We aimed at analyzing the structure of extracellular polysaccharide A from Grifola frondosa (EXGFP-A) and testing its immunomodulatory activity. Structural analysis shows that EXGFP-A was a contained α-D-glucoside bond and pyranose ring. GC analysis reveals that EXGFP-A was mainly composed of rhamnose, arabinose, xylose, mannose, glucose, galactose, by the molar ratio of 0.28:0.31:0.30:0.06:7.98:0.61. The results of MTT(3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay indicates when EXGFP-A was at a concentration of 80 μg/mL and treatment time of 48 h, RAW264.7 cells proliferation index reached a maximum of 137.5%. Meanwhile, the AO staining showed that EXGFP-A activated RAW264.7 cells and improved the level of intracellular nucleic acid metabolism. In addition, in a certain range of concentration, EXGFP-A was able to increase the release of NO in RAW264.7 cells, and upregulate the mRNA expression of immunological factor TNF-α, IL-1β, IL-6, IL-12, IFN-γ and iNOS of RAW264.7 cells. Our results confirm that EXGFP-A had immunomodulatory activity. Our findings provided scientific basis for the structural analysis and application of Grifola frondosa polysaccharide.
对灰树花胞外多糖A 组分 (EXGFP-A) 进行结构分析和免疫活性的研究。结构分析结果表明，EXGFP-A 是一种主要含有葡萄糖的吡喃型中性多糖。气相结果说明EXGFP-A 的单糖组成为鼠李糖、阿拉伯糖、木糖、甘露糖、葡萄糖、半乳糖，摩尔比为0.28∶0.31∶0.30∶0.06∶7.98∶0.61。MTT(3-(4,5-Dimethylthiazol-2-yl)-2,5- diphenyltetrazo-lium bromide)比色实验表明，当EXGFP-A 浓度为80 μg/mL，作用48 h 时，RAW264.7细胞增殖指数达到最大值，为137.5%。吖啶橙 (AO) 染色结果表明EXGFP-A 能够激活RAW264.7 细胞，增强细胞内部核酸代谢水平。EXGFP-A 在一定浓度范围内，可以提高RAW264.7 细胞中NO 的释放量，上调细胞内TNF-α、IL-1β、IL-6、IL-12、IFN-γ 细胞因子以及细胞中iNOS 的mRNA 水平的表达。结果表明，EXGFP-A具有一定的免疫调节活性，为灰树花胞外多糖的结构分析和应用提供了科学依据。.
Keywords: Grifola frondosa polysaccharide; RAW264.7 cells; immunomodulatory activity; structural analysis.