BACKGROUND: Chronic rejection is the leading cause of progressive allograft function decline. Studies have demonstrated that CD40-CD40L-induced paired immunoglobulin-like receptor-A (PIR-A) is the MHC-I receptor necessary for the specific memory response in macrophages of mice with chronic rejection. However, the underlying mechanisms remain unclear. METHODS: BALB/c mouse hearts were transplanted into C57BL/6, RelB RESULTS: CD40 and PIR-A were highly expressed and colocalized in macrophage-infiltrating allograft in the mouse model. CD40-CD40L blockade inhibited PIR-A expression and prolonged allograft survival. Conditional deletion of Pira in recipient's macrophages inhibited chronic rejection and promoted long-term allograft acceptance. Mechanistically, CD40 may activate transcription factor NF-κB2 translocation into the nucleus to up-regulate PIR-A expression, promoting chronic rejection of cardiac transplantation. NF-κB2 regulated PIR-A expression by binding to the intergenic region of Pira. CONCLUSIONS: Our data suggest that Pira is a potential target to induce long-term allograft tolerance. CD40 may activate transcription factor NF-κB2 translocation into the nucleus to up-regulate PIR-A expression, promoting chronic rejection of cardiac transplantation. The study findings provide novel therapeutic opportunities to promote transplant survival in clinical settings.