Background/Aim: We have documented crucial role of TIM-3 dependent T cell dysfunction in mouse orthotopic liver transplantation (OLT). CEACAM1 (CD66a; CM1; Carcinoembryonic Antigen-Related Cell Adhesion Molecule 1) has been recently identified as a cellular TIM-3 ligand, essential for T cell priming/negative immune regulation in autoimmune and neoplastic disorders. The role of TIM-3 – CEACAM1 signaling in transplantation remains unknown. In this study, we analyzed the impact of donor liver CM1 on host T cell activation and hepatocyte stress response in OLT. Methods/Results: We used a mouse OLT model with extended cold storage (18h). Groups of wild-type (WT) and CM1-deficient (CM1-KO) livers transplanted to WT recipients (C57/BL6 to C57/BL6) were collected at 6h post-OLT (n=5/gr). Disruption of graft CM1 signaling exacerbated hepatocellular damage, evidenced by sALT levels (IU/L; 14,395±3,811 vs. 9,082±1,696; p<0.05), Suzuki's histological grading (p<0.01) and frequency of TUNEL+ cells (p<0.05). FACS-assisted PBL analysis in CM1-KO→WT hosts showed depressed TIM3+ (22.9% vs. 70.9%) and PD1+ (22.9% vs. 68.9%) but increased Tbet+ (27.1% vs 9.6%) T cell staining, as compared to WT→WT. CM1-KO liver grafts had consistently increased levels of IL2, IL1β, TNFα, CXCL1, CXCL2 (p<0.05). Then, we assessed the role of CM1 signaling in primary hepatocyte stress response to H₂O₂ in vitro. Indeed, CM1-deficient hepatocytes were more susceptible to H₂O₂-induced apoptosis, evidenced by increased cleaved caspase 8/3, frequency of TUNEL+ cells and elevated LDH activity in culture medium (p<0.05). Of note, CM1-KO hepatocytes exhibited lower mRNA levels coding for cytoprotective IGF1 gene (p<0.05). Finally, to assess putative clinical relevance of donor liver-dependent CM1 cytoprotection, we analyzed 21 post-transplant human liver biopsies (Western blots) divided into high (n=10) vs. low (n=11) CM1 expression groups. Indeed, serum transaminase levels at POD1 in the “low” CM1 group were markedly higher as compared with the “high” CM1 group (AST: 545±213 vs 277±73, ALT: 469±128 vs 267±65). Conclusion: Graft CM1 plays a key role in maintaining OLT function not only by priming TIM-3 dependent T cell inactivation (exhaustion?) but also by promoting hepatocyte resistance against stress-induced apoptosis.

CITATION INFORMATION: Nakamura K, Kageyama S, Kujawski M, Sosa R, Reed E, Zarrinpar A, Busuttil R, Kupiec-Weglinski J. TIM-3 – CEACAM1 Checkpoint Regulation of T-Cell Activation and Hepatocellular Function in Liver Transplantation. Am J Transplant. 2017;17 (suppl 3).

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