Acute vascular rejection (AVR) and systemic inflammation in xenograft recipients (SIXR) negatively impact the xenografts survival, and novel immunosuppressants are required to improve survival outcomes. We previously reported that TJ-M2010-5, a myeloid differentiation factor 88 (MyD88) inhibitor, exerts excellent anti-rejection effects in allogeneic transplantation. The aim of the present study was to evaluate the efficacy of TJ-M2010-5 in preventing AVR and SIXR and to investigate whether combined treatment of TJ-M2010-5 with anti-CD154 antibody (MR1) could prolong xenograft survival furthermore.A model involving heart transplantation from Sprague-Dawley rats to BALB/c mice was established in vivo, and the xenografts developed typical AVR. Bone marrow-derived dendritic cells and macrophages were cultured to study the underlying mechanisms induced by rat cardiomyocyte lysate stimulation in vitro.TJ-M2010-5 monotherapy prolonged xenograft survival, although combination treatment with MR1 further enhanced the anti-AVR and anti-SIXR effects with about 21 days graft survival, compared to monotherapy. TJ-M2010-5 reduced dendritic cell and macrophage activation induced by xenotransplantation, downregulated CD80/CD86 expression, suppressed B-cell activation and anti-donor antibody generation, reduced pro-inflammatory cytokine production and tissue factor expression, and attenuated epigenetic modifications underlying interleukin-6 and tumor necrosis factor-α production in macrophages by inhibiting nuclear factor kappa B nuclear translocation.TJ-M2010-5 attenuated AVR and SIXR and contributed to xenograft survival by inhibiting dendritic cell and macrophage activation. A dual-system inhibition strategy combining TJ-M2010-5 with anti-CD154 antibody achieved better results in xenotransplantation.Copyright © 2024. Published by Elsevier B.V.