A Shift in the Intestinal Microflora towards Pro-Inflammatory Bacteria and Signaling Via Toll-Like-Receptor 2 Triggers Early Onset of Graft-Versus-Host Disease.

Pattern recognition receptors such as Toll-like-receptors (TLR) and NOD proteins bind specifically to microbial components and activate innate immune responses. Reducing the bacterial load by antibiotic treatment has been shown to ameliorate the severity of graft-versus-host-disease (GvHD) after allogenic stem cell transplantation. The impact of bacteria and TLRs on the induction of T-cell-alloreactivity and GvHD was investigated in a murine transplantation model. C57B/10 wild-type (wt) mice served as recipients of MHC-mismatched Balb/c grafts after conditioning with treosulfan and cyclophosphamide. Analyses of the gut microbiota in fecal samples showed a pronounced increase of luminal load of both gram-positive and gram-negative bacteria within 11 days after stem cell transplantation. The increase in the Escherichia coli load was significantly correlated with the severity of graft-versus-host-disease and with decreased survival rates. Analysis of the kinetics of fecal E. coli load and the histologial and clinical GvHD scores revealed that the increase of the fecal E. coli load is detected simultanously to the onset of clinical overt GvHD. Because TLR 2 and 4 represent the major receptors for gram-positive and gram-negative bacteria, respectively, the incidence and severity of GvHD was investigated in TLR2, TLR4 and TLR2/4 deficient mice on a C57B/10 background. The onset of GvHD was delayed in recipient mice lacking TLR2 and both TLR2 and 4 (p<0.05) whereas a single TLR4 deficiency had no influence. Interestingly, there was no correlation of E. coli load and GvHD in TLR2-deficient mice.Taken together, these results support potential roles of TLR2-dependent signaling and gut bacterial components in the induction and early onset of intestinal GvHD. In expansion of these findings, our data elucidate the role of the innate immune system and its receptors for the induction of alloreactive T-cell responses, providing evidence that modulation of the intestinal gut flora might prevent or reduce GvHD in humans.