Fiber rich food suppressed airway inflammation, GATA3 + Th2 cells, and FcεRIα+ eosinophils in asthma.

Background: Allergic Asthma is a disease presenting various endotypes and no current therapies act curative but alleviate disease symptoms. Dietary interventions are gaining increasing importance in regulating immune responses. Furthermore, short chain fatty acids (SFCA), as the main products of dietary fiber's fermentation by the gut bacteria, ameliorate the pathogenesis and disease burden of different illnesses including asthma. Nevertheless, the connection and crosstalk between the gut and lung is poorly understood. Objective: In this work, the role of high fiber diet on the development of allergic asthma at baseline and after exacerbation of disease induced by respiratory viruses was investigated. Methods: Hereby, SCFA in serum of asthmatic and non-asthmatic pre-school children before and after airway disease symptoms were analyzed. Moreover, the effect of high fiber diet in vivo in a murine model of house dust mite extract (HDM) induced allergic asthma and in the end in isolated lung and spleen cells infected ex vivo with Rhinovirus was analyzed. Results: In this study, a decrease of the SCFA 3-Hydroxybutyric acid in serum of asthmatic children after symptomatic episodes at convalescent visit as compared to asthmatic and control children at baseline visit was observed. In experimental asthma, in mice fed with high fiber diet, a reduced lung GATA3 + Th2 type mediated inflammation, mucus production and collagen deposition and expression of Fc epsilon receptor Ia (FcεRIa) in eosinophils was observed. By contrast, the CD8+ memory effector T cells were induced in the lungs of asthmatic mice fed with high fiber diet. Then, total lung cells from these asthmatic mice fed with either standard food or with fiber rich food were infected with RV ex vivo. Here, RV1b mRNA was found significantly reduced in the lung cells derived from fiber rich food fed mice as compared to those derived from standard food fed asthmatic mice. Looking for the mechanism, an increase in CD8+ T cells in RV infected spleen cells derived from fiber rich fed asthmatic mice, was observed. Conclusion: Convalescent preschool asthmatic children after a symptomatic episode have less serum ß-Hydroxybutyric acid as compared to control and asthmatic children at baseline visit. Fiber rich diet associated with anti-inflammatory effects as well as anti-allergic effects by decreasing Type 2 and IgE mediated immune responses and inducing CD8+ memory effector T cells in a murine model of allergic asthma. Finally, ex vivo infection with Rhinovirus (RV) of total lung cells from asthmatic mice fed with fiber rich food led to a decreased RV load as compared to mice fed with standard food. Moreover, spleen cells derived from asthmatic mice fed with fiber rich food induced CD8+ T cells after ex vivo infection with RV. Clinical implications: Dietary interventions with increased content in natural fibers like pectins would ameliorate asthma exacerbations. Moreover, respiratory infection in asthma downregulated SCFA in the gut contributing to asthma exacerbations.

Etrolizumab-s fails to control E-Cadherin-dependent co-stimulation of highly activated cytotoxic T cells.

Despite promising preclinical and earlier clinical data, a recent phase III trial on the anti-ß7 integrin antibody etrolizumab in Crohn's disease (CD) did not reach its primary endpoint. The mechanisms leading to this outcome are not well understood. Here we characterize the ß7+ T cell compartment from patients with CD in comparison to cells from individuals without inflammatory bowel disease. By flow cytometric, transcriptomic and functional profiling of circulating T cells, we find that triple-integrin-expressing (α4+ß7+ß1hi) T cells have the potential to home to the gut despite α4ß7 blockade and have a specific cytotoxic signature. A subset of triple-integrin-expressing cells readily acquires αE expression and could be co-stimulated via E-Cadherin-αEß7 interactions in vitro. Etrolizumab-s fails to block such αEß7 signalling at high levels of T cell stimulation. Consistently, in CD patients treated with etrolizumab, T cell activation correlates with cytotoxic signatures. Collectively, our findings might add one important piece to the puzzle to explain phase III trial results with etrolizumab, while they also highlight that αEß7 remains an interesting target for future therapeutic approaches in inflammatory bowel disease.

Long-term outcomes of cyclosporin induction and ustekinumab maintenance combination therapy in patients with steroid-refractory acute severe ulcerative colitis.

Background: Effective management of patients with acute severe ulcerative colitis (ASUC) is a major challenge and there remains a paucity of available maintenance treatment options after efficacious cyclosporin induction therapy. Objectives: We investigated the long-term effectiveness and safety of cyclosporin and ustekinumab combination therapy in patients with steroid refractory ASUC. Design: Monocentric, prospective study. Methods: We included patients with steroid refractory ASUC with multiple failed prior advanced therapies, who were treated with cyclosporin and ustekinumab combination therapy. Results: Among the 11 included patients, 10 had prior failure to infliximab and 8 failed at least three previous biological therapies. The mean baseline Mayo and Lichtiger scores were 10.9 (9-12) and 13.3 (11-14), respectively. Ustekinumab was initiated 3.2 weeks (1-8) after initiation of cyclosporin treatment and combination therapy was continued for a mean of 11.5 (4-20) weeks. Clinical response was achieved in six patients at week 16 and clinical steroid-free clinical remission in five patients at week 48. Endoscopic remission was achieved in five patients at week 16 and together with histological remission in five patients at week 52. Intestinal ultrasound demonstrated mean bowel wall thickening in the sigmoid colon of 5.5 mm at baseline and 3.5 mm at week 52, respectively. Two patients had to undergo colectomy (mean 4.5 months, range 3-6) and three stopped ustekinumab therapy due to ineffectiveness. Overall, combination therapy was well tolerated. Conclusion: Combination of cyclosporin and ustekinumab therapy allowed nearly half of ASUC patients to reach clinical and endoscopic remission after 52 weeks, warranting further studies. Trial registration: Not applicable.

Effects of cyclosporin and ustekinumab combination therapy in acute severe ulcerative colitis In this study, we looked at how to treat patients with a severe form of ulcerative colitis, a type of inflammatory bowel disease, when the usual treatments don't work. We tested a combination of two drugs, cyclosporine and ustekinumab, to see if it could help these patients in the long term. We included eleven patients who had already tried many other treatments and didn't get better. Most of them had also tried a drug called infliximab and had failed at least three other biological therapies. At the start, these patients were very sick, with high scores on disease activity measures. We gave them ustekinumab in addtion after a therapy with cyclosporin had been started before. The combination therapy continued for an average of almost 12 weeks. After 16 weeks, six patients showed improvement in their symptoms, and five were able to stop taking steroids. Five patients also had their colon lining looking healthy again when we looked inside with a scope after 16 weeks. And after 52 weeks, five patients had normal colon lining and healthy tissue under the microscope. Ultrasound showed that the thickness of their colon wall had decreased. Unfortunately, two patients had to have surgery to remove their colon, and three had to stop taking ustekinumab because it didn't help them. Overall, the combination therapy was safe and well-tolerated. In conclusion, combining cyclosporin and ustekinumab helped about half of the patients with severe ulcerative colitis get better and have healthy colon lining after 52 weeks. This suggests that more research is needed to understand the benefits of this treatment in these patients.

Blocking GPR15 counteracts integrin-dependent T cell gut homing in vivo.

BACKGROUND AND AIMS: The G protein coupled receptor GPR15 is expressed on and functionally important for T cells homing to the large intestine. However, the precise mechanisms by which GPR15 controls gut homing have been unclear. Thus, we aimed to elucidate these mechanisms as well as to explore the potential of targeting GPR15 for interfering with T cell recruitment to the colon in IBD. METHODS: We used dynamic adhesion and transmigration assays as well as a humanized in vivo model of intestinal cell trafficking to study GPR15-dependent effects on gut homing. Moreover, we analysed GPR15 and integrin expression in patients with and without IBD cross-sectionally and longitudinally. RESULTS: GPR15 controlled T cell adhesion to MAdCAM-1 and VCAM-1 upstream of α4ß7 and α4ß1 integrin, respectively. Consistently, high co-expression of these integrins with GPR15 was found on T cells from patients with IBD and GPR15 also promoted T cell recruitment to the colon in humanized mice. Anti-GPR15 antibodies effectively blocked T cell gut homing in vitro and in vivo. In vitro data as well as observations in a cohort of patients treated with vedolizumab suggest that this might be more effective than inhibiting α4ß7. CONCLUSIONS: GPR15 seems to have a broad, but organ-selective impact on T cell trafficking and is therefore a promising target for future therapy of IBD. Further studies are needed.

ATP citrate lyase (ACLY)-dependent immunometabolism in mucosal T cells drives experimental colitis in vivo.

OBJECTIVE: Mucosal T cells play a major role in inflammatory bowel disease (IBD). However, their immunometabolism during intestinal inflammation is poorly understood. Due to its impact on cellular metabolism and proinflammatory immune cell function, we here focus on the enzyme ATP citrate lyase (ACLY) in mucosal T cell immunometabolism and its relevance for IBD. DESIGN: ACLY expression and its immunometabolic impact on colitogenic T cell function were analysed in mucosal T cells from patients with IBD and in two experimental colitis models. RESULTS: ACLY was markedly expressed in colon tissue under steady-state conditions but was significantly downregulated in lamina propria mononuclear cells in experimental dextran sodium sulfate-induced colitis and in CD4+ and to a lesser extent in CD8+ T cells infiltrating the inflamed gut in patients with IBD. ACLY-deficient CD4+ T cells showed an impaired capacity to induce intestinal inflammation in a transfer colitis model as compared with wild-type T cells. Assessment of T cell immunometabolism revealed that ACLY deficiency dampened the production of IBD-relevant cytokines and impaired glycolytic ATP production but enriched metabolites involved in the biosynthesis of phospholipids and phosphatidylcholine. Interestingly, the short-chain fatty acid butyrate was identified as a potent suppressor of ACLY expression in T cells, while IL-36α and resolvin E1 induced ACLY levels. In a translational approach, in vivo administration of the butyrate prodrug tributyrin downregulated mucosal infiltration of ACLYhigh CD4+ T cells and ameliorated chronic colitis. CONCLUSION: ACLY controls mucosal T cell immunometabolism and experimental colitis. Therapeutic modulation of ACLY expression in T cells emerges as a novel strategy to promote the resolution of intestinal inflammation.

NLRP3 Inhibition Leads to Impaired Mucosal Fibroblast Function in Patients with Inflammatory Bowel Diseases.

BACKGROUND AND AIMS: Inflammatory bowel diseases (IBD) are characterized by mucosal inflammation and sequential fibrosis formation, but the exact role of the hyperactive NLRP3 inflammasome in these processes is unclear. Thus, we studied the expression and function of the NLRP3 inflammasome in the context of inflammation and fibrosis in IBD. METHODS: We analysed intestinal NLRP3 expression in mucosal immune cells and fibroblasts from IBD patients and NLRP3-associated gene expression via single-cell RNA sequencing and microarray analyses. Furthermore, cytokine secretion of NLRP3 inhibitor treated blood and mucosal cells, as well as proliferation, collagen production, and cell death of NLRP3 inhibitor treated intestinal fibroblasts from IBD patients were studied. RESULTS: We found increased NLRP3 expression in the inflamed mucosa of IBD patients and NLRP3 inhibition led to reduced IL-1ß and IL-18 production in blood cells and diminished the bioactive form of mucosal IL-1ß. Single cell analysis identified overlapping expression patterns of NLRP3 and IL-1ß in classically activated intestinal macrophages and we also detected NLRP3 expression in CD163+ macrophages. In addition, NLRP3 expression was also found in intestinal fibroblasts from IBD patients. Inhibition of NLRP3 led to reduced proliferation of intestinal fibroblasts, which was associated with a marked decrease in production of collagen type I and type VI in IBD patients. Moreover, NLRP3 inhibition in intestinal fibroblasts induced autophagy, a cellular process involved in collagen degradation. CONCLUSIONS: In the presented study, we demonstrate that inhibiting NLRP3 might pave the way for novel therapeutic approaches in IBD, especially to prevent the severe complication of intestinal fibrosis formation.

IL-20 controls resolution of experimental colitis by regulating epithelial IFN/STAT2 signalling.

OBJECTIVE: We sought to investigate the role of interleukin (IL)-20 in IBD and experimental colitis. DESIGN: Experimental colitis was induced in mice deficient in components of the IL-20 and signal transducer and activator of transcription (STAT)2 signalling pathways. In vivo imaging, high-resolution mini-endoscopy and histology were used to assess intestinal inflammation. We further used RNA-sequencing (RNA-Seq), RNAScope and Gene Ontology analysis, western blot analysis and co-immunoprecipitation, confocal microscopy and intestinal epithelial cell (IEC)-derived three-dimensional organoids to investigate the underlying molecular mechanisms. Results were validated using samples from patients with IBD and non-IBD control subjects by a combination of RNA-Seq, organoids and immunostainings. RESULTS: In IBD, IL20 levels were induced during remission and were significantly higher in antitumour necrosis factor responders versus non-responders. IL-20RA and IL-20RB were present on IECs from patients with IBD and IL-20-induced STAT3 and suppressed interferon (IFN)-STAT2 signalling in these cells. In IBD, experimental dextran sulfate sodium (DSS)-induced colitis and mucosal healing, IECs were the main producers of IL-20. Compared with wildtype controls, Il20-/-, Il20ra-/- and Il20rb-/- mice were more susceptible to experimental DSS-induced colitis. IL-20 deficiency was associated with increased IFN/STAT2 activity in mice and IFN/STAT2-induced necroptotic cell death in IEC-derived organoids could be markedly blocked by IL-20. Moreover, newly generated Stat2ΔIEC mice, lacking STAT2 in IECs, were less susceptible to experimental colitis compared with wildtype controls and the administration of IL-20 suppressed colitis activity in wildtype animals. CONCLUSION: IL-20 controls colitis and mucosal healing by interfering with the IFN/STAT2 death signalling pathway in IECs. These results indicate new directions for suppressing gut inflammation by modulating IL-20-controlled STAT2 signals.

Diagnostic Endoscopic Ultrasound in Pancreatology: Focus on Normal Variants and Pancreatic Masses.

Background: Endoscopic ultrasound (EUS) is a main tool in gastroenterology for both diagnosis and exclusion of pancreatic pathology. It allows minimally invasive assessment of various diseases or anatomic variations affecting the pancreas also with the help of new Doppler technologies, elastography, contrast-enhanced imaging including post hoc image processing with quantification analyses, three-dimensional reconstruction, and artificial intelligence. EUS also allows interventional direct access to the pancreatic parenchyma and the retroperitoneal space, to the pancreatic duct, pancreatic masses, cysts, and vascular structures. Summary: This review aimed to summarize new developments of EUS in the field of pancreatology. We highlight the role of EUS in evaluating pancreatic pathology by describing normal anatomic variants like pancreas divisum, pancreatic lipomatosis, pancreatic fibrosis in the elderly and characterizing pancreatic masses, both in the context of chronic pancreatitis and within healthy pancreatic parenchyma. EUS is considered the optimal imaging modality for pancreatic masses of uncertain dignity and allows both cytological diagnosis and histology, which is essential not only for neoplastic conditions but also for tailoring therapy for benign inflammatory conditions. Key Messages: EUS plays an indispensable role in pancreatology and the development of new diagnostic and interventional approaches to the retroperitoneal space and the pancreas exponentially increased over the last years. The development of computer-aided diagnosis and artificial intelligence algorithms hold the potential to overcome the obstacles associated with interobserver variability and will most likely support decision-making in the management of pancreatic disease.

Endoscopic Ultrasound in Pancreatology: Focus on Inflammatory Diseases and Interventions.

Background: Endoscopic ultrasound (EUS) is a main tool in pancreatology for both diagnosis and therapy. It allows minimally invasive differentiation of various diseases, with a minimal degree of inflammation or anatomic variations. EUS also enables interventional direct access to the pancreatic parenchyma and the retroperitoneal space, the pancreatic duct, the pancreatic masses, cysts, vascular structures for diagnostic and therapeutic purposes. Summary: This review aimed to summarize the new developments of EUS in the field of pancreatology, with special interest on inflammation and interventions. EUS enables way to perform pseudocyst drainage, necrosectomy, transenteral drainage and transenteric access of the main pancreatic duct, or the direct visualization or therapy of vascular structures adjacent to the pancreas. Key Messages: EUS has a deep impact on pancreatology, and the development of new diagnostic and interventional approaches to the retroperitoneal space and the pancreas has increased in the last years exponentially, allowing minimal invasive diagnostics and therapy and avoiding surgery and percutaneous therapy.

Healing of the epithelial barrier in the ileum is superior to endoscopic and histologic remission for predicting major adverse outcomes in ulcerative colitis.

Background: Achieving endoscopic remission is a key therapeutic goal in patients with ulcerative colitis (UC) that is associated with favorable long-term disease outcomes. Here, we prospectively compared the predictive value of endoscopic and/or histologic remission against ileal barrier healing for predicting long-term disease behavior in a large cohort of UC patients in clinical remission. Methods: At baseline, UC patients in clinical remission underwent ileocolonoscopy with assessment of ileal barrier function by confocal endomicroscopy. Endoscopic and histologic disease activity and ileal barrier healing were scored using validated scores. During subsequent follow-up (FU), patients were closely monitored for clinical disease activity and occurrence of major adverse outcomes (MAO) defined as the following: disease relapse; UC-related hospitalization; UC-related surgery; necessity for initiation or dose escalation of systemic steroids, immunosuppressants, small molecules or biological therapy. Results: Of the 73 UC patients included, 67% experienced MAO during a mean FU of 25 months. The probability of MAO-free survival was significantly higher in UC patients with endoscopic and/or histologic remission compared to patients with endoscopically and/or histologically active disease. Ileal barrier healing on endomicroscopy was highly accurate for predicting the further course of UC and outcompeted endoscopic and histologic remission for predicting MAO-free survival. Conclusion: Ileal barrier healing in clinically remittent UC patients can accurately predict future MAO development and is superior in its predictive capabilities than endoscopic and histologic remission. Ileal barrier healing therefore represents a novel and superior surrogate parameter for stratification of UC patients according to their risk for development of complicated disease behavior. Clinical trial registration: https://classic.clinicaltrials.gov/ct2/show/NCT05157750, identifier NCT05157750.

IL-3 receptor signalling suppresses chronic intestinal inflammation by controlling mechanobiology and tissue egress of regulatory T cells.

IL-3 has been reported to be involved in various inflammatory disorders, but its role in inflammatory bowel disease (IBD) has not been addressed so far. Here, we determined IL-3 expression in samples from patients with IBD and studied the impact of Il3 or Il3r deficiency on T cell-dependent experimental colitis. We explored the mechanical, cytoskeletal and migratory properties of Il3r -/- and Il3r +/+ T cells using real-time deformability cytometry, atomic force microscopy, scanning electron microscopy, fluorescence recovery after photobleaching and in vitro and in vivo cell trafficking assays. We observed that, in patients with IBD, the levels of IL-3 in the inflamed mucosa were increased. In vivo, experimental chronic colitis on T cell transfer was exacerbated in the absence of Il-3 or Il-3r signalling. This was attributable to Il-3r signalling-induced changes in kinase phosphorylation and actin cytoskeleton structure, resulting in increased mechanical deformability and enhanced egress of Tregs from the inflamed colon mucosa. Similarly, IL-3 controlled mechanobiology in human Tregs and was associated with increased mucosal Treg abundance in patients with IBD. Collectively, our data reveal that IL-3 signaling exerts an important regulatory role at the interface of biophysical and migratory T cell features in intestinal inflammation and suggest that this might be an interesting target for future intervention.

Adhesion GPCR Gpr126 (Adgrg6) Expression Profiling in Zebrafish, Mouse, and Human Kidney.

Adhesion G protein-coupled receptors (aGPCRs) comprise the second-largest class of GPCRs, the most common target for approved pharmacological therapies. aGPCRs play an important role in development and disease and have recently been associated with the kidney. Several aGPCRs are expressed in the kidney and some aGPCRs are either required for kidney development or their expression level is altered in diseased kidneys. Yet, general aGPCR function and their physiological role in the kidney are poorly understood. Here, we characterize in detail Gpr126 (Adgrg6) expression based on RNAscope® technology in zebrafish, mice, and humans during kidney development in adults. Gpr126 expression is enriched in the epithelial linage during nephrogenesis and persists in the adult kidney in parietal epithelial cells, collecting ducts, and urothelium. Single-cell RNAseq analysis shows that gpr126 expression is detected in zebrafish in a distinct ionocyte sub-population. It is co-detected selectively with slc9a3.2, slc4a4a, and trpv6, known to be involved in apical acid secretion, buffering blood or intracellular pH, and to maintain high cytoplasmic Ca2+ concentration, respectively. Furthermore, gpr126-expressing cells were enriched in the expression of potassium transporter kcnj1a.1 and gcm2, which regulate the expression of a calcium sensor receptor. Notably, the expression patterns of Trpv6, Kcnj1a.1, and Gpr126 in mouse kidneys are highly similar. Collectively, our approach permits a detailed insight into the spatio-temporal expression of Gpr126 and provides a basis to elucidate a possible role of Gpr126 in kidney physiology.

Case Report: IBD-like colitis following CAR T cell therapy for diffuse large B cell lymphoma.

Chimeric antigen receptor (CAR) T cell therapy has become a new mainstay in the treatment of several hematologic malignancies, but the spectrum of associated complications is still incompletely defined. Here, we report the case of a 70-year-old female patient treated with tisagenlecleucel for diffuse large B cell lymphoma (DLBCL), who developed chronic diarrhea with characteristics of inflammatory bowel disease (IBD)-like colitis. CAR T cells were substantially enriched in the colon lamina propria and other diagnoses were ruled out. Thus, we conclude that IBD-like colitis in this patient was associated to CAR T cell therapy and needs to be considered as a rare potential complication.

Washout appearance of hepatocellular carcinomas using standardized contrast-enhanced ultrasound (CEUS) including an extended late phase observation - Real-world data from the prospective multicentre DEGUM study.

BACKGROUND: Non-invasive diagnosis of hepatocellular carcinoma (HCC) in contrast-enhanced ultrasound (CEUS) is based on the combination of arterial phase hyperenhancement (APHE) and subsequent late (>60 seconds) and mild contrast washout (WO). Whereas APHE is seen in the majority of HCC, wash-out pattern may vary in onset and intensity. In some HCC lesions, even no washout is seen at all. OBJECTIVE: Our prospective multicentre DEGUM HCC CEUS study aimed at identifying typical and atypical washout appearance of HCC in a real-life setting. METHODS: High-risked patients for HCC with focal liver lesions upon B-mode ultrasound were recruited prospectively. In a multicentre real-life setting, a standardised CEUS examination including an extended late phase up to 6 minutes was performed. CEUS patterns of HCC were recorded, and onset and intensity of washout appearance were assessed with respect to patient and tumour characteristics. Histological findings served as reference standard. RESULTS: In 230/316 HCC (72.8%), a CEUS pattern of APHE followed by WO was observed. In 158 cases (68.7%), WO was typical (onset > 60 seconds, mild intensity). 72 cases (31.3%) showed marked and / or early WO, whereas in 41 HCCs (13%), APHE was followed by sustained isoenhancement.Atypical WO upon CEUS was associated with macroinvasion of the liver vessels, portal vein thrombosis and diffuse growth pattern, but not tumour size and histological grading. CONCLUSIONS: In a prospective multicentre real-life setting, APHE is followed by atypical washout appearance or no washout at all in almost half of the HCCs with APHE. The examiner has to bear in mind that in spite of a characteristic APHE in HCCs, washout appearance can be atypical in CEUS, especially in HCCs with macrovascular invasion or diffuse growth pattern.

Protective effect of the novel calcineurin inhibitor voclosporin in experimental colitis.

Background and aims: Acute severe steroid-refractory ulcerative colitis remains a medically challenging condition with frequent need of surgery. It can be treated with the calcineurin inhibitor cyclosporine A with the need for therapeutic drug monitoring and significant toxicity. Recently, a novel calcineurin inhibitor, voclosporin, has been approved for the treatment of lupus nephritis with no need for therapeutic drug monitoring and an improved long-term safety profile. However, the therapeutic effect of voclosporin in acute severe steroid-refractory ulcerative colitis is still uncertain. We aimed to assess the therapeutic potential of voclosporin to ameliorate inflammation in an experimental model of colitis. Methods: We used the dextran sodium sulfate-induced model of colitis in C57BL/6 J wildtype mice treated with either cyclosporine A, voclosporin or solvent control. We employed endoscopy, histochemistry, immunofluorescence, bead-based multiplex immunoassays and flow cytometry to study the therapeutic effect of calcineurin inhibitors in a preventive setting. Results: Acute colitis was induced by dextran sodium sulfate characterized by weight loss, diarrhea, mucosal erosions and rectal bleeding. Both cyclosporine A and voclosporin strongly ameliorated the course of disease and reduced colitis severity in a similar manner. Conclusion: Voclosporin was identified as biologically effective in a preclinical model of colitis and may be a potential therapeutic option in treating acute severe steroid-refractory ulcerative colitis.

Differential Effects of Ontamalimab Versus Vedolizumab on Immune Cell Trafficking in Intestinal Inflammation and Inflammatory Bowel Disease.

BACKGROUND AND AIMS: The anti-MAdCAM-1 antibody ontamalimab demonstrated efficacy in a phase II trial in ulcerative colitis and results of early terminated phase III trials are pending, but its precise mechanisms of action are still unclear. Thus, we explored the mechanisms of action of ontamalimab and compared it to the anti-α4ß7 antibody vedolizumab. METHODS: We studied MAdCAM-1 expression with RNA sequencing and immunohistochemistry. The mechanisms of action of ontamalimab were assessed with fluorescence microscopy, dynamic adhesion and rolling assays. We performed in vivo cell trafficking studies in mice and compared ontamalimab and vedolizumab surrogate [-s] antibodies in experimental models of colitis and wound healing. We analysed immune cell infiltration under anti-MAdCAM-1 and anti-α4ß7 treatment by single-cell transcriptomics and studied compensatory trafficking pathways. RESULTS: MAdCAM-1 expression was increased in active inflammatory bowel disease. Binding of ontamalimab to MAdCAM-1 induced the internalization of the complex. Functionally, ontamalimab blocked T cell adhesion similar to vedolizumab, but also inhibited L-selectin-dependent rolling of innate and adaptive immune cells. Despite conserved mechanisms in mice, the impact of ontamalimab-s and vedolizumab-s on experimental colitis and wound healing was similar. Single-cell RNA sequencing demonstrated enrichment of ontamalimab-s-treated lamina propria cells in specific clusters, and in vitro experiments indicated that redundant adhesion pathways are active in these cells. CONCLUSIONS: Ontamalimab has unique and broader mechanisms of action compared to vedolizumab. However, this seems to be compensated for by redundant cell trafficking circuits and leads to similar preclinical efficacy of anti-α4ß7 and anti-MAdCAM-1 treatment. These results will be important for the interpretation of pending phase III data.

Manual cell selection in single cell transcriptomics using scSELpy supports the analysis of immune cell subsets.

Introduction: Single cell RNA sequencing plays an increasing and indispensable role in immunological research such as in the field of inflammatory bowel diseases (IBD). Professional pipelines are complex, but tools for the manual selection and further downstream analysis of single cell populations are missing so far. Methods: We developed a tool called scSELpy, which can easily be integrated into Scanpy-based pipelines, allowing the manual selection of cells on single cell transcriptomic datasets by drawing polygons on various data representations. The tool further supports the downstream analysis of the selected cells and the plotting of results. Results: Taking advantage of two previously published single cell RNA sequencing datasets we show that this tool is useful for the positive and negative selection of T cell subsets implicated in IBD beyond standard clustering. We further demonstrate the feasibility for subphenotyping T cell subsets and use scSELpy to corroborate earlier conclusions drawn from the dataset. Moreover, we also show its usefulness in the context of T cell receptor sequencing. Discussion: Collectively, scSELpy is a promising additive tool fulfilling a so far unmet need in the field of single cell transcriptomic analysis that might support future immunological research.

Anti-ß7 integrin treatment impedes the recruitment on non-classical monocytes to the gut and delays macrophage-mediated intestinal wound healing.

BACKGROUND: Closing mucosal defects to reach mucosal healing is an important goal of therapy in inflammatory bowel disease (IBD). Among other cells, monocyte-derived macrophages are centrally involved in such intestinal wound healing. We had previously demonstrated that the anti-α4ß7 integrin antibody vedolizumab blocks the recruitment of non-classical monocytes as biased progenitors of wound healing macrophages to the gut and delays wound healing. However, although important for the interpretation of disappointing results in recent phase III trials in IBD, the effects of the anti-ß7 antibody etrolizumab on wound healing are unclear so far. METHODS: We analyzed the expression of etrolizumab targets on human and mouse monocyte subsets by flow cytometry and assessed their function in adhesion and homing assays. We explored wound-associated monocyte recruitment dynamics with multi-photon microscopy and compared the effects of etrolizumab and vedolizumab surrogate (-s) antibodies on experimental wound healing and wound-associated macrophage abundance. Finally, we investigated wound healing macrophage signatures in the large intestinal transcriptome of patients with Crohn's disease treated with etrolizumab. RESULTS: Human and mouse non-classical monocytes expressed more αEß7 integrin than classical monocytes and were a target of etrolizumab-s, which blocked non-classical monocyte adhesion to MAdCAM-1 and E-Cadherin as well as gut homing in vivo. Intestinal wound healing was delayed on treatment with etrolizumab-s along with a reduction of peri-lesional wound healing macrophages. Wound healing macrophage signatures in the colon of patients with Crohn's disease were substantially down-regulated on treatment with etrolizumab, but not with placebo. CONCLUSIONS: Combined blockade of αEß7 and α4ß7 with etrolizumab seems to exceed the effect of anti-α4ß7 treatment on intestinal wound healing, which might help to inform further investigations to understand the recent observations in the etrolizumab phase III trial program.

Artificial intelligence for the classification of focal liver lesions in ultrasound - a systematic review.

Focal liver lesions are detected in about 15% of abdominal ultrasound examinations. The diagnosis of frequent benign lesions can be determined reliably based on the characteristic B-mode appearance of cysts, hemangiomas, or typical focal fatty changes. In the case of focal liver lesions which remain unclear on B-mode ultrasound, contrast-enhanced ultrasound (CEUS) increases diagnostic accuracy for the distinction between benign and malignant liver lesions. Artificial intelligence describes applications that try to emulate human intelligence, at least in subfields such as the classification of images. Since ultrasound is considered to be a particularly examiner-dependent technique, the application of artificial intelligence could be an interesting approach for an objective and accurate diagnosis. In this systematic review we analyzed how artificial intelligence can be used to classify the benign or malignant nature and entity of focal liver lesions on the basis of B-mode or CEUS data. In a structured search on Scopus, Web of Science, PubMed, and IEEE, we found 52 studies that met the inclusion criteria. Studies showed good diagnostic performance for both the classification as benign or malignant and the differentiation of individual tumor entities. The results could be improved by inclusion of clinical parameters and were comparable to those of experienced investigators in terms of diagnostic accuracy. However, due to the limited spectrum of lesions included in the studies and a lack of independent validation cohorts, the transfer of the results into clinical practice is limited.