Immunomodulatory effects of sulfated polysaccharides of pine pollen on mouse macrophages

被引:65
作者
Geng, Yue [1 ]
Xing, Li [1 ]
Sun, Mengmeng [1 ]
Su, Fangchen [1 ]
机构
[1] Shandong Normal Univ, Coll Life Sci, Key Lab Anim Resistance Biol Shandong Prov, Jinan 250014, Peoples R China
关键词
Sulfated polysaccharide; Macrophages; Ca2+](i); ACTIVATION; CALCIUM;
D O I
10.1016/j.ijbiomac.2016.06.021
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
This study was undertaken to explore the effects of sulfated polysaccharide (SPPM60-D) from masson pine pollen on [Ca2+](i) and immune function of RAW264.7 macrophages. SPPM60-D was obtained by subjecting Masson pine pollen to boiling water and alcohol precipitation, 60% ethanol grading precipitation, Sephacryl S-400HR purification, and chlorosulfonic acid-pyridine method sulfation. An 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay was used to measure the effect of SPPM60-D on relative activity and proliferation of RAW264.7 cells, and a fluorescence spectrophotometer was used to determine [Ca2+](i). Phagocytosis of neutral red was used to determine phagocytosis capacity. Adherence, scratch healing, and transwell assays were used to assess migration and adhesion abilities of macrophages. An enzyme-linked immuno sorbent assay (ELISA) assay was used to assess the secretion of cytokines and inflammatory mediators. A dexamethasone (DEX) inhibition method was used to measure the recovery of RAW264.7 immune activity by SPPM60-D. SPPM60-D significantly increased relative activity, proliferation, and [Ca2+](i) levels of mouse RAW264.7 cells. It also significantly enhanced the immune function of macrophages from normal and immune-suppressed mouse. The results showed that SPPM60-D mainly bound to TLR4 on macrophages. This activated the TLR4-PI3K-PLC-IP3R signaling pathway, leading to the opening of calcium release-activated calcium channels (CRAC), increasing [Ca2+](i) and activating the macrophages, and thus improving immunity. (C) 2016 Published by Elsevier B.V.
引用
收藏
页码:846 / 855
页数:10
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