Effects of Angelica polysaccharide on differentiation and function in M2 macrophages

Abstract

Objective To investigate the effect of Angelica polysaccharide (APS) on the differentiation and function of M2 macrophages and underlying molecular mechanism. Methods Mouse bone marrow derived macrophages (BMDM) and M2 macrophages were induced and treated with APS (0, 80, 160, 320 μg/mL); Mouse peritoneal macrophages were isolated and treated with APS (0, 160 μg/mL). Flow cytometry (FCM) was used to detect mannose receptor(MR), CD11b, F4/80, CD163, and ARG-1 expression levels, apoptosis, and phagocytic ability of M2 macrophages and peritoneal macrophages. Mice were randomly divided into APS gavage group and control group, APS was intragastrically administered to mice, and macrophage MR expression level in blood and spleen were detected by FCM. Fluorescence microscopy was used to observe the morphology of BMDM-differentiated M2 macrophages. RT-qPCR was employed to detect the mRNA expression levels of MR and ARG-1 in M2 macrophages. Immunofluorescence assay was performed to detect the expression of the proteins related to molecular mechanism of differentiation and function of M2 macrophages. Results Compared with the 0 μg/mL APS group, the MR expression level in the M2 macrophages was decreased with the increase of APS concentration within a certain concentration range (80~320 μg/mL), and the MR expression level in peritoneal macrophages was also decreased in the 160 μg/mL APS treatment group (P < 0.01). The expression level of macrophage MR was also significantly decreased in peripheral blood and spleen in the APS gavage mice than the control group (P < 0.05). Compared with the 0 μg/mL APS group, the expression levels of CD11b, F4/80, and CD163 in the macrophages were increased in the 80 ~ 320 μg/mL APS treatment groups (P < 0.01). The morphology of macrophage had changed, from mostly spindle-shaped and pseudopodia to mostly round or irregular, and even a few cells with pseudopodia. APS induced apoptosis in M2 macrophages (P < 0.05). Compared with the 0 μg/mL APS group, M2 macrophages treated with 160 μg/mL APS had an increased ability to phagocytose fluorescent microspheres (P < 0.01), but the expression level of ARG-1 was decreased (P < 0.01). The mRNA expression of MR and ARG-1 in M2 macrophages was decreased (P < 0.05). The mean fluorescence intensity of phosphate acidified-signal transducers and activators of transcription 6(p-STAT6)-positive signals in M2 macrophages was significantly reduced in the 160 μg/mL APS-treated group (P < 0.05). Conclusion APS has bidirectional regulation on the differentiation and function of M2 macrophages, which may be associated with its downregulation of signal transducers and activators of transcription 6(STAT6) signaling pathway.

Keywords

• angelica polysaccharide
• mannose receptor
• m2 macrophages
• signal transducers and activators of transcription 6