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MyD88/TLR9 mediated immunopathology and gut microbiota dynamics in a novel murine model of intestinal graft-versus-host disease
  1. Markus M Heimesaat1,
  2. Axel Nogai2,
  3. Stefan Bereswill1,
  4. Rita Plickert1,
  5. André Fischer1,
  6. Christoph Loddenkemper3,
  7. Ulrich Steinhoff4,
  8. Sandrine Tchaptchet5,
  9. Eckhard Thiel2,
  10. Marina A Freudenberg5,
  11. Ulf B Göbel1,
  12. Lutz Uharek2
  1. 1Institut für Mikrobiologie und Hygiene, Charité-Universitätsmedizin, Campus Benjamin Franklin, Berlin, Germany
  2. 2Medizinische Klinik III (Hämatologie, Onkologie und Transfusionsmedizin), Charité - Universitätsmedizin Berlin, Berlin, Germany
  3. 3Institut für Pathologie, Charité - Universitätsmedizin Berlin, Berlin, Germany
  4. 4Max Planck Institut für Infektionsbiologie, Berlin, Germany
  5. 5Max Planck Institut für Immunbiologie, Freiburg, Germany
  1. Correspondence to Dr Markus M Heimesaat, Institut für Mikrobiologie und Hygiene, Charité - Universitätsmedizin Berlin, Charité Centrum 5, Campus Benjamin Franklin, Hindenburgdamm 27, D-12203 Berlin, Germany; markus.heimesaat{at}charite.de

Abstract

Background The bacterial microflora aggravates graft-versus-host-disease (GvHD) after allogeneic stem cell transplantation, but the underlying mechanisms of manifestations of intestinal GvHD (iGvHD) in the gut remain poorly understood.

Aim To analyse the gut flora composition and the impact of bacterial sensing via Toll-like receptors (TLRs) in iGvHD.

Methods By mimicking clinical low-intensity conditioning regimens used in humans, a novel irradiation independent, treosulfan and cyclophosphamide-based murine allogeneic transplantation model was established. A global survey of the intestinal microflora by cultural and molecular methods was performed, the intestinal immunopathology in TLR-deficient recipient mice with iGvHD investigated and finally, the impact of anti-TLR9 treatment on iGvHD development assessed.

Results The inflammatory responses in iGvHD were accompanied by gut flora shifts towards enterobacteria, enterococci and Bacteroides/Prevotella spp. Analysis of iGvHD in MyD88-/-, TRIF-/-, TLR2/4-/-, and TLR9-/- recipient mice showed that bacterial sensing via TLRs was essential for iGvHD development. Acute iGvHD was characterised by increasing numbers of apoptotic cells, proliferating cells, T cells and neutrophils within the colon. These responses were significantly reduced in MyD88-/-, TLR2/4-/-, TRIF-/- and TLR9-/- mice, as compared with wild-type controls. However, TRIF-/- and TLR2/4-/- mice were not protected from mortality, whereas TLR9-/- mice displayed increased survival rates. The important role of TLR9-mediated immunopathology was independently confirmed by significantly reduced macroscopic disease symptoms and colonic apoptosis as well as by reduced T-cell and neutrophil numbers within the colon after treatment with a synthetic inhibitory oligonucleotide.

Conclusions These results emphasise the critical role of gut microbiota, innate immunity and TLR9 in iGvHD and highlight anti-TLR9 strategies as novel therapeutic options.

  • Gut flora
  • innate immunity
  • TLR9
  • synthetic inhibitory oligonucleotide
  • intestinal GvHD
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Footnotes

  • MMH, AN, SB contributed equally to the work.

  • Funding This work was funded by grants of the Fritz-Thyssen-Stiftung (Grant No 10.05.2.194) and German Research Foundation (DFG) to UBG and AF (SFB633/ TP A7), MMH (SFB633/ TP B6) and LU (SFB633/ TP B11).

  • Competing interests None.

  • Provenance and peer review Not commissioned; externally peer reviewed.

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