CD34+ cell-derived endothelial cells orchestrate vascular and immune remodeling in the transplanted liver.

BACKGROUND & AIMS: Endothelial cell (EC) damage is an initiating event in acute cellular rejection after liver transplantation (LT). However, the origin and characteristics of post-transplant neonatal ECs remain controversial. We aimed to uncover the mechanisms underlying EC-T cell interactions after transplantation and to develop an EC-targeted strategy to alleviate transplant rejection.

METHODS

Leveraging single-cell RNA sequencing from 13 human and 4 murine liver allografts, we mapped the functional atlas of ECs. Allogeneic orthotopic LT in CAG;R26-tdTomato and Cd34-CreERT2;R26-tdTomato mice confirmed the cellular origin of ECs. We used CellChat, multiplex immunohistochemistry, and in vitro co-culture models to investigate the mechanism of EC-T cell interactions. Using a platelet-based bio-delivery system, we achieved targeted delivery of a CXCL12 monoclonal antibody (αCXCL12).

RESULTS

ECs in transplanted livers exhibited a dual origin, being derived from both donor and recipient cells. Recipient-derived ECs were characterized by high CD34 expression, high stemness, and pro-inflammatory characteristics. Using genetic lineage-tracing mice combined with an orthotopic LT model, we found that recipient CD34+ cell-derived ECs peaked at 2 weeks post-LT and declined by 4 weeks, consistent with the temporal pattern of rejection. We further identified that CD34+ ECs recruit and potentiate Th1 and cytotoxic CD8+ T cells via the CXCL12-CXCR4 axis and co-stimulatory molecules. In turn, cytotoxic CD8+ T cells induced pyroptosis of CD34+ cell-derived ECs through the Caspase1-GSDMD pathway. Platelets loaded with αCXCL12 specifically targeted ECs in the transplanted liver, reducing T-cell infiltration and mitigating rejection.

CONCLUSIONS

We identified a population of recipient-derived CD34+ ECs that exacerbates acute rejection by activating T cells through the CXCL12-CXCR4 axis. We further developed a platelet-based delivery strategy that precisely targets EC-derived CXCL12 and effectively prevents acute cellular rejection.

IMPACT AND IMPLICATIONS: We integrated single-cell transcriptomic data from human and murine liver allografts to delineate the dual cellular origins and functional atlas of endothelial cells. Using lineage-tracing mice in an allogeneic orthotopic liver transplantation model, we tracked the fate of CD34+ cells and demonstrated the contribution of recipient-derived CD34-lineage endothelial cells to liver allograft angiogenesis. These CD34-lineage endothelial cells recruited T cells through the CXCL12-CXCR4 axis and activated them via co-stimulatory molecules. Furthermore, a platelet-based biological delivery strategy targeting CXCL12 in CD34-lineage endothelial cells alleviated T cell-mediated rejection. This study provides an endothelial cell-centered perspective and proposes a potential novel immunosuppressive strategy for liver transplantation.