Intestinal Inflammation Modulates the Epithelial Response to Butyrate in Patients With Inflammatory Bowel Disease

Background: Butyrate-producing gut bacteria are reduced in patients with active inflammatory bowel disease (IBD), supporting the hypothesis that butyrate supplementation may be beneficial in this setting. Nonetheless, earlier studies suggest that the oxidation of butyrate in IBD patients is altered. We propose that inflammation may decrease epithelial butyrate consumption. Methods: Non-IBD controls and IBD patients were recruited for the study. Stool samples were used for short-chain fatty acid and bacterial butyryl CoA:acetate CoA-transferase quantification. Colonic biopsies and ex vivo differentiated epithelial organoids (d-EpOCs) treated with bu- tyrate and/or tumor necrosis factor alpha (TNFα) were used for analyzing the expression of transporters MCT1 and ABCG2, metabolic enzyme ACADS, and butyrate receptor GPR43, and for butyrate metabolism and consumption assays. Results: We observed that lower stool content of butyrate-producing bacteria in active IBD patients did not correlate with decreased bu- tyrate concentrations. Indeed, the intestinal epithelial expression of MCT1, ABCG2, ACADS, and GPR43 was altered in active IBD patients. Nonetheless, d-EpOCs derived from IBD patients showed SLC16A1 (gene encoding for MCT1 protein), ABCG2, ACADS, and GPR43 expres- sion levels comparable to controls. Moreover, IBD- and non-IBD-derived d-EpOCs responded similarly to butyrate, as assessed by transcriptional regulation. TNFα significantly altered SLC16A1, ABCG2, and GPR43 transcription in d-EpOCs, mimicking the expression profile observed in biopsies from active IBD patients and resulting in reduced butyrate consumption. Conclusions: We provide evidence that the response to butyrate is not intrinsically altered in IBD patients. However, TNFα renders the epithe- lium less responsive to this metabolite, defeating the purpose of butyrate supplementation during active inflammation

Differences in peripheral and tissue immune cell populations following haematopoietic stem cell transplantation in Crohn's disease patients

Background and aims: recent studies have shown the efficacy of autologous haematopoietic stem cell transplantation [HSCT] in severely refractory Crohn's disease [CD] patients. HSCT is thought to eliminate auto-reactive cells; however, no specific studies of immune reconstitution in CD patients are available. Methods: we followed a group of CD patients [n = 18] receiving autologous HSCT, with 50% of them achieving endoscopic drug-free remission. To elucidate the mechanisms driving efficacy, we monitored changes after HSCT in blood and intestine immune-cell composition. CD patients [n = 22] receiving anti-tumour necrosis factor [TNF]-α were included for comparison. Results: severe immune ablation followed by HSCT induced dramatic changes in both peripheral blood T and B cells in all patients regardless of the efficacy of the treatment. Endoscopic remission at week 52 following HSCT was associated with significant intestinal transcriptional changes. A comparison of the remission signature with that of anti-TNFα identified both common and unique genes in the HSCT-induced response. Based on deconvolution analysis of intestinal biopsy transcriptome data, we show that response to HSCT, but not to anti-TNFα, is associated with an expansion of naïve B-cells, as seen in blood, and a decrease in the memory resting T-cell content. As expected, endoscopic remission, in response to both HSCT and anti-TNFα, led to a significant reduction in intestinal neutrophil and M1 macrophage content. Conclusions: peripheral blood immune remodelling after HSCT does not predict efficacy. In contrast, a profound intestinal T-cell depletion that is maintained long after transplant is associated with mucosal healing following HSCT, but not anti-TNFα

Immune mechanisms involved in inducing remission in Crohn’s disease patients undergoing hematopoietic stem cell transplant

[eng] Crohn’s disease [CD] is a chronic inflammatory disease of the intestinal tract with considerable heterogeneity among affected patients in terms of disease phenotype and therapeutic responses. Despite the increase in the number of drugs approved for the management of CD, a significant percentage of patients remain unresponsive or lose response over time to treatments, and eventually require surgery to control disease activity and/or complications. Nonetheless, in a fraction of these refractory patients, intestinal resection may not be possible due to disease location, extension or previous surgeries. For such patients, autologous hematopoietic stem cell transplantation [HSCT] represents a potential salvage therapy despite the risks associated with this procedure. Stem cell transplantation is an accepted therapy for hematological disorders, aplastic anemia and immunodeficiencies. In the context of autoimmune diseases, the serendipitous benefits of transplantation were initially reported in patients suffering from both immune-mediated diseases and hematological disorders. This led to trials that have shown the efficacy of autologous HSCT in treating an array of autoimmune diseases including refractory severe multiple sclerosis, systemic lupus erythematosus, juvenile idiopathic arthritis, rheumatoid arthritis and, more recently, CD. The benefit of HSCT in autoimmunity is thought to originate from the ability of intense immune depletion to eliminate auto-reactive cells regardless of their specificity. This would lead to de novo generation of immune cells that could re-establish tolerance, although no objective evidence of this ‘resetting’ has been reported thus far. To explore this hypothesis, we group of CD patients [n = 18] receiving autologous HSCT, with 50% of them achieving endoscopic drug-free remission. To elucidate the mechanisms driving efficacy, we monitored changes after HSCT in blood and intestine immune-cell composition. CD patients [n = 22] receiving anti-tumor necrosis factor [TNF]-α were included for comparison. Severe immune ablation followed by HSCT induced dramatic changes in both peripheral blood T and B cells in all patients regardless of the efficacy of the treatment. Endoscopic remission at week 52 following HSCT was associated with significant intestinal transcriptional changes. A comparison of the remission signature with that of anti-TNFα identified both common and unique genes in the HSCT-induced response. Based on deconvolution analysis of intestinal biopsy transcriptome data, we show that response to HSCT, but not to anti-TNFα, is associated with an expansion of naïve B-cells, as seen in blood, and a decrease in the memory resting T- cell content. As expected, endoscopic remission, in response to both HSCT and anti-TNFα, led to a significant reduction in intestinal neutrophil and M1 macrophage content. HSCT dramatically renewed the pre-existing T-cell receptor [TCR] repertoire; however, persistent high-frequency TCR clones were detected in all patients in both blood and biopsy throughout the first year of follow-up. The number of persistent resident T cell clonotypes in tissue after treatment decreased in patients that achieve endoscopic remission. Furthermore, we found that low TCR peripheral diversity at baseline, and up to 1 year after HSCT, were associated with a lack of response to the immunoablative protocol. Peripheral blood immune remodeling after HSCT does not predict efficacy. In contrast, a profound intestinal T-cell depletion that is maintained long after transplant is associated with mucosal healing following HSCT, but not anti-TNFα.[spa] Se ha demostrado la eficacia del trasplante hematopoyético de células madre (THCM) en pacientes con enfermedad de Crohn severamente refractaria. La hipótesis aceptada es que el proceso de THCM elimina células auto-reactivas, devolviendo así la tolerancia. No obstante, no existen estudios específicos y exhaustivos sobre la reconstitución inmune en pacientes con enfermedad de Crohn tratados con THCM. En esta tesis, monitorizamos una cohorte de 18 pacientes con enfermedad de Crohn refractaria que recibieron trasplante autólogo de células madre hematopoyéticas, de los cuales el 50% consiguieron la remisión endoscópica sin tratamiento de mantenimiento. Para elucidar los mecanismos que determinan la eficacia, monitorizamos los cambios después de THCM en la composición de células inmunes tanto en sangre como en tejido intestinal. Como resultado, observamos que la ablación inmune producida durante el TCMH induce cambios dramáticos en las células T y B en sangre en todos los pacientes, independientemente de la eficacia del tratamiento. La remisión endoscópica en la semana 52 posterior al TCMH se asoció con cambios transcripcionales intestinales significativos. Una comparación de la transcripción en remisión con la de la remisión inducida por fármaco identificó genes comunes y únicos en la respuesta inducida por el TCMH. A través de un análisis de deconvolución de los datos del transcriptoma de la biopsia intestinal, mostramos que la respuesta al TCMH, pero no a fármaco inmuno- modulador, se asocia con una expansión de las células B naïve y una disminución en el contenido de células T memoria. Como se esperaba, la remisión endoscópica, en respuesta a tanto a TCMH como a fármaco, condujo a una reducción significativa en el contenido de neutrófilos intestinales y macrófagos M1 La remodelación inmune de la sangre periférica después del TCMH no predice la eficacia. No obstante, la depleción de las células T intestinales mantenida un año después del trasplante se asocia con la curación mucosa después del TCMH

Copy number variations of colorectal cancer by whole exome sequencing data

Curs 2013-2014Colorectal cancer (CRC) is the third most common cancer and the fourth leading cause of cancer death worldwide. About 85% of the cases of CRC are known to have chromosomal instability, an allelic imbalance at several chromosomal loci, and chromosome amplification and translocation. The aim of this study is to determine the recurrent copy number variant (CNV) regions present in stage II of CRC through whole exome sequencing, a rapidly developing targeted next-generation sequencing (NGS) technology that provides an accurate alternative approach for accessing genomic variations. 42 normal-tumor paired samples were sequenced by Illumina Genome Analyzer. Data was analyzed with Varscan2 and segmentation was performed with R package R-GADA. Summary of the segments across all samples was performed and the result was overlapped with DEG data of the same samples from a previous study in the group1. Major and more recurrent segments of CNV were: gain of chromosome 7pq(13%), 13q(31%) and 20q(75%) and loss of 8p(25%), 17p(23%), and 18pq(27%). This results are coincident with the known literature of CNV in CRC or other cancers, but our methodology should be validated by array comparative genomic hybridisation (aCGH) profiling, which is currently the gold standard for genetic diagnosis of CNV.Director/a: Victor Moreno, M. Luz Call

Multi-omic modelling of inflammatory bowel disease with regularized canonical correlation analysis

Personalized medicine requires finding relationships between variables that influence a patient's phenotype and predicting an outcome. Sparse generalized canonical correlation analysis identifies relationships between different groups of variables. This method requires establishing a model of the expected interaction between those variables. Describing these interactions is challenging when the relationship is unknown or when there is no pre-established hypothesis. Thus, our aim was to develop a method to find the relationships between microbiome and host transcriptome data and the relevant clinical variables in a complex disease, such as Crohn's disease

Dissecting common and unique effects of anti-alpha4beta7 and anti-tumor necrosis factor treatment in ulcerative colitis

Background and Aims:Vedolizumab is an anti-α4β7 antibody approved for the treatment of ulcerative colitis [UC]. Although it is assumed that vedolizumab blocks intestinal homing of lymphocytes, its effects on different intestinal cell populations are not fully stablished. In order to establish the unique mechanisms of action of vedolizumab in UC patients, we compared its effects to those induced by anti-tumour necrosis factor [TNF]. Methods:patients with active UC [endoscopic Mayo score >1] starting vedolizumab [n = 33] or anti-TNF [n = 45] and controls [n = 22] were included. Colon biopsies [at weeks 0, 14 and 46] and blood samples [at weeks 0, 2, 6, 14, 30 and 46] were used for cell phenotyping, transcriptional analysis [qPCR], and to measure receptor occupancy. Results:Vedolizumab, in contrast to anti-TNF, significantly reduced the proportion of α4β7+ cells within intestinal T subsets while preserving the percentage of α4β7+ plasma cells. The marked decrease in α4β7 did not change the percentage of colonic αEβ7+ cells [at 46 weeks]. Both vedolizumab and anti-TNF significantly downregulated inflammation-related genes in the colon of responders [Mayo score < 2]. Moreover, both treatments significantly decreased the percentage of intestinal, but not blood, total lymphocytes [T and plasma cells], as well as the proportion of α4β1+ cells within intestinal T lymphocytes. Conclusions:Our data show that while vedolizumab and anti-TNF block two unrelated targets, they induce remarkably similar effects. On the other hand, vedolizumab's unique mechanism of action relies on blocking intestinal trafficking of α4β7 T cells, despite effectively binding to B and plasma cells that express α4β7