Regulation of M1‑type and M2‑type macrophage polarization in RAW264.7 cells by Galectin‑9
Published online on: October 4, 2017
Generally considered as a potent pro‑inflammatory signal, β‑galactosidelectin suppresses T cell receptor activation, can both promote and inhibit integrin‑mediated adhesion and is required in nuclear pre‑mRNA splicing. Galectin‑9 (Gal‑9), a member of β‑galactoside lectin, is involved many processes of T cell‑mediated diseases (such as autoimmune diseases and asthma) and immunomodulation of macrophages. Macrophages are involved in the occurrence of inflammation, development and digestion and other stages. At different stages of the inflammatory response, macrophages exhibit different phenotypes, but mainly two subtypes, classically (M1) or alternatively (M2) polarization. The purpose of this work is to investigate the effect of overexpression or knockdown of Gal‑9 on the macrophage polarization. Macrophage polarization was detected by flow cytometric profiling of secreted cytokines and specific surface markers expression, including nitric oxide synthase 2 (NOS2) and mannose receptor 1 (CD206). Protein and mRNA expression levels of TNF‑α, TGF‑β, IL‑6, IL‑10, NF‑κB, signal transducer and activator of transcription (Stat)1 and Stat3 were determined by ELISA, western blot analysis or qRT‑PCR. Our results implied that differentiation of the mouse macrophage line RAW264.7 into M1‑type and M2‑type macrophages is followed by marked variations of Gal‑9 expression. Furthermore, its overexpression and secretion are tightly associated with M2‑type macrophages, whereas its downregulation promotes macrophages to polarize into M1‑type macrophages, which confirmed by elevated CD206 and NOS2, respectively. In response to the changes of Gal‑9 expression, cytokines, transcription factors and regulators, including TNF‑α, IL‑6, NF‑κB, Stat1, TGF‑β, IL‑10, and Stat3, were tightly regulated and significantly associated with classically and alternatively activated macrophages. Consistent with characteristics of M1‑type macrophages, the transcriptional or translational expression levels or activity of TNF‑α, IL‑6, Stat1 and NF‑κB were markedly increased with knockdown of Gal‑9 in macrophages. By contrast, the expression levels or activity of TGF‑β, IL‑10 and Stat3 were clearly elevated in macrophages with Gal‑9 overexpression, which is closely related with M2‑type macrophages. Specific expression and secretion patterns of cytokines, transcription factors and regulators in M1‑type and M2‑type macrophages contribute to better understanding the role of Gal‑9 in regulation in macrophages. This study provides a new insight that Gal‑9 may be a new immunomodulatory target for macrophages.