In the first funding period, we observed that neutrophils mediate cellular communication between the ileum and the mesenteric lymph nodes during early graft-versus-host disease (GVHD). Phenotypically different subsets of neutrophils could be distinguished. Single cell isolation and RNA-sequencing revealed an immuno-regulatory phenotype of neutrophils located in the intestinal tract and mesenteric lymph nodes. We hypothesize thus that one of the factors leading to the immunopathology of GVHD is an impaired tolerogenic effect of a subset of neutrophils on the alloreactive T cell response. Why the immune reaction by tolerogenic neutrophils is impaired is to date unclear. Current GVHD treatments fail to enhance the function of tolerogenic neutrophils, but rather cause global unspecific impairment of immune reactions. We plan now to characterize the distinct signalling / transcriptional events in neutrophil subsets. To test for a potential translation into the clinic, we will assess the phenotype and function of human neutrophils in patients with acute steroid refractory GVHD. Overall, we plan to characterize the immuno-regulatory function of neutrophil subsets in GVHD.