Abstract
Allograft rejection following solid-organ transplantation is a major cause of graft dysfunction and mortality. Current approaches to diagnosis rely on histology, which exhibits wide diagnostic variability and lacks access to molecular phenotypes that may stratify therapeutic response. Here, we leverage image-based spatial transcriptomics at sub-cellular resolution in longitudinal human cardiac biopsies to characterize transcriptional heterogeneity in 62 adult and pediatric heart transplant (HT) recipients during and following histologically-diagnosed rejection. Across 28 cell types, we identified significant differences in abundance in CD4 + and CD8 + T cells, fibroblasts, and endothelial cells across different biological classes of rejection (cellular, mixed, antibody-mediated). We observed a broad overlap in cellular transcriptional states across histologic rejection severity and biological class and significant heterogeneity within rejection severity grades that would qualify for immunomodulatory treatment. Individuals who had resolved rejection after therapy had a distinct transcriptomic profile relative to those with persistent rejection, including 216 genes across 6 cell types along pathways of inflammation, IL6-JAK-STAT3 signaling, IFNα/IFNγ response, and TNFα signaling. Spatial transcriptomics also identified genes linked to long-term prognostic outcomes post-HT. These results underscore importance of subtyping immunologic states during rejection to stratify immune-cardiac interactions following HT that are therapeutically relevant to short- and long-term rejection-related outcomes.