Pediatric Gastroenterology & Nutrition
Mucosal Immunology and Intestinal Homeostasis
Within the laboratory of Pediatrics the research division Gastroenterology and Nutrition is headed by Janneke N. Samsom. The Samsom lab has a focus on mucosal immunology.
The mucosal surfaces of the gastrointestinal tract have developed as the largest surface area of the body that is in contact with the external environment. Not only do most pathogens enter through this site, the intestinal mucosa is continuously exposed to a large variety of harmless antigens, such as dietary proteins and constituents of commensal bacteria. To regulate this high antigenic pressure the human gut holds approximately 50 x109 lymphocytes, a large proportion of all immune cells in the body. In recent years it has become clear that both innate and acquired regulatory immune responses are essential for the development of oral tolerance to harmless antigens.
Our research is focused on identifying immune regulatory processes* that are pivotal to intestinal homeostasis. By developing multiple murine models, we have identified that functional mucosal regulatory T cells differentiate in mucosa draining lymphoid tissue in the presence of specific mucosal factors. We have established a humanized mouse model of gluten tolerance and discovered that tolerance to gluten is mediated by IL-10 secreting regulatory T cells, a finding opening new avenues of research for celiac disease*. In our translational inflammatory bowel disease* (IBD) work we focus on dissecting inflammatory pathways that classify IBD patients in subgroups and yield parameters to characterize their disease subtype. Through analysis of infants with genetic deficiencies we are uncovering pathways that are pivotal for intestinal tolerance and are identifying patterns of inflammation that evolve from a particular defect.
Translation to the clinic
The solid, very direct link between the research lab and the clinical Pediatric gastroenterology department is of great value. Together with the group of Prof. J.C. Escher and Dr. L. de Ridder, our research has a bidirectional approach: key questions from the clinic concerning pathology, diagnosis and treatment of IBD are translated to an experimental setting and vice versa fundamental data are implemented in patient diagnostics and design for new treatment strategies. In parallel, similar initiatives are developed in the field of celiac disease via a strong collaboration with dr. M. Groeneweg in the Maasstad Hospital and by taking advantage of our Dutch Celiac Disease Consortium network involving the LUMC, UMCG, WUR and AMC. This translation is even further extended in the new TIMID consortium, a disease overarching research initiative that our lab coordinates.
The clinical department of Pediatric Gastroenterology of the Sophia Children’s hospital is involved in multiple international drug studies in pediatric IBD patients, both initiated by industry as well as investigator-initiated. The department has the largest cohort of pediatric IBD patients in the Netherlands. Hankje C. Escher is the initiator and leader of EUROKIDS, a web-based European registry of new pediatric IBD patients. This web registry is ongoing since 2004 and currently harbors data on demographics, diagnostic work-up, and disease phenotype of over 4000 patients. As a member of the Horizon 2020 EU PIBD Set Quality consortium (PHC-18-2015) the laboratory and clinic are joining forces to develop new investigative tools as classifiers and/or potential predictors of response to therapy in pediatric IBD.
Immune regulation in the gastrointestinal tract
A large part of our published work focuses on the mechanisms that underlie mucosal regulatory T cell (Treg) differentiation. We have shown that the microenvironment, in particular the nature of the antigen presenting cell and cofactors such as cyclooxygenase derived prostaglandins and Fc receptor mediated signaling in the intestinal draining lymphoid tissue are crucial for inducible Foxp3+ Treg differentiation. Continuous surveillance by suppressive Treg cells prevents inflammatory responses to luminal antigens thereby maintaining intestinal homeostasis. However, the nature of the antigenic pressure varies at different locations of the intestinal tract. In agreement with this strong microenvironmental control our research shows that small intestinal and colonic Treg induction and function varies considerably. This led to the discovery that, in mice disruption of IL-10-driven- signaling in antigen presenting cells leads to spontaneous inflammation in the small intestine but not the colon. However, when colonized with Helicobacter hepaticus deficiency of IL-10 signaling in antigen presenting cells causes severe colitis. Using these mouse systems and cells from an IL-10R deficient patient we have performed in depth analysis of how the IL-10 pathway controls inflammatory Th1 and Th17 cells in small intestine and colon.
Celiac disease (CeD) is the most common food intolerance in the Western hemisphere: approximately 1:200 of the population is intolerant to gluten proteins found in wheat, rye and barley. Due to severe inflammation small intestinal villi, that are required for nutrient uptake, regress leading to malabsorption. Histologically a strong infiltration of lymphocytes (denoted as intraepithelial lymphocytes; IEL) is seen in the small intestinal epithelium. Serology shows these patients have circulating IgA antibodies to gluten and autoantibodies against tissue transglutaminase (TG2). The precise role of this humoral response in disease development is unclear. However, the anti-TG2 response is not coincidental as gluten proteins can be deamidated by TG2 in the gastrointestinal tract. Due to deamidation, negative charges are introduced into gluten peptides that facilitate their binding to the disease-predisposing HLA-DQ2 or HLA-DQ8 molecules which can trigger inflammatory interferon-g (IFN-g) and interleukin-21-driven CD4+ T-cell responses in the small intestine. The only available treatment for CeD is a lifelong gluten free diet. Importantly, when insufficiently treated, the disease may lead to a series of complications, ranging from deficiencies associated with small intestinal malabsorption to malignancy in later life.
In our patient studies we first focused on identifying the features that characterize pediatric CeD when compared to adult CeD. These results demonstrated that an inteleukin-17A-independent increase in interleukin-21 production by CD4+ Thelper cells is characteristic of pediatric CdD when compared to disease in adults. Moreover, in children, the percentages of peripheral blood CD4+Foxp3+ Treg cells were comparable between CeD patients and healthy age-matched controls. In adult patients on gluten-free diet and in refractory CD increased percentages of circulating natural CD62L+Foxp3+ Treg, but normal mucosally-imprinted CD62LnegCD38+Foxp3+ Treg frequencies were observed.
To unravel why CD patients fail to mount tolerance to gluten we have first established a humanized mouse model of gluten tolerance and discovered that tolerance to gluten is mediated by IL-10 secreting Foxp3 negative T cells. Next, we have dissected the mechanisms that underlie the differentiation of these tolerogenic IL-10 secreting cells and are currently investigating how disruption of IL-10 signaling leads to gluten-induced enteropathy in mice.
Inflammatory bowel disease
Inflammatory bowel disease (IBD) is a chronic disease of the intestinal tract caused by aberrant inflammatory immune responses to harmless microbiota. Two clinical subtypes of disease Crohn’s disease (CD) and Ulcerative colitis are distinguished. Both innate and adaptive immune responses contribute to the disease pathogenesis. However, excessive responses by inflammatory T-cells secreting interferon-γ (IFN-γ) or interleukin-17 (IL-17) drive the chronicity of IBD. Currently, patients are invariably treated with immune suppressants. Unfortunately, in approximately 30% of patients this therapy is insufficient entailing a risk of operation to remove the chronically inflamed intestinal segments.
In our translational work we have focused on dissecting inflammatory pathways that classify IBD patients in subgroups and yield parameters to characterize their disease subtype and therapy responsiveness. Since 2009 we have started our longitudinal IBD studies for which we are collecting patient peripheral blood and biopsies at diagnosis and fixed intervals during treatment. Using our newly acquired fundamental mucosal immunology knowledge phenotypic markers are designed to classify disease subtypes.
For example in our mouse systems we have identified that the mucosal factors TGF- β and retinoic acid induced a very selective expression pattern on differentiating CD4 + T cells in the mesenteric lymph node (4). This TGF- β – and retinoic acid- induced surface expression of CD62LnegCD38+ was also found to occur on human cells. In consequence we investigated whether the CD62LnegCD38+ phenotype could be used as a marker to identify circulating mucosal T cells in peripheral blood. In a collaborative study we established that indeed the CD62LnegCD38 + phenotype very selectively identifies gluten-specific circulating T cells in peripheral blood of bread challenged Celiac Disease patients. The value of defining this new subset lies in lowering the threshold for detection of luminal antigen specific T cells within the circulating CD4+ T-cell pool. This is of pivotal importance for functional studies as this method removes 90-95% of CD4+ non-gut imprinted effector T cells and naive T cells from the analysis. Current studies aim at characterizing circulating mucosal T cells from IBD patients and identifying their microbial specificity which is expected to be pivotal for identifying disease subtypes.
More recently, our translational work has expanded to studying IBD that arises in infants with genetic defects amongst which the IL-10R deficiency. Thorough analysis of these infants reveals pathways that are pivotal for intestinal tolerance in human and uncover patterns of inflammation that evolve from a particular defect. In turn, these patterns can be used to identify whether similar disease subtypes can be found in adolescent IBD and stem from a similar immune dysfunction.
Disease Overarching Research
Recently, we have initiated a large national consortium named TIMID: T cell driven Immune Mediated Inflammatory Diseases in which we are investigating which immune patterns are shared by patients with an immune mediated disease independent of their diseased organ. In a unique interdisciplinary setting with renowned scientists from 6 Dutch universities and 5 companies we are investigating whether a derailed host-gut microbiota immune interaction can be used to classify IMID patients (see https://timid.eu)