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Research Activities of the
Laboratory for Experimental Allogeneic BMT

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Ernst Holler, Günther Eissner, Joachim Hahn, Gerhard Hildebrandt, Matthias Grube, Ute Schulz, Altug Kesikli, Heike Bremm, Silvia Haffner, Verena Burger, Stephanie Moritz, Constanze Winter, Nancy Hahn, Sabine Schemm

For about 30 years allogeneic stem cell transplantation (SCT), among other curative approaches, is a well established method for the treatment of hematological malignancies. However, due to the disparity in major and minor histocompatibility antigens between donor and recipient, transplant-related complications (TRC), such as the graft-versus-host disease (GvHD) may arise. It is likely that the pathophysiology of GvHD consists of three phases. The first phase is triggered by conditioning-mediated inflammation and recipient cell apoptosis (Host tissue damage) with a contribution of proinflammatory cytokines. In the second acute phase one can find a predominance of Th1 cytokines, such as TNF-a and IFN-g accompanied by massive cellular infiltrates in the inflamed tissues, along with a massive donor T cell activation. Thirdly, during progression to the third chronic or effector phase there is a cytokine switch to the Th2 cytokines IL-4 and IL-10. In addition, tissue destruction takes place with a predominance of the gastrointenstinal tract and the lung. In all three phases the endothelium plays a pivotal role as an antigen presenting, a cytokine producing and an apoptosis target organ.

  
In our lab we are addressing the following questions using experimental models as well as translational research on clinical samples (clinical studies are not displayed here):

  • Pathophysiology of endothelial complications with a focus on cytokines and adhesion molecules
  • Monitoring of endothelial complications with functional assays of pts.´sera and with the detection of circulating endothelial cells (CEC)
  • Influence of alternative conditioning regimens on the vitality, activation and allogenicity of endothelial and epithelial cells, investigation of protective reagents, such as Defibrotide and Oligotide
  • Animal models for allogeneic stem cell transplantation (SCT), focus on pulmonary and endothelial complications
  • Monitoring of organ-specific alloreactions in the lung (clinical samples); pathophysiology and pathogenesis of pulmonary complications in the late phase post SCT
  • In vitro monitoring for GvHD and GvL by the help of in vitro predictive tests (skin explant assay, cytokine secretions assay, cytokine gene polymorphisms, minor Hag typing)
  • Studies on tolerance induction by regulatory T cells and anergizing dendritic cells
  • Bidirectional cross-talk between monocytes and endothelial cells under normal and proinflammatory conditions, focus on signal transduction mechanisms of tumor necrosis factor alpha (TNF)
  • Assessment of donor-recipient chimerism by the help of an PCR approach for the detection of variable numbers of tandem repeats (VNTR)
  • Minor histocompatibility typing of HLA-A2 positive donors and recipients
  • Analysis of immune recontitution post SCT by the help of T cell receptor excsion circles (TREC) determination
  • Differentiation of endothelial-like cells (ELC) from adult peripheral monocytic precursors as a source of patient-specific endothelial cells for transplant monitoring
  • PCR analyses of single-nucleotide polymorphisms in the NOD2/CARD15 gene and correlation with clinical outcome

Some of our key findings are:

  • Prognostic significance of TNF and interleukin 10 (IL-10) in the course of BMT
  • Balance between Th1 and Th2 cytokines in GvH and GvL
  • The endothelium is a target organ of BMT conditioning in vivo and in vitro with a crucial involvement of transmembrane TNF (mTNF)
  • mTNF is able to elicit reverse signals in monocytic cells that lead to LPS resistance, this type of anergy is caused by an exhaustion of the signal pathway of mitogen-activated protein kinases (MAPK). Reverse signalling of mTNF also occurs in T cells where it upregulates CD8+ and downregulates CD4+ activity.
  • Histopathological damage and keratinocyte apoptosis in the skin explant assay correlate with clinical outcome and the development of clincal GvHD in certain diagnostic subgroups
  • Correlation of keratinocyte apoptosis in the skin explant assay and clinical GvHD
  • Fludarabine as a compound of novel conditioning regimens induces activation and damage as well as allogenicity in human endothelial and epithelial cells, protective effect of Defibrotide and its low molecular weight derivative Oligotide
  • The presence of endothelial apoptosis inducing factor(s) in pts.´ sera correlates with episodes of clinical endothelial complications, including microangiopathy, and with acute GvHD
  • Single-nucleotide polymorphisms (SNP) in the NOD2/CARD15 gene in both donor and host associate with intestinal GvHD and transplant-related mortality. This observation has now been confirmed in 2 independent patient cohorts in a European-wide study.
  • Defibrotide, despite its endothelial protective function, also has anti-angiogenic functions in vitro and in vivo which is highly likely due to migration inhibition of endothelial cells, rather than due to proliferation inhibition.
Patient-specific ELC for transplant monitoring
Patient-specific ELC for transplant monitoring

  
Selected references (2001 to date):

  • Jarvis M., Schulz U., Dickinson A.M., Sviland L., Jackson G., Konur A., Wang X.N., Hromadnikova I., Kolb H.J., Eissner G., and Holler E. The Detection of Apoptosis in a Human in Vitro Skin Explant Assay for Graft Versus Host Reactions. J Clin Pathol 55:127, 2001
  • Eissner G, Multhoff G, Gerbitz A, Kirchner S, Haffner S, Sondermann D, Bauer S, Andreesen R, Holler E: Fludarabine induces apoptosis, activation and allogenicity in human endothelial and epithelial cells, protective role of defibrotide. Blood 100:334, 2002
  • Holler E. Cytokines, Viruses, and Graft-Versus-Host Disease. Curr Opin Hematol 9:479, 2002
    Gerbitz A, Schultz M, Wilke A, Linde HJ, Scholmerich J, Andreesen R, Holler E. Probiotic effects on experimental graft-versus-host disease: let them eat yogurt. Blood 103:4365, 2004
  • Holler E, Rogler G, Herfarth H , Brenmoehl J, Wild PJ, Hahn J, Eissner G, Schölmerich J, Andreesen R: Both, donor and recipient CARD15/NOD2 mutations associate with transplant-related mortality and GvHD following allogeneic stem cell transplantation. Blood 104:889, 2004
  • Gerbitz A, Nickoloff BJ, Olkiewitz K, Willmarth NE, Hildebrandt G, Liu C, Kobzik L, Eissner G, Holler E, Ferrara JLM, Cooke KR: A role for TNFa mediated endothelial apoptosis in the development of experimental idiopathic pneumonia syndrome. Transplantation 78:494, 2004
  • Eissner G, Kolch W, Scheurich P: Ligands working as receptors: Reverse signaling by members of the TNF superfamily enhances the plasticity of the immune system. Cytokine Growth Factor Rev 15:353, 2004
  • Rogler G, Holler E. Can NOD2/CARD15 mutations predict intestinal graft-versus-host disease and aid our understanding of Crohn's disease? Nat Clin Pract Gastroenterol Hepatol. 1:62-3, 2004.
  • Eissner G, Iacobelli M, Blüml S, Burger V, Haffner S, Andreesen R, Holler E: Oligotide, a defibrotide derivative, protects human microvascular endothelial cells against fludarabine-induced activation, damage and allogenicity. Bone Marrow Transplant 35:915-920, 2005
  • Vudattu NK, Holler E, Ewing P, Schulz U, Haffner S, Burger V, Kirchner S, Andreesen R, Eissner G: Reverse signaling of membrane integrated tumor necrosis factor (mTNF) differentially regulates alloresponses of CD4+ and CD8+ T cells against human microvascular endothelial cells. Immunology 115:536-543, 2005
  • Holler E, Rogler G, Brenmoehl J, Hahn J, Herfarth H, Greinix H, Dickinson AM, Socie G, Wolff D, Fischer G, Jackson G, Rocha V, Steiner B, Eißner G, Marienhagen J, Schoelmerich J, Andreesen R. Prognostic significance of NOD2/CARD15 variants in HLA-identical sibling hematopoietic stem cell transplantation: Effect on long term outcome is confirmed in 2 independent cohorts and may be modulated by the type of gastrointestinal decontamination. Blood 107:4189-4193, 2006

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Letzte Aktualisierung: 21.10.2008 | Online-Redaktion
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