Bestimmung der relativen Zellverteilung peripher zirkulierender T-Zell-Subpopulationen unter besonderer Berücksichtigung des Chemokin- und Homing-Rezeptorprofils bei einem pädiatrischen CED-Patientenkollektiv unter Infliximab- Therapie

Zusammenfassung (Abstract) Hintergrund und Ziele (Background) The pathological characteristics of inflammatory bowel disease (IBD) are connected to mucosal leukocyte infiltration, which contributes to the progression and persistence of the disease via chemokine receptors expressed by pro- or anti-inflammatory cells. The recruitment of leukocytes to the intestinal mucosa entails the local intestinal expression of these chemokines, which interact with receptors located on the leukocytes' surface. In this longitudinal study, CD4+-T helper cells were examined throughout the course of an infliximab therapy in the peripheral blood in a cohort of pediatric IBD patients. The aim was to determine the individual chemokine and homing receptor expression profiles and the effects of an anti- TNF-α therapy throughout the course of this study. Methoden (Methods) A total of 42 pediatric patients who received a therapy with infliximab were recruited (Crohn's disease, n=30, ulcerative colitis, n=12). 14 healthy, age-matched children served as control subjects. Peripheral blood mononuclear cells (PBMCs) were collected before starting anti-TNF- α therapy and after three, six and twelve months on therapy. Multicolor flow cytometry was performed to assess the surface expression of a4b7, CXCR5, CCR6, CD62L, CCR5, CXCR3 and CCR9 after previous cellular immunomagnetic isolation using CD4-specific MACS beads. Responders to successful infliximab therapy were classified as those who achieved a fecal calprotectin level below 100 μg/g or levels <10% of baseline at the end of the 12-month follow- up period. The study was approved by the scientific ethics committee of the Friedrich- Alexander-University Erlangen-Nuremberg (#347_15B). Ergebnisse (Results) A key finding of this study is the differential expression and alteration in expression of three chemokine receptors, namely a4b7, CCR6 and CCR5 among those investigated. After 12 months of infliximab therapy, the expression of the integrin receptor a4b7 in responder-patients (RS) was downregulated to a greater extent than in non-responder patients (NRS). This was observed in the Tconv-Memory, Teff-Memory and Treg-Memory cell populations. Furthermore, we demonstrate that CCR6 expression on peripheral CD4+-T-cells in the Crohn’s disease (CD) population and ulcerative colitis (UC) population increased considerably during the early stages of therapy. This tendency was significant in the Crohn's disease (CD) population. This trend was also observed for CCR6+ effector T cells in responder patients who sustained a state of remission. In the responders, this upregulation of CCR6 expression was followed by a downregulation after 6 months of therapy while an increased CCR6 expression was still detected in the memory T-cell population of the non-responders. A decreased expression of CD62L on naïve CD4+ T cells was indicative for the presence of an ulcerative colitis (ROC analysis: sensitivity 93%, specificity 86%, cut off MFI<4377) when compared to healthy subjects. Our results further indicate for CCR5 that between the start and the end of an infliximab therapy, the expression of CCR5 on Teff-memory cells was comparably low in the responding population compared to the healthy control and the non-responder population. Prior to infliximab therapy, it could be observed that CCR5 was predominantly expressed on memory- T-cells, whilst CCR5 expression on naïve-T-cells was negligible in all inflammatory bowel disease patients, in both the responder- and non- responder patients as well as in and the healthy control patients. Schlussfolgerung (Conclusion) The homing and chemokine receptor profiles of peripheral CD4+-T-helper cells altered under an infliximab therapy in a cohort of pediatric patients over a 12-month period. In the therapy responding population, expression of important homing and chemokine receptors, such as a4b7 and CCR6, was predominantly downregulated. As this contrasts with the non-responding population, who predominantly experienced upregulation of these chemokine receptors as well as continuing disease activity, this observation may indicate unaltered inflammatory activity and thus therapy failure.