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

Macrophages 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.