MyD88/TLR9 mediated immunopathology and gut microbiota dynamics in a novel murine model of intestinal graft-versus-host disease.
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.


OBJECTIVE
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.
Heimesaat
Markus M
MM
Institut für Mikrobiologie und Hygiene, Charité-Universitätsmedizin, Campus Benjamin Franklin, Berlin, Germany. markus.heimesaat@charite.de
Nogai
Axel
A
Bereswill
Stefan
S
Plickert
Rita
R
Fischer
André
A
Loddenkemper
Christoph
C
Steinhoff
Ulrich
U
Tchaptchet
Sandrine
S
Thiel
Eckhard
E
Freudenberg
Marina A
MA
Göbel
Ulf B
UB
Uharek
Lutz
L
D000208:Acute Disease; D000818:Animals; D017209:Apoptosis / Q000276:immunology; D016026:Bone Marrow Transplantation; D049109:Cell Proliferation; D003092:Colitis / Q000276:immunology / Q000382:microbiology* / Q000473:pathology / Q000517:prevention & control; D004195:Disease Models, Animal; D005260:Female; D006086:Graft vs Host Disease / Q000276:immunology* / Q000382:microbiology* / Q000473:pathology / Q000517:prevention & control; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D053594:Myeloid Differentiation Factor 88 / Q000172:deficiency / Q000276:immunology*; D016376:Oligonucleotides, Antisense / Q000627:therapeutic use; D013154:Spleen / Q000637:transplantation; D051217:Toll-Like Receptor 9 / Q000172:deficiency / Q000276:immunology*; D019172:Transplantation Conditioning
C507780:Myd88 protein, mouse; D053594:Myeloid Differentiation Factor 88; D016376:Oligonucleotides, Antisense; C495370:Tlr9 protein, mouse; D051217:Toll-Like Receptor 9