Macrophage/monocyte-specific deletion of Ras homolog gene family member A (RhoA) downregulates fractalkine receptor and inhibits chronic rejection of mouse cardiac allografts

International audienceBACKGROUND: The cellular and molecular mechanisms of chronic rejection of transplanted organs remain obscure; however, macrophages are known to play a critical role in the injury and repair of allografts. Among multiple factors influencing macrophage infiltration to allografts, the fractalkine chemokine (C-X3-C motif) ligand 1(CX3CL1)/chemokine (C-X3-C motif) receptor 1 (CX3CR1) signaling pathway and actin cytoskeleton, which is regulated by a small guanosine-5׳-triphosphatase Ras homolog gene family member A (RhoA), are of the utmost importance. To define the role of macrophage/RhoA pathway involvement in chronic rejection, we generated mice with monocyte/macrophage-specific deletion of RhoA.METHODS: Hearts from BALB/c (H-2d) donors were transplanted into RhoA(flox/flox) (no Cre) and heterozygous Lyz2(Cre+/-)RhoA(flox/flox) recipients treated with cytotoxic T-lymphocyte-associated protein 4 immunoglobulin to inhibit early T-cell response. Allografts were assessed for chronic rejection and monocyte/macrophage functions.RESULTS: The deletion of RhoA inhibited macrophage infiltration, neointimal hyperplasia of vasculature, and abrogated chronic rejection of the allografts. The RhoA deletion downregulated G protein-coupled fractalkine receptor CX3CR1, which activates the RhoA pathway and controls monocyte/macrophage trafficking into the vascular endothelium. This in turn promotes, through overproliferation and differentiation of smooth muscle cells in the arterial walls, neointimal hyperplasia.CONCLUSIONS: Our finding of codependence of chronic rejection on monocyte/macrophage CX3CR1/CX3CL1 and RhoA signaling pathways may lead to the development of novel anti-chronic rejection therapies

Dissonant response of M0/M2 and M1 bone-marrow-derived macrophages to RhoA pathway interference

International audienceMacrophages have a multitude of functions in innate and adaptive immune response and organ and tissue homeostasis. Many experimental studies are performed on bone-marrow-derived macrophages differentiated in vitro into M1 (inflammatory) and M2 (anti-inflammatory) subtypes that express different molecular markers pertaining to their prospective functions. Macrophage phenotype, polarity and functions depend on the actin cytoskeleton, which is regulated by small GTPase RhoA, its downstream effector ROCK, and non-apoptotic Caspase-3. We generated transgenic mice with the macrophage-specific deletion of RhoA and compared the effect of Rho pathway interference (RhoA deletion and ROCK and Caspase-3 inhibition) on the phenotype, polarity and expression of subtype-specific molecular markers of bone-marrow-derived M0, M1 and M2 macrophages. We show that M0 and M2 macrophages have a radically different phenotype and polarity from M1 macrophages, and that this is mirrored in dissonant response to RhoA pathway interference. The RhoA pathway interference induces extreme elongation (hummingbird phenotype) of M0 and M2 but not M1 macrophages and inhibits the expression of M2-specific but not M1-specific molecular markers. These dramatic differences in the response of M0/M2 versus M1 macrophages to the same molecular cues ought to be important considerations in the interpretation of experimental data and therapeutic use of bone-marrow-derived macrophages