Abstract
The Notch pathway plays critical roles in the differentiation and polarized activation of macrophages; however, the downstream molecular mechanisms underlying Notch activity in macrophages remain elusive. Our previous study has identified a group of microRNAs that mediate Notch signaling to regulate macrophage activation and tumor-associated macrophages (TAMs). In this study, we demonstrated that miR-148a-3p functions as a novel downstream molecule of Notch signaling to promote the differentiation of monocytes into macrophages in the presence of granulocyte macrophage colony-stimulating factor (GM-CSF). Meanwhile, miR-148a-3p promoted M1 and inhibited M2 polarization of macrophages upon Notch activation. Macrophages overexpressing miR-148a-3p exhibited enhanced ability to engulf and kill bacteria, which was mediated by excessive production of reactive oxygen species (ROS). Further studies using reporter assay and Western blotting identified Pten as a direct target gene of miR-148a-3p in macrophages. Macrophages overexpressing miR-148a-3p increased their ROS production through the PTEN/AKT pathway, likely to defend against bacterial invasion. Moreover, miR-148a-3p also enhanced M1 macrophage polarization and pro-inflammatory responses through PTEN/AKT-mediated upregulation of NF-κB signaling. In summary, our data establish a novel molecular mechanism by which Notch signaling promotes monocyte differentiation and M1 macrophage activation through miR-148a-3p, and suggest that miR-148a-3p-modified monocytes or macrophages are potential new tools for the treatment of inflammation-related diseases.
Highlights
Macrophages have pivotal roles in development, homeostasis, and host defense
To assess whether miR-148a-3p could serve as a molecule downstream of Notch in macrophage regulation, we examined the expression of miR-148a-3p in macrophages with disrupted or ectopically activated Notch signaling, which is clarified by the expression level of HES1, one downstream molecule of Notch signaling (Figures S1A,B in Supplementary Material)
In BM-derived macrophages (BMDMs) polarized with PBS (M0), LPS + interferon γ (IFN-γ) (M1), or IL-4 (M2), Recombination signal-binding protein Jκ (RBP-J) deficiency significantly reduced the expression of miR-148a-3p (Figure 1B)
Summary
Macrophages have pivotal roles in development, homeostasis, and host defense. Ontogenically, macrophages originate from embryonic primitive and definitive hematopoiesis, as well as hematopoiesis in bone marrow (BM) in adults [1, 2]. Hematopoietic stem cells in BM give rise to common myeloid progenitors, which produce monocytes through several intermediates, Abbreviations: BM, bone marrow; BMDMs, BM-derived macrophages; iNOS, inducible nitric oxide synthase; FIH1, factor inhibiting hypoxia-inducible factor 1; MR, mannose receptor; ROS, reactive oxygen species; RBP-J, recombination signalbinding protein Jκ; NIC, Notch intracellular domain; TAMs, tumor-associated macrophages. Lipopolysaccharide (LPS) and interferon γ (IFN-γ)-stimulated macrophages, or M1 macrophages, upregulate the expression of tumor necrosis factor (TNF)-α, interleukin (IL)-6, IL-12, and inducible nitric oxide synthase (iNOS, official symbol NOS2), accompanied by increased antigen presentation and bactericidal capacity. IL-4-activated macrophages (M2 macrophages) express higher levels of immunosuppressive cytokines, such as IL-10 and transforming growth factor β, together with arginase-1, mannose receptor (MR; official symbol MRC1), and other molecules involved in anti-inflammation and tissue remodeling [3, 4]. The multi-modality of macrophage activation has been implicated in many human diseases, including inflammatory diseases and cancer [5]
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