Objective. Chronic rejection of transplanted organs remains the main obstacle in the long-term success of organ transplantation. Chronic rejection occurs in 40-45% of recipients at 5 years following transplantation. Thus, our goal was to uncover the mechanisms of chronic rejection, which will undoubtedly revolutionize heart transplant medicine. It is known that macrophages play a critical role in the injury and repair of transplanted organs. Novel anti-rejection strategies in organ transplantation target macrophages to abrogate transplant rejection. Multiple macrophage functions depend on actin cytoskeleton, which is under the control of small GTPase RhoA and its downstream effector ROCK1. Methods We generated mice with macrophage specific deletion of RhoA. Hearts from BALB/c (H-2d) donors were transplanted into RhoAflox/flox (no Cre), heterozygous Lyz2Cre+/-RhoA+/flox and Lyz2Cre+/-RhoAflox/flox recipients treated with CTLA4-Ig to inhibit early T cell response, and transplanted hearts were pathologically assessed for signs of chronic rejection. Macrophage responses to RhoA deletion were studied by actin staining, immunostaining, PCR, Western blotting, flow cytometry, and phagocytosis, matrix degradation and cell migration in vitro assays. Results. Macrophage-specific deletion of RhoA in conjunction with CTLA4-Ig abrogated chronic rejection of allografts and inhibited macrophage infiltration into the grafts. RhoA deletion increased ROCK1 protein level and its activity and affected macrophage structure, polarity and actin-related functions. Increased ROCK1 activity was partially dependent on non-apoptotic Caspase-3 cleavage and was sensitive to Caspase-3 inhibition. Conclusions. These novel findings indicate that interference with the RhoA pathway inhibits macrophage infiltration of the graft and modifies macrophage polarity and functions. This, in turn, will help to design novel macrophage-targeted anti-rejection therapies.

CITATION INFORMATION: Liu Y, Chen W, Li X, Ghobrial R, Kloc M. Macrophage Specific Deletion of RhoA Impedes Macrophage Functions and Abrogates Chronic Rejection of Cardiac Allograft. Am J Transplant. 2016;16 (suppl 3).

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