Our projects are externally funded by the Deutsche Forschungsgemeinschaft (DFG) through an individual project grant and the Cluster of Excellence Inflammation-at-Interfaces (ExC 306) as well as the Else-Kröner-Fresenius Stiftung.
PD Dr. Hans-Heinrich Oberg (DFG, ExC), Dr. Christian Peters (EKFS), Dr. Guranda Chitadze (ExC), M.Sc. Jaydeep Bhat (ExC), M.Sc. Léonce Kouakanou (DAAD), M.Sc. Daniel Gonnermann (DFG), cand. med. Theo Nerdal, cand. med. Valentina Biel, Hoa Ly (MTLA), Monika Kunz (MTLA)
1. Reciprocal interactions between neutrophils and gamma/delta T lymphocytes (DFG Ka 502/16-1)
Human Vgamma9Vdelta2 (Vg9Vd2) T cells recognize pyrophosphate molecules secreted by many bacteria. Homologous molecules produced by eukaryotic cells are also recognized by gd T cells but require 3-log higher concentrations. Tumor cells overproduce endogenous pyrophosphates that can be sensed by Vg9Vd2 T cells as tumor-associated antigens, which formally explains the role of these gd T cells in both anti-infective and anti-tumor immunity. Importantly, the cellular production of gd T cell-stimulating pyrophosphates can be drastically increased by aminobisphosphonates which interfere with the mevalonate pathway involved in pyrophosphate production. We have synthesized a fluorescent derivative („FluorZOL“) of zoledronate, the most potent gd T cell-stimulating aminobisphosphonate. We found that among blood leukocytes, neutrophils and monocytes but not lymphocytes efficiently take up FluorZOL. However, despite efficient uptake of zoledronate, neutrophils are unable to support gd T cell expansion in vitro. In addition, we observed that following uptake of zoledronate, neutrophils actually inhibit gd T cell expansion, due to the production of inhibitory molecules including reactive oxygen species. Within this project, we aim to study in detail the reciprocal regulatory interactions between neutrophils and gd T cells. A precise understanding might help to design strategies to prevent the functional anergy and/or disappearance of gd T cells following i.v. application of zoledronate and related aminobisphosphonates. Moreover, we also study the capacity of various leukocyte subpopulations to present microbial and eukaryotic pyrophosphates („phosphoantigens“) to Vg9Vd2 T cells, with a special focus on the role of the butyrophilin 3A1 (CD277) antigen.
Contributors: PD Dr. Hans-Heinrich Oberg (DFG), Dr. Christian Peters (EKFS), cand. med. Theo Nerdal, cand. med. Valentina Biel, Monika Kunz, Hoa Ly
External collaborations: Dr. Shirin Kalyan (Vancouver, BC), Prof. Thisbe Lindhorst (Institute of Organic Chemistry, University of Kiel), Prof. Daniel Olive (Marseille, France), Prof. Jorma Määttä (Kuopio, Finland)
2. Regulatory function of human gamma/delta T cells (Else-Kröner-Fresenius Stiftung 2013 A276)
Human gd T cells display a broad range of effector functions including cytokine production, cytotoxicity, antigen presentation and regulatory activity. This project focusses on the characterization of the regulatory activity and cytokine production of human gd T cells. The major goal is the identification of molecular pathways involved in the induction of suppressive activity and differential cytokine induction. To this end, purified gd T cells are cultured in vitro under differential priming conditions in the absence or presence of various cytokines and T cell receptor stimuli. The phenotype of the cultured gd T cells will be analyzed at the transcriptional level and micro-RNA and protein expression, as well as the regulatory activity. This project will help to define the pathways for the generation of suppressive gd T cells and to understand in more detail the functional plasticity of gd T cells. From a translational point of view, this is important in order to optimize the potential application of gd T cells in tumor immunotherapy by preventing suppressive activity of gd T cells. With regard to autoimmune disorders, however, it might be appropriate to intentionally induce suppressive activity in gd T cells.
Contributors: Dr. Christian Peters (EKFS), M.Sc. Léonce Kouakanou (DAAD), Ina Martens (Medical Faculty)
This project is a close collaboration with the AG Wesch
3. Functional plasticity and epigenetic profiling of human gamma/delta T cells (ExC 306 project N TP1)
Based on T cell receptor V gene usage, human gd T cells basically come in two flavours: While Vg9Vd2 T cells dominate among peripheral blood gd T cells, Vd1 cells account for the majority among intraepithelial gd T cells, notably in the small intestine. Moreover, gd T cells can display a surprisingly large functional plasticity. One of the basic principles of gene regulation is epigenetic modulation at the level of histone acetylation, DNA methylation, and other mechanisms. The aim of this project is to gain information on the functional plasticity and epigenetic profiling of human gd T cells in comparison to other immune cell populations, notably NKT cells. To this end, we perform (i) genome-wide methylation profiling of total alpha/beta versus total gamma/delta T cells to identify the role of specific genes in ab versus gd T cell specification; (ii) integrative epigenomic analysis using ATAC sequencing; and (iii) clonal analysis of plasticity of naive Vg9Vd2 T cells in response to various priming conditions. Moreover, we will perform RNA analysis including miRNA and lnRNA to obtain a full appreciation of differences between ab and gd T cells. Furthermore, the analysis of Vd1 and Vd2 T cells isolated from inflamed tissue of patients with inflammatory bowel diseases and the direct comparison with other innate-like cells such as iNKT cells will provide additional information on the functional plasticity of gd T cells in inflammation.
Contributor: M.Sc. Jaydeep Bhat (ExC 306), Hoa Ly
External collaborations: Prof. Ole Ammerpohl and Prof. Reiner Siebert (Institute of Human Genetics, University of Kiel), Prof. Sebastian Zeissig (1st Medical Department, UKSH Campus Kiel), Prof. Philipp Rosenstiel and Dr. Maren Falk-Paulsen (Institute of Clinical Molecular Biology, UKSH Campus Kiel)
4. Functional immunophenotyping of therapeutic antibody responses (ExC 306 project R TP2 )
Anti-TNFalpha and anti-integrin antibodies have dramatic clinical effects in many but not all patients with inflammatory bowel disease (IBD) and Psoriasis. Moreover, some IBD and Psoriasis patients are refractory to this treatment, show an insufficient response, or develop for unknown reasons a secondary resistance to anti-TNF antibodies. To better understand the effects of such biologicals on the human immune system, we will carry out an in-depth functional immunophenotyping in IBD and Psoriasis patients before and at defined intervals after initiation of antibody therapy. We will perform on mucosal and skin biopsies and blood samples (i) multicolor FACS analysis for cell surface markers and intracellular antigens, (ii) multiplex cytokine analysis, (iii) quantitative single cell analysis of lymphocyte/monocyte activation by ImageStream® cytometry, and (iv) T-cell receptor spectratyping. Furthermore, the in vitro activation of blood T cells and the activity of regulatory T cells will be studied. The aim of our studies is (i) to understand the impact of anti-cytokine/anti-integrin antibody therapies on the human immune system; (ii) to identify immune system parameters that characterize responders versus non-responders; and (iii) eventually to develop a diagnostic immune profile that will allow (in conjunction with a transcriptomic profile) to predict the responsiveness to biologicals in individual IBD patients.
Contributor: Dr. Guranda Chitadze (ExC 306)
External collaborations: This subproject TP 2 is part of the ExC 306 joint project R coordinated by Prof. Dr. Stefan Schreiber (1st Medical Department, UKSH Campus Kiel). Co-investigators are Prof. Dr. Sebastian Zeissig (1st Medical Department, UKSH Campus Kiel) and Prof. Stephan Weidinger (Dept. of Dermatology, UKSH Campus Kiel)
5. Cluster Laboratory VII Human Immunophenotyping (ExC 306 CL VII)
The Cluster Laboratory VII Human Immunophenotyping (CLVII) is a platform structure of the Cluster of Excellence Inflammation-at-Interfaces (ExC 306) jointly organized by the Dept. of Dermatology UKSH Campus Lübeck (Prof. Dr. D. Zillikens) and the Institute of Immunology UKSH Campus Kiel (Prof. Dr. D. Kabelitz). The Kiel Campus provides high speed cell sorting (BD FACSAria), up to 11-colour flow cytometry (BD LSR Fortessa) and ImageStream cytometry (EMD Millipore ImageStream Mark II) (see the CLVII web site http://www.immunophenotyping.uni-luebeck.de/index.php?id=5). The ImageStream cytometer combines features of both laserscan microscopy (single cell imaging) and flow cytometry (quantitative analysis) and offers unique possibilities for the analysis of processes such as receptor internalization, colocalization, protein-protein interaction, synapse formation, and many others (see https://www.amnis.com/imagestream.html).
Contributors: PD Dr. Hans-Heinrich Oberg (ExC), Sandra Ussat (Medical Faculty, ExC)