Differential effects of Smad2 and Smad3 in regulation of macrophage phenotype and function in the infarcted myocardium

TGF-βs regulate macrophage responses, by activating Smad2/3. We have previously demonstrated that macrophage-specific Smad3 stimulates phagocytosis and mediates anti-inflammatory macrophage transition in the infarcted heart. However, the role of macrophage Smad2 signaling in myocardial infarction remains unknown. We studied the role of macrophage-specific Smad2 signaling in the healing infarct, and we explored the basis for the distinct effects of Smad2 and Smad3. Infarct macrophages exhibited both Smad2 and Smad3 activation. In contrast to the effects of Smad3 loss, myeloid cell-specific Smad2 disruption had no effects on mortality, ventricular dysfunction and adverse remodeling, after myocardial infarction. Phagocytic removal of dead cells, macrophage and myofibroblast infiltration, collagen deposition, angiogenesis and scar remodeling were not affected by macrophage Smad2 loss. In isolated macrophages, TGF-β1, -β2 and -β3, activated both Smad2 and Smad3, whereas BMP6 triggered only Smad3 activation. Smad2 and Smad3 had similar patterns of nuclear translocation in response to TGF-β1. Smad3, and not Smad2, was the main mediator of transcriptional effects of TGF-β on macrophages and Smad3 loss resulted in enrichment of genes associated with RAR/RXR signaling, cholesterol biosynthesis and lipid metabolism. In conclusion, the in vivo and in vitro effects of TGF-β on macrophage function involve Smad3, and not Smad2. This study is designed to compare gene expression profiles between Smad2fl/fl and MyS2KO (Smad2fl/fl + LyzMCre), and Smad3fl/fl and MyS3KO (Smad3fl/fl + LyzMCre) bone marrow derived macrophages in the presence or absence of 10 ng/ml TGF-β1. The loss of Smad2 and Smad3 in MyS2KO, MyS3KO macrophages has been documented by qPCR and western blot.