Immunomodulatory effects of sulfated polysaccharides of pine pollen on mouse macrophages

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• 作者：Yue Geng ; ^{gengy@sdnu.edu.cn" class="auth_mail" title="E-mail the corresponding author} ; ^{yuegeng@sohu.com" class="auth_mail" title="E-mail the corresponding author} ; Li Xing ; Mengmeng Sun ; Fangchen Su
• 关键词：Sulfated polysaccharide ; Macrophages ; [Ca2+]i
• 刊名：International Journal of Biological Macromolecules
• 出版年：2016
• 出版时间：October 2016
• 年：2016
• 卷：91
• 期：Complete
• 页码：846-855
• 全文大小：2862 K

文摘

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.