Alternatively Activated (M2) Macrophage Phenotype Is Inducible by Endothelin-1 in Cultured Human Macrophages

• Published in: PloS one Vol. 11; no. 11; p. e0166433
• Main Authors: Soldano, Stefano, Pizzorni, Carmen, Paolino, Sabrina, Trombetta, Amelia Chiara, Montagna, Paola, Brizzolara, Renata, Ruaro, Barbara, Sulli, Alberto, Cutolo, Maurizio
• Format: Journal Article
• Language: English
• Published: United States Public Library of Science 15.11.2016; Public Library of Science (PLoS)
• Subjects: 12-O-Tetradecanoylphorbol-13-acetate; Acetic acid; Aging; Antigens, CD - genetics; Antigens, CD - immunology; Antigens, Differentiation, Myelomonocytic - genetics; Antigens, Differentiation, Myelomonocytic - immunology; Biology and Life Sciences; Cancer therapies; CD163 antigen; Cell Differentiation - drug effects; Cell Line; Chemokine CCL22 - genetics; Chemokine CCL22 - immunology; Cytokines; Diabetes mellitus; Diabetes mellitus (non-insulin dependent); Endothelin; Endothelin 1; Endothelin Receptor Antagonists - pharmacology; Endothelin-1 - pharmacology; Fibrosis; Gelatinase B; Gene expression; Gene Expression Regulation; Genes; Genetic aspects; Genotype & phenotype; Growth factors; Humans; Immunocytochemistry; Interleukin 10; Interleukin 4; Interleukin-10 - genetics; Interleukin-10 - immunology; Interleukin-4 - pharmacology; Interleukins; Internal medicine; Laboratories; Lectins, C-Type - genetics; Lectins, C-Type - immunology; Macrophage Activation - drug effects; Macrophages; Macrophages - cytology; Macrophages - drug effects; Macrophages - immunology; Mannose; Mannose-Binding Lectins - genetics; Mannose-Binding Lectins - immunology; Markers; Matrix Metalloproteinase 9 - genetics; Matrix Metalloproteinase 9 - immunology; Medicine; Medicine and Health Sciences; Metalloproteinase; Monocytes; Monocytes - cytology; Monocytes - drug effects; Monocytes - immunology; Monosaccharides; Pathogenesis; Peripheral blood; Phenotype; Polymerase chain reaction; Primary Cell Culture; Receptor, Endothelin A - genetics; Receptor, Endothelin A - immunology; Receptors; Receptors, Cell Surface - genetics; Receptors, Cell Surface - immunology; Research and Analysis Methods; Rheumatology; Scavenger receptors; Scavenger Receptors, Class A - genetics; Scavenger Receptors, Class A - immunology; Scleroderma (Disease); Studies; Sulfonamides - pharmacology; Systemic sclerosis; Tetradecanoylphorbol Acetate - pharmacology; Transforming Growth Factor beta1 - genetics; Transforming Growth Factor beta1 - immunology; Transforming growth factor-b1; Transforming growth factors; Type 2 diabetes; Western blotting; Italy
• ISSN: 1932-6203; 1932-6203
• DOI: 10.1371/journal.pone.0166433

Alternatively activated (M2) macrophages are phenotypically characterized by the expression of specific markers, mainly macrophage scavenger receptors (CD204 and CD163) and mannose receptor-1 (CD206), and participate in the fibrotic process by over-producing pro-fibrotic molecules, such as transforming growth factor-beta1 (TGFbeta1) and metalloproteinase (MMP)-9. Endothelin-1 (ET-1) is implicated in the fibrotic process, exerting its pro-fibrotic effects through the interaction with its receptors (ETA and ETB). The study investigated the possible role of ET-1 in inducing the transition from cultured human macrophages into M2 cells. Cultured human monocytes (THP-1 cell line) were activated into macrophages (M0 macrophages) with phorbol myristate acetate and subsequently maintained in growth medium (M0-controls) or treated with either ET-1 (100nM) or interleukin-4 (IL-4, 10ng/mL, M2 inducer) for 72 hours. Similarly, primary cultures of human peripheral blood monocyte (PBM)-derived macrophages obtained from healthy subjects, were maintained in growth medium (untreated cells) or treated with ET-1 or IL-4 for 6 days. Both M0 and PBM-derived macrophages were pre-treated with ET receptor antagonist (ETA/BRA, bosentan 10-5M) for 1 hour before ET-1 stimulation. Protein and gene expression of CD204, CD206, CD163, TGFbeta1 were analysed by immunocytochemistry, Western blotting and quantitative real time polymerase chain reaction (qRT-PCR). Gene expression of interleukin(IL)-10 and macrophage derived chemokine (CCL-22) was evaluated by qRT-PCR. MMP-9 production was investigated by gel zymography. ET-1 significantly increased the expression of M2 phenotype markers CD204, CD206, CD163, IL-10 and CCL-22, and the production of MMP-9 in both cultures of M0 and PBM-derived macrophages compared to M0-controls and untreated cells. In cultured PBM-derived macrophages, ET-1 increased TGFbeta1 protein and gene expression compared to untreated cells. The ET-1-mediated effects were contrasted by ETA/BRA treatment in both cultured cell types. ET-1 seems to induce the M2 phenotype in cultured human macrophages, a process apparently contrasted by the action of the ETA/BRA, suggesting possible clinical implications in those fibrotic diseases characterized by increased ET-1 concentrations, such as systemic sclerosis but also type 2 diabetes.