*Purpose: Single-cell RNA sequencing is an emerging, powerful genomic technology that can describe heterogenous cellular populations of whole solid organs with high resolution. We report on preliminary single-cell RNA sequencing analysis of liver biopsy tissue obtained at time of transplant from three enrolled living-donor liver transplant (LDLTx) patients infused with donor-derived regulatory dendritic (DCreg) cells as part of an ongoing in-human phase I/II clinical trial (NCT03164265).

*Methods: DCregs generated from prospective living donor elutriated monocytes were infused into three LDLTx recipients enrolled in our study 7 days prior to transplantation. An 18-gauge core needle biopsy of the donor hepatic allograft was obtained 1-hour after reperfusion in the recipient and sent for tissue digestion and processing. A subsequent single cell genomics library was prepared using the 10x Genomics platform. Data was normalized and clustered to identify distinct resident cell populations in the transplanted livers after DCreg infusion.

*Results: We generated 6,440 single cell transcriptomes of transplant hepatic tissue from three trial LDLTx recipients and performed unsupervised clustering analysis of gene expression. We uniquely identify 4 clusters of granulocytes/neutrophils and 4 clusters of NK cells in our transcriptional profile that has not been previously reported in single cell analysis on normal human liver. We also demonstrate the presence of a CD16+ monocyte population that represents a distinct non-classical subset of monocytes. Regulatory gene expression profiles across several innate and adaptive immune cell types were represented, including CTLA4 which may indicate the presence of a regulatory T cell population.

*Conclusions: A large number of immune cell subtypes are represented in the immediate post-transplant reperfused liver, distinct from that previously reported in normal human liver. Regulatory gene expression profiles are present and may indicate a heterogenous immune response to DCreg infusion, liver transplantation, or both. This preliminary study provides a proof of concept for the role of next-generation sequencing in defining changes to tissue immune cell landscape as response to stimuli.

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