Alloantigen Infusion Selectively Activates the Transcriptome of Type 2 Conventional Dendritic Cells"

Recent studies have revealed novel molecular mechanisms by which innate monocytic cells acutely recognize and respond to alloantigen, and the significance of this to allograft rejection and immunologic tolerance. What remains unclear is the single cell heterogeneity of the innate allo-response, and particularly the contribution of dendritic cell (DC) subsets. To investigate the acute response of innate immune cells and DC subsets to exposure of alloantigen, experimental rodents were administered an intravenous injection of live allogenic cells versus isogenic cells. In parallel, we also infused apoptotic allogenic and isogenic cells, which have been reported to modulate immunity. Forty-eight hours after injection, recipient spleens were harvested, pooled by condition, enriched for DCs, and subjected to single cell mRNA sequencing and downstream analysis. Injection of live splenic cells induced a greater quantity of transcriptional changes across all DC subsets as identified by differential gene expression analysis and compared to apoptotic cell injection. In the setting of live cell infusion, type 2 conventional dendritic cells (cDC2s) were more transcriptionally responsive with a Ccr2+ cDC2 subcluster uniquely responding to the presence of alloantigen compared to isogenic control. Candidate receptors of allorecognition in other innate populations were interrogated amongst cDC2s and A-type paired immunoglobulin-like receptors (PIR-As) was found to be increased specifically in the cDC2 population following allo-exposure at the mRNA level and by flow cytometry. These results illuminate previously unclear distinctions between therapeutic infusions of live versus apoptotic allogenic cells and newly implicate cDC2 cells in innate allorecognition in the context of transplantation. Overall design: Splenic cells magnetically enriched for DCs and CD45+ cells from mice infused with live allo- (LA, Balbc), live iso- (LI, B6), apopototic allo- (AA, Balbc), apoptotic iso- (AI, B6) antigen or saline control (C) 48 hours prior and subjected to single cell RNA sequencing.