aryl hydrocarbon receptor (AhR) plays a crucial role in the differentiation of TolDCs, although the mechanism remains vague. This study aimed to evaluate the role of AhR in TolDCs formation, which may affect Th17/Treg balance in CD. Colon biopsy specimens were obtained from healthy controls and patients with CD. Wild type (WT) and AhR-/- mice were induced colitis by drinking dextran sulphate sodium (DSS) with or without 6-formylindolo 3,2-b carbazole (FICZ) treatment. Wild type and AhR-/- bone marrow-derived cells (BMDCs) were cultured under TolDCs polarization condition. Ratios of DCs surface markers were determined by flow cytometry. Enzyme-linked immunosorbent assay (ELISA) was performed to quantify the levels of interleukin (IL)-1β, transforming growth factor (TGF)-β and IL-10. Tolerogenic dendritic cells differentiated from BMDCs of WT or AhR-/- mice were adoptively transferred to DSS-induced WT colitis mice. Patients with CD showed less AhR expression and activation in their inflamed colon regions. Compared with WT mice, AhR-/- mice experienced more severe colitis. Tolerogenic dendritic cells and Tregs were both decreased in the colon of AhR-/- colitis mice, while Th17 cells were upregulated. In vitro, compared with WT DCs, AhR-deficient DCs led to less TolDC formation. Furthermore, intestinal inflammation in WT colitis mice, which transferred with AhR-/- TolDCs, showed no obvious improvement compared with those transferred with WT TolDCs, as evidenced by no rescues of Th17/Treg balance. Activation of AhR attenuates experimental colitis by modulating the balance of TolDCs and Th17/Treg. The AhR modulation of TolDCs may be a viable therapeutic approach for CD.

Identification of differentially expressed genes associated with ferroptosis in Crohn's disease.

Ferroptosis-related genes may play a critical regulatory role in the pathogenesis of Crohn's disease (CD). The purpose of the present study was to identify genes expressed in CD that are associated with ferroptosis, and to provide guidance in the diagnosis and therapy of CD. CD mRNA expression data were initially gathered from the Gene Expression Omnibus (GEO) database. GSE75214 and GSE102133 datasets were selected as the major targets and were analyzed for differentially expressed genes (DEGs). Subsequently, R software was used to analyze the common genes among the DEGs between CD and ferroptosis-related genes. Gene Ontology enrichment analysis and Kyoto Encyclopedia of Genes and Genome pathway analysis were conducted to identify related pathways and functions. Protein-protein interaction (PPI) analysis was performed to identify target genes. The DSigDB website was used to predict potential target drugs for hub genes. Reverse transcription-quantitative (RT-q) PCR was employed to detect the expression of these ferroptosis-related genes in clinical samples obtained from healthy controls and patients with CD. According to the two GEO datasets, 13 ferroptosis DEGs (11 upregulated genes and two downregulated genes) were identified in CD with thresholds of P<0.05 and |log2 fold change|>1, and were selected for further analysis. PPI analysis indicated the mutual effects among these genes and filtered out five hub genes. The top 10 potential targeted drugs were selected. The qPCR results showed that the expression levels of three genes, namely, IL-6, prostaglandin-endoperoxide synthase 2 (PTGS2) and dual oxidase 2 (DUOX2), were different between CD samples and healthy samples. This result was consistent with the results obtained from the bioinformatics analysis. In conclusion, bioinformatics analysis identified a total of 13 ferroptosis-associated genes in CD. Further verification by qPCR showed that IL-6, PTGS2 and DUOX2 may affect the process of CD by regulating ferroptosis. These findings might provide new biomarkers, diagnostic and therapeutic markers for CD.

Antibiotics for inflammatory bowel disease: Current status.

There is abundant literature reporting about the use of antibiotics in inflammatory bowel disease (IBD), but their role in the management of IBD is not entirely clear. Diverse infectious organisms have been implicated in the pathogenesis of Crohn's disease. Also, infections are believed to be a trigger for flares of ulcerative colitis. The benefit of the routine use of antibiotics in IBD is equivocal. However, there are certain situations, where antibiotics have a clear role and evidence of benefit: perianal fistula, intra-abdominalabscesses in Crohn's disease, acute pouchitis and infection-related flares. However, there is a lack of supportive evidence for the routine use of antibiotics in all disease-related flares. Evidence indicates a lack of benefit of intravenous antibiotics in acute severe ulcerative colitis and only limited benefit in active ulcerative colitis. Limited evidence suggests the role of a combination of oral antibiotics in pediatric ulcerative colitis. Certain targeted antibiotic regimens have been used in IBD. In ulcerative colitis, limited evidence suggests the benefit of the use of an antibiotic cocktail directed against Fusobacterium varium. Therapy directed against Escherichia coli does not seem to have a benefit in inflammatory Crohn's disease. In Crohn's disease, antimycobacterial therapy may result in symptomatic improvement but no durable benefit. Antitubercular therapy (ATT), on the contrary, may result in fibrotic transformation, suggesting a need to avoid misdiagnosis and limit the duration of ATT in Crohn's disease. This review assesses the published literature with respect to antibiotic use and provides guidance to clinicians in appropriate antibiotic use in various situations in the setting of IBD.

P452 Serum Amyloid A for Predicting Surgery and Disease Exacerbation in Patients with Newly Diagnosed Crohn’s Disease

Abstract Background Crohn's disease (CD) is characterized by relapsing-remitting course with highly heterogenous prognosis, ranging from quiescent disease to complicated course with stricturing, penetrating or need of abdominal surgery. Identifying biomarkers that determine their disease course can enable implementing more aggressive preventive strategies. Serum amyloid A (SAA) has been demonstrated to involve in the pathogenesis of Crohn’s disease (CD); however, its prognostic value is still uncertain. This study was determined to explore the prognostic value of SAA for CD-related surgery and disease exacerbation in newly diagnosed CD. Methods We conducted a retrospective cohort study and recruited newly-diagnosed CD patients at the First Affiliated Hospital of Sun Yat-sen University. Level of SAA was detected at diagnosis. The primary outcome was CD-related surgery. The secondary outcome was disease exacerbation, defining as newly developed structuring or penetrating lesions. Association between variables and outcome was evaluated through Kaplan–Meier method and Cox proportional hazard regression analysis. For continuous variables, the optimal cutoff values were determined by using the survminer R package in the analysis of primary outcome. The prognostic model was developed by cox proportional-hazards model. Concordance-index (c-index) was used to assess the predictive ability of the prognostic model. Results During 2187.6 person-years (median age, 28 years, 72.4% male), 87 surgery and 153 disease exacerbation events were documented. After adjustment, a 14% higher risk for surgery (adjusted hazard ratio, aHR [95% confidence interval]: 1.14 [1.05–1.23]; p=0.001) and a 12% higher risk for disease exacerbation (1.12 [1.05–1.19]; p&amp;lt;0.001) were observed with a 100-unit increase in SAA level. Patients with SAA levels ≥89.2 mg/L had aHRs of 2.08 [1.31–3.28] and 1.72 [1.22–2.41] for CD-related surgery and disease exacerbation, respectively. The association between SAA and outcomes was assessed as linear by the restricted cubic spline. Adding SAA into the model including known risk factors, including age at diagnosis ≥40 years, disease behavior, disease location, and initial biological treatment, led to significant improvement in its predictive ability for surgery (net reclassification index and integrated discrimination index, p&amp;lt;0.001) and disease exacerbation (net reclassification index and integrated discrimination index, p&amp;lt;0.001). Sensitivity analyses showed robust results. Conclusion Our study shows that higher serum SAA level at diagnosis was associated with poorer prognosis, including risk of surgery and disease exacerbation. And prognostic models based on SAA and known risk factors could better predict prognosis.

P583 Infliximab Monitoring in Crohn's Disease and Development of Neural Networks Using Clinical Variables to Predict Disease Activity and Immunogenicity

Abstract Background Infliximab (IFX) is a biological drug that inhibits the action of tumor necrosis factor-alpha (TNFα), a pro-inflammatory cytokine enrolled in the pathogenesis of Crohn's disease (CD). Trough-level drug monitoring (TDM) has been increasingly used in clinical practice to optimize biological therapy and improve precision medicine. Therefore, this study aimed to analyze the serum level of IFX and antidrug antibodies (ADA) in active and in remission CD patients and develop prediction models via neural networks using a combination of clinical variables. Methods Seventy-five CD patients who underwent IFX therapy in the maintenance phase were included. Disease activity was defined by endoscopic and/or radiological criteria, comprising remission (CDR) and active (CDA) groups. Serum IFX levels were measured by ELISA (Promonitor ®) and rapid lateral flow test (Quantum-Blue®). ADA was measured using ELISA (Promonitor ®). For statistical analysis, a non-parametric test was used, with adopted p≤0.05. To create a prediction model, four sets of neural networks were constructed for the following output variables: remission/activity, presence or not of ADA, therapeutic level according to ELISA, and therapeutic level according to the lateral flow test. The neural networks and ROC curves were designed using the Keras package in Python software. The study was approved by the Research Ethics Committee. Results No differences were observed in the IFX level when comparing the CDA and CDR groups in both tests (p&amp;gt; 0.05). There was no statistical difference in IFX levels according to the use of immunosuppressants (IMS) (p&amp;gt;0.05). ADA levels &amp;gt; 5AU/mL were detected in only 11 (14.6%) patients. All patients with positive ADA had infratherapeutic IFX levels in both tests, and 72.7% were on combination therapy with IMS. Of the 4 neural networks developed, two, whose output variables were “remission/activity” and “presence or not of ADA,” showed excellent performance, with AUC ranging from 82% to 92% and 100%, respectively. Conclusion The introduction of IFX monitoring may allow more individualized and precise therapeutic management. Regarding the performance characteristics of the neural networks, an AUC &amp;gt;80% was evident in two of them. Through clinical and non-invasive laboratory variables, disease activity was determined. Furthermore, the finding of combinations of variables that determine the ADA level, and at the same time not having demonstrated good models for IFX serum level measurement, means that the latter continues to be necessary for clinical practice. Otherwise, ADA levels, which indicate immunogenicity, can be predicted by non-invasive clinical variables.

DOP19 High dimensional profiling of IL-23-responsive cells revealed molecular signatures that predict response to anti-IL-23 therapy in patients with Crohn’s Disease

Abstract Background The interleukin (IL)-23 signaling plays a crucial role in the pathogenesis of Crohn’s disease (CD). IL-23 activates T-helper (Th) 17 cells, IL-17-secreting CD8 T (Tc)17 cells, gd T cells, nature killer (NK) T cells, and group 3 innate lymphoid cells (ILC3) to secrete proinflammatory cytokines including IL-17, IFN-g, and TNF-a. Risankizumab is the first selective IL-23 antagonist approved for moderate-severe CD. However, there is currently no biomarker to predict response to anti-IL-23 antibodies in IBD. Here, we characterized the cell type- and cytokine-specific pathways in IL-23-responsive cells that predict response to risankizumab in CD patients. Methods Adult patients with a history of ileal, colonic, or ileocolonic CD were included. Active CD in terminal ileum and/or colon were confirmed by colonoscopy and/or MRI. Blood samples were collected within 3 months before and 4, 8, 12, and 52 weeks after the initiation of risankizumab. Peripheral blood mononuclear cells (PBMC) were isolated and cryopreserved for batch analysis using mass cytometry (CyTOF). Pre-treatment PBMCs were stimulated ex vivo with IL-23, and stained for lineage markers to identify Th17, Tc17, gd T, NKT cells, and ILC3, as well as intracellular cytokines including IL-17, IFN-g, TNF-a, IL-22, IL-6, and GM-CSF, using a pre-optimized and validated antibody panel. The CyTOF data were analyzed using the viSNE tool in Cytobank and FlowJo. Patients’ responses were evaluated 12-40 weeks after the initiation of risankizumab based on clinical symptoms and endoscopic/radiographic assessment. Results Among IL-23-responsive cells, increased frequency of NKT cells was identified in the peripheral blood of non-responders before initiation of risankizumab (Figure 1A). In non-responders, Tc17 cells, gd T cells, and ILC3 demonstrated significant elevations in TNF-a, a key pathogenic mediator in CD. gd T cells from non-responders also showed increased IL-17F before treatment. In contrast, NKT cells from non-responders expressed significantly lower GM-CSF, which was previously shown to alleviate intestinal inflammation in patients and mouse models (Figure 1B). Ongoing study is investigating the cytokine production in IL-23-responsive cells after the initiation of risankizumab, as well as before and after starting ustekinumab, an anti-IL-12/IL-23 antibody. Conclusion High dimensional profiling of pre-treatment PBMCs revealed molecular signatures, e.g., elevated TNF-a in multiple IL-23-responsive cells, which are associated with resistance to risankizumab in CD patients. Those findings may help identify response biomarkers for IL-23 antagonists and provide a rationale for using anti-IL-23/anti-TNF combination therapy in some patients with refractory CD.

The role of serum bilirubin in the pathogenesis of Crohn’s disease in children and assessment of its activity

Introduction. The article is devoted to the identification of a marker of imbalance of oxidative stress and the antioxidant system in children with Crohn’s disease. The putative level of total bilirubin as an integrative method for assessing Crohn’s disease activity is highly correlated with indicators of oxidative stress and the antioxidant system (ischemia, modifying albumin, malondialdehyde, catalase, sulfhydryl groups and glutathione). Purpose: to explain the relationship between the level of total bilirubin and the activity of the inflammatory process in Crohn’s disease, which allows the introduction of the studied integrative indicator in clinical practice. Materials and methods. Based on the results of scientific studies of biomarkers of oxidative stress (albumin-modifying ischemia - plasma IMA, malondialdehyde - plasma MDA) and the antioxidant system (plasma SH-groups, catalase and reduced erythrocyte glutathione), as well as a simultaneous study of the level of total bilirubin in children diagnosed with Crohn’s disease. To search for a differential method for assessing the level of Crohn’s disease activity by the degree of bilirubin, use binary logistic regression. Results and survey. The results of the study showed a clear relationship between the increase in bilirubin levels and the severity of oxidative stress. It has been established that with a decrease in the level of total bilirubin, there is a predominance of oxidative stress over the antioxidant system, which leads to an increase in inflammation activity. It has been proven that the level of total bilirubin can be used in the complex activity of diseases as a minimally invasive, as well as an objective and acute marker. The sensitivity and specific sensitivity of this protocol is 90.9% and 93.3% respectively, which allows the use of total bilirubin as a marker of inflammation in Crohn’s disease.

Longitudinal dynamics of gut bacteriome and mycobiome interactions pre- and post-visceral surgery in Crohn's disease.

Alterations of the gut microbiome are involved in the pathogenesis of Crohn's disease (CD). The role of fungi in this context is unclear. This study aimed to determine postoperative changes in the bacterial and fungal gut communities of CD patients undergoing intestinal resection, and to evaluate interactions between the bacteriome and mycobiome and their impact on the patients' outcome. We report a subgroup analysis of a prospective cohort study, focusing on 10 CD patients whose fecal samples were collected for bacterial 16S rRNA and fungal ITS2 genes next-generation sequencing the day before surgery and on the 5th or 6th postoperative day. No significant differences in bacterial and fungal diversity were observed between preoperative and postoperative stool samples. By in-depth analysis, significant postoperative abundance changes of bacteria and fungi and 17 interkingdom correlations were detected. Network analysis identified 13 microbial clusters in the perioperative gut communities, revealing symbiotic and competitive interactions. Relevant factors were gender, age, BMI, lifestyle habits (smoking, alcohol consumption) and surgical technique. Postoperative abundance changes and identified clusters were associated with clinical outcomes (length of hospital stay, complications) and levels of inflammatory markers. Our findings highlight the importance of dissecting the interactions of gut bacterial and fungal communities in CD patients and their potential influence on postoperative and disease outcomes.

Review article: The complex interplay between diet and Escherichia coli in inflammatory bowel disease.

Although no causative microbe has been yet identified or successfully targeted in the treatment of inflammatory bowel disease (IBD), the role of Escherichia coli in the pathogenesis of Crohn's disease has attracted considerable interest. In this review, we present a literature overview of the interactions between diet and E. coli and other Proteobacteria in the aetiology, outcomes and management of IBD and suggest future research directions. An extensive literature search was performed to identify in vitro studies and research in animal models that explored mechanisms by which dietary components can interact with E. coli or Proteobacteria to initiate or propagate gut inflammation. We also explored the effect diet and dietary therapies have on the levels of E. coli or Proteobacteria in patients with IBD. Preclinical data suggest that the Western diet and its components influence the abundance, colonisation and phenotypic behaviour of E. coli in the gut, which may in turn initiate or contribute to gut inflammation. In contrast, the Mediterranean diet and specific dietary fibres may abrogate these effects and protect from inflammation. There are limited data from clinical trials, mostly from patients with Crohn's disease during treatment with exclusive enteral nutrition, with findings often challenging observations from preclinical research. Data from patients with ulcerative colitis are sparse. Preclinical and some clinical trial data suggest that E. coli and other Proteobacteria interact with certain dietary components to promote gut inflammation. Well-designed clinical trials are required before dietary recommendations for disease management can be made.

Identification of differences in gene expression implicated in the Adherent-Invasive Escherichia coli phenotype during in vitro infection of intestinal epithelial cells.

Adherent-invasive Escherichia coli (AIEC) is strongly associated with the pathogenesis of Crohn's disease (CD). However, no molecular markers currently exist for AIEC identification. This study aimed to identify differentially expressed genes (DEGs) between AIEC and non-AIEC strains that may contribute to AIEC pathogenicity and to evaluate their utility as molecular markers. Comparative transcriptomics was performed on two closely related AIEC/non-AIEC strain pairs during Intestine-407 cell infection. DEGs were quantified by RT-qPCR in the same RNA extracts, as well as in 14 AIEC and 23 non-AIEC strains to validate the results across a diverse strain collection. Binary logistical regression was performed to identify DEGs whose quantification could be used as AIEC biomarkers. Comparative transcriptomics revealed 67 differences in expression between the two phenotypes in the strain pairs, 50 of which (81.97%) were corroborated by RT-qPCR. When explored in the whole strain collection, 29 DEGs were differentially expressed between AIEC and non-AIEC phenotypes (p-value < 0.042), and 42 genes between the supernatant fraction of infected cell cultures and the cellular fraction containing adhered and intracellular bacteria (p-value < 0.049). Notably, six DEGs detected in the strain collection were implicated in arginine biosynthesis and five in colanic acid synthesis. Furthermore, two biomarkers based on wzb and cueR gene expression were proposed with an accuracy of ≥ 85% in our strain collection. This is the first transcriptomic study conducted using AIEC-infected cell cultures. We have identified several genes that may be involved in AIEC pathogenicity, two of which are putative biomarkers for identification.

Transcriptome profiling of intact bowel wall reveals that PDE1A and SEMA3D are possible markers with roles in enteric smooth muscle apoptosis, proliferative disorders, and dysautonomia in Crohn's disease.

Background: Inflammatory bowel disease (IBD) is a complex and multifactorial inflammatory condition, comprising Crohn's disease (CD) and ulcerative colitis (UC). While numerous studies have explored the immune response in IBD through transcriptional profiling of the enteric mucosa, the subtle distinctions in the pathogenesis of Crohn's disease and ulcerative colitis remain insufficiently understood. Methods: The intact bowel wall specimens from IBD surgical patients were divided based on their inflammatory status into inflamed Crohn's disease (iCD), inflamed ulcerative colitis (iUC) and non-inflamed (niBD) groups for RNA sequencing. Differential mRNA GO (Gene Ontology), and KEGG (Kyoto Encyclopedia of Genes and Genomes), and GSEA (Gene Set Enrichment Analysis) bioinformatic analyses were performed with a focus on the enteric autonomic nervous system (ANS) and smooth muscle cell (SMC). The transcriptome results were validated by quantitative polymerase chain reaction (qPCR) and immunohistochemistry (IHC). Results: A total of 2099 differentially expressed genes were identified from the comparison between iCD and iUC. Regulation of SMC apoptosis and proliferation were significantly enriched in iCD, but not in iUC. The involved gene PDE1A in iCD was 4-fold and 1.5-fold upregulated at qPCR and IHC compared to that in iUC. Moreover, only iCD was significantly associated with the gene sets of ANS abnormality. The involved gene SEMA3D in iCD was upregulated 8- and 5-fold at qPCR and IHC levels compared to iUC. Conclusion: These findings suggest that PDE1A and SEMA3D may serve as potential markers implicated in enteric smooth muscle apoptosis, proliferative disorders, and dysautonomia specifically in Crohn's disease.