IRE1α/XBP1 suppression reprograms macrophage plasticity and Treg induction to enhance ethylene-carbodiimide-fixed donor splenocytes-induced cardiac allograft tolerance.

In this study, we elucidate how IRE1α/XBP1 suppression reprograms macrophage plasticity and regulatory T cells (Tregs) induction to amplify ethylene-carbodiimide-fixed donor splenocytes (ECDI-SPs)-mediated cardiac allograft tolerance. Murine cervical heterotopic heart transplantation models revealed prolonged cold ischemia time (4 h) impaired ECDI-SPs-mediated graft protection, correlating with elevated IL-6 and suppressed IL-10 level. Pharmacological IRE1α inhibition via 4μ8C bidirectionally regulated macrophage plasticity: suppressing LPS + IFNγ-induced M1 polarization while amplifying IL-4/IL-13-driven M2 polarization. Mechanistically, 4μ8C enhanced efferocytosis and expanded CD206+ M2 macrophages, accompanied by reduced pro-inflammatory cytokines and anti-inflammatory cytokine elevation. Furthermore, co-culture experiments demonstrated that 4μ8C-treated bone marrow-derived macrophages promoted the differentiation of CD4+CD25+Foxp3+ Tregs from CD4+ T cells, which was dependent on secretion of IL-10 and TGF-β from macrophages. Combined therapy synergistically extended cardiac allograft survival, reducing leukocyte infiltration while amplifying M2 macrophages and Tregs. Our findings establish that IRE1α/XBP1 suppression coordinates macrophage/Treg crosstalk to optimize ECDI-SPs efficacy, providing a translatable strategy to overcome ischemia-reperfusion injury and enhance transplant outcomes.