🎗Liver-resident natural killer cells promote immune tolerance after liver transplantation via the PDL1/PD1 axis: Toward engineered cell therapy

Introduction: Immune tolerance has been the "holy grail" in transplantation immunity, making its safe induction a critical clinical goal after liver transplantation (LT). Liver-resident natural killer (lrNK) cells exhibit inherent regulatory properties, but their exact role in immune tolerance remains unclear. In this study, we aimed to elucidate the contribution of lrNK cells to tolerance induction and develop an lrNK-based cell therapy to promote long-term graft acceptance.
Methods: Mouse orthotopic LT models were established to characterize spontaneous tolerance dynamics. Single-cell RNA sequencing (scRNA-seq) was performed on human and mouse grafts to profile immune cell landscape changes after LT. LrNK cell function was validated via mouse models, combined with in vitro experiments. We induced "lrNK-like cells" using cytokine stimulation, and assessed their safety and therapeutic efficacy in mouse models.
Results: In the allogeneic mouse orthotopic LT model, transplanted livers spontaneously established immune tolerance after a brief period of rejection (Figure 1A-E). Both human and mouse samples revealed significant lrNK cell expansion in the tolerance liver (Figure 1F-G and 1I-K). Transcriptomic and functional enrichment analyses confirmed lrNK cells were enriched in immune-suppressive genes and tolerance-associated pathways (Figure 1H and 1L-M). These results suggested that lrNK may be correlated with tolerance establishment. Depletion of donor-derived lrNK cells exacerbated early acute rejection but did not prevent long-term tolerance development (Figure 1N-R), which might due to the compensatory infiltration of recipient-derived lrNK cells (Figure 1S). In contrast, depletion of recipient-derived lrNK cells resulted in fatal, severe acute rejection (Figure 1T-W), confirming recipient lrNK cells as essential mediators of spontaneous tolerance.

CellChat ligand-receptor analysis and mechanistic studies further confirmed that lrNK cells suppressed T cell proliferation via the PD-L1/PD-1 axis, and blocking PD-L1 could reverse this inhibitory effect (Figure 2A-C). Trajectory analysis of scRNA-seq data showed that lrNK cells could differentiate from cNK cells, which might be related to IL-2 and IL-15 (Figure 2D-F). Using IL-2 and IL-15 stimulation, we successfully induced cNK cells to differentiate into induction NK (iNK) cells with “lrNK-like” phenotypic and functional features, including high PDL1 expression (Figure 2G-J). Additionally, we found that adoptive transfer of iNK cells into LT recipients reduced graft rejection (Figure 2K-O).

Conclusion: Recipient-derived lrNK cells play a central role in mediating immune tolerance post-LT by suppressing T cell responses via the PDL1/PD1 axis. In vitro-engineered iNK cells represent a safe and effective cell-based therapy to reduce acute rejection and promote immune tolerance, offering a promising translational strategy to improve long-term outcomes for LT recipients.