*Purpose: Infiltration of allografts by inflammatory macrophages is frequently associated with chronic graft loss In both animal models and clinical settings, but the mechanisms that control such inflammatory macrophages remain poorly defined.

*Methods: In the present study we examined the BET family protein BRD4 (an epigenetic modifier) in regulation of macrophage phenotypes in a heart transplant model and how changes in macrophage phenotypes affected transplant survival.

*Results: We firstly established an in vitro model of macrophage polarization and used the chemical inhibitor JQ1, which blocks BRD4 binding to the acetylated histones, to examine the impact of BRD4 inhibition on macrophage phenotypes. We found that JQ1 potently suppressed the induction of inflammatory macrophages, and most prominently the induction of the M2 phenotype. Further analysis by RNA-seq showed that the expression of M2-associated transcription factors KLF4, KLF9, IRF4 and ppar-γ was markedly suppressed upon BRD4 inhibition, demonstrating that BRD4 likely controls the chromatin accessibility and transcriptional programs of M2 cells. Additionally, ChIP-qPCR experiments showed that binding of M2-related transcription factors at the Arg-1 locus was markedly reduced after JQ1 treatment, and the expression of Arg-1 enhancer RNA (eRNA) was abolished after JQ1 treatment. Studies using gene knockout mice showed that the nuclear localization of p-STAT6 was profoundly reduced after JQ1 treatment. Importantly, in a fully MHC mismatched heart transplant model (BALB/c to B6), we observed that treatment of recipient mice with a low dose of CTLA4-Ig inhibited the acute rejection but allowed the development of chronic allograft rejection, which was accompanied by extensive macrophage infiltration in the grafts. Surprisingly, additional treatment of transplant recipients with the BRD4 inhibitor JQ1 induced long-term heart allograft survival (>100 days) with no histological signs of chronic rejection. Analysis of graft-infiltrating cells showed that the most striking effect of JQ1 treatment was the inhibition of M2 macrophages in vivo.

*Conclusions: These novel data suggest that BRD4 inhibition is a new strategy for preventing chronic allograft rejection and a key underlying mechanism for graft survival is the suppression of inflammatory macrophages.

« Back to 2022 American Transplant Congress