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目的探讨哺乳动物雷帕霉素靶蛋白(mTOR)信号通路在慈菇多糖提高RAW 264.7巨噬细胞功能中的作用。方法体外培养RAW 264.7巨噬细胞,分别用ELISA和Griess法检测不同剂量慈菇多糖(5、50、500μg/m L)和香菇多糖(100μg/m L)对巨噬细胞分泌白介素2(IL-2)、肿瘤坏死因子α(TNF-α)、一氧化氮(NO)的影响;用RT-PCR和Western blot检测各组巨噬细胞mTOR/第10染色体缺失与张力蛋白同源的磷酸酶基因(PTEN)mRNA和蛋白表达变化。结果与对照组比较,慈菇多糖低、中、高剂量组,香菇多糖组巨噬细胞内IL-2含量[分别为(68.216±1.609)、(70.850±6.669)、(89.634±8.399)、(84.367±8.165)pg/m L]上升;慈菇多糖中、高剂量组,香菇多糖组巨噬细胞内TNF-α含量[分别为(46.989±7.895)、(70.739±7.452)、(48.125±6.757)pg/m L]上升,慈菇多糖高剂量组巨噬细胞内NO值[为(7.199±1.242)μmol/L]上升,差异均有统计学意义(均P<0.01);慈菇多糖中、高剂量组RAW 264.7细胞mTOR mRNA表达低于对照组(P<0.05);慈菇多糖高剂量组、香菇多糖组mTOR蛋白表达低于对照组(P<0.05)。结论慈菇多糖可能通过mTOR相关信号通路提高巨噬细胞功能。
Objective To investigate the role of mammalian rapamycin target protein (mTOR) signaling pathway in the enhancement of the function of Sagittaria polysaccharides in RAW 264.7 macrophages. Methods RAW 264.7 macrophages were cultured in vitro. The levels of interleukin 2 (IL-2β) secreted by macrophages were detected by ELISA and Griess method with different dosages of fructus polysaccharide (5,50 and 500 μg / ml) and lentinan 2, TNF-α and NO were detected by enzyme-linked immunosorbent assay (ELISA). The phosphatase gene homologue of mTOR / chromosome 10 in macrophages was detected by RT-PCR and Western blot (PTEN) mRNA and protein expression changes. Results Compared with the control group, the levels of IL-2 in macrophages of low, medium and high dose group of Lentinus edodes polysaccharides (68.216 ± 1.609, (70.850 ± 6.669), (89.634 ± 8.399, 84.367 ± 8.165) pg / m L]. The content of TNF-α in the macrophages in the medium, high dose and lentinan groups [(46.989 ± 7.895), (70.739 ± 7.452) and (48.125 ± 6.757 ) pg / m L], the intracellular macrophage NO level [(7.199 ± 1.242) μmol / L] was increased in the high dose group (all P <0.01) , And the mRNA expression of mTOR in RAW 264.7 cells in high dose group was lower than that in control group (P <0.05). The expression of mTOR protein in high dose group and lentinan group was lower than that in control group (P <0.05). Conclusion Lentinus edodes polysaccharides may enhance the function of macrophages through mTOR related signaling pathway.