The role of endogenous Smad7 in regulating macrophage phenotype following myocardial infarction [JunLi_MyS7KOinvivo]. The role of endogenous Smad7 in regulating macrophage phenotype following myocardial infarction [JunLi_MyS7KOinvivo]

Induction of the inhibitory Smad, Smad7 serves as a negative feedback mechanism that restrains TGF-b-mediated actions in injured tissues. In inflammatory cells, Smad7 has been suggested to exert both pro-inflammatory actions attributed to inhibition of TGF-b-induced suppression of inflammation, and anti-inflammatory effects due to disruption of the TAK-1/NF-kB system. Myocardial infarction triggers a macrophage-driven inflammatory response that plays a central role in cardiac repair, but also contributes to adverse remodeling and fibrosis. We hypothesized that Smad7 upregulation in infarct macrophages may play a modulatory role in cardiac repair, by restraining effects of TGF-b. To test the hypothesis, we investigated the response of Myeloid cell-specific Smad7 knockout mice (MyS7KO) following myocardial infarction protocols, and we examined the in vitro effects of Smad7 in isolated macrophages. Smad7 was upregulated in a subset of infarct macrophages, peaking 7 days after infarction. Myeloid cell-specific Smad7 loss did not affect baseline macrophage gene expression and had no significant effects on homeostatic functions. Although RNA-seq analysis predicted that, in the absence of Smad7, infarct macrophages may have attenuated activation of inflammatory pathways and suppressed TREM1 signaling, myeloid cell-specific Smad7 loss had no significant effects on ventricular dysfunction, adverse remodeling, scar remodeling and collagen deposition after myocardial infarction. In isolated macrophages, TGF-b attenuated pro-inflammatory cytokine and chemokine expression, modulated synthesis of matrix remodeling genes, and had profound effects on macrophage profile, inducing genes associated with activation of sphingosine-1 phosphate and integrin signaling pathways, and inhibiting cholesterol biosynthesis genes. However, RNA-seq and PCR array experiments showed that Smad7 loss has minimal effects on TGF-b-mediated macrophage responses, restraining synthesis of only a small fraction of TGF-b-induced genes, such as Itga5, Olfml3 and Fabp7. Smad7 absence did not affect the anti-inflammatory actions of TGF-b in TNF-stimulated cells. In conclusion, our findings suggest a limited role for macrophage Smad7 in regulation of post-infarction inflammation and repair, and demonstrate that the anti-inflammatory effects of TGF-b in macrophages are not restrained by endogenous Smad7 induction. Overall design: In order to study the role of Smad7 in myeloid cells in vivo, we generated mice with myeloid specific loss of Smad7 in lysozyme M+ cells (MyS7KO). We used a transgenic mouse line in which Cre recombinase is driven by the LyzM promoter (Jackson Laboratory, stock No: 004781). LyzM-Cre mice were bred with Smad7fl/fl mice (Jackson Laboratory, stock No: 017008) to generate LyzM-Cre;Smad7fl/fl animals (MyS7KO) and corresponding Smad7fl/fl control littermates. A model of non-reperfused myocardial infarction was induced by permanent ligation of left anterior descending coronary artery and samples were collected for baseline mice, and 3 and 7 days after coronary occlusion.