High-Resolution Phenotyping of Early Acute Rejection Reveals a Conserved Alloimmune Signature

Alloimmune responses in acute rejection are complex, involving multiple interacting cell types and pathways. Deep profiling of these cell types has been limited by technology that lack the capacity to resolve this high dimensionality. Single-cell mass cytometry was used to characterize the alloimmune response in early acute rejection, measuring 37 parameters simultaneously, across multiple time-points in two models; a well-established murine cardiac and a novel murine vascularized composite allotransplant (VCA). Unsupervised hierarchical clustering was used to group related cell types defined by combinatorial expression of surface and intracellular proteins, along with markers of effector function and activation. This expression profile was mapped to visualize structural changes in antigen composition across cell types, revealing unique phenotypic signatures in alloimmune T cells, NK cells, and myeloid subsets that were conserved and that firmly distinguished rejecting from non-rejecting grafts.