短管兔耳草多糖的结构表征及其免疫调节活性研究

doi:10.11843/j.issn.0366-6964.2021.011.028

摘要/Abstract

摘要： 本试验旨在从短管兔耳草（Lagotis brevituba Maxim）中分离纯化多糖，初步分析多糖的结构表征并研究其体外免疫调节活性。采用水提醇沉法从短管兔耳草中提取粗多糖，经DEAE-52纤维素离子交换柱进行分离纯化，得到短管兔耳草多糖（Lagotis brevituba Maxim polysaccharide，LMP）。采用高效凝胶色谱法（HPGPC）、傅里叶变换-红外光谱法（FT-IR）、紫外光谱法（UV）等方法对其结构进行初步分析，并以小鼠骨髓来源树突状细胞（murine bone marrow derived dendritic cells，BMDCs）为靶细胞，采用MTT法、流式细胞术（FCM）、酶联免疫吸附试验（ELISA）等对其免疫活性进行测定。结果显示：从短管兔耳草中分离到不含核酸和蛋白质的多糖LMP，得率为（18.5±1.7）%，糖含量为（89.7±1.9）%，平均分子量为3.18 ku，平均粒径为1 483.89 nm，Zeta电位为-14.81 mV。扫描电镜（SEM）和原子力显微镜（AFM）结果显示，LMP的表面光滑，呈片状分布，单链的高度在5.4 nm左右。3.13～50.00 μg·mL^-1 LMP能显著促进BMDCs增殖（P<0.05）及其表面分子（CD80、CD86、MHC-Ⅰ和MHC-Ⅱ）的表达（P<0.05）和细胞因子（TNF-α、IL-12）的分泌（P<0.05），同时显著降低BMDCs的吞噬能力（P<0.05）。LMP能促进BMDCs表型和功能的成熟，具有较好的免疫调节活性，可将其作为潜在的免疫调节剂。

关键词: 短管兔耳草, 多糖, 结构特征, 树突状细胞, 免疫活性

Abstract: The purpose of this study was to isolate the polysaccharide from Lagotis brevituba Maxim (LMP) and to investigate the structural features and its immunomodulatory activity in vitro. LMP was obtained by water extraction and ethanol precipitation, and purified sequentially by DEAE-52 column chromatography. Subsequently, high performance gel permeation chromatography (HPGPC), fourier transform infrared spectroscopy (FT-IR), ultraviolet spectroscopy (UV) were employed to characterize the structural properties of LMP. The immunomodulatory activities of LMP were studied by MTT, FCM and ELISA, and using murine bone marrow derived dendritic cells (BMDCs) as the target cells. The results showed that LMP was obtained from Lagotis brevituba Maxim and contained no nucleic acid and protein. The yield and total carbohydrate content of LMP were 18.5%±1.7% and 89.7%±1.9%, respectively, and the average molecular weight of LMP was 3.18 ku. In addition, the average particle size and Zeta potentials of LMP were 1 483.89 nm and-14.81 mV, respectively. SEM and AFM images showed that LMP was a sheet-like appearance with smooth surface and uniformly dispersed in height of 5.4 nm. At 3.13-50 μg·mL^-1, LMP promoted the cell proliferation of BMDCs, and significantly increased the expression of surface markers (CD80, CD86, MHC-Ⅰ and MHC-Ⅱ) (P<0.05) and the secretion content of cytokines (TNF-α and IL-12) (P<0.05). Moreover, LMP decreased the phagocytosis of BMDCs (P<0.05). LMP showed better immune activity and it could induce the maturation of the phenotype and functions of BMDCs, which would be potentially developed as an effective immunomodulatory agent.

Key words: Lagotis brevituba Maxim, polysaccharides, characterization, dendritic cells, immunomodulation

中图分类号:

S853.74

余蕊宏, 孟祯, 孙梦珂, 陈仕雄, 张浚文, 黄一帆, 秦韬, 任喆. 短管兔耳草多糖的结构表征及其免疫调节活性研究[J]. 畜牧兽医学报, 2021, 52(11): 3270-3281.

YU Ruihong, MENG Zhen, SUN Mengke, CHEN Shixiong, ZHANG Junwen, HUANG Yifan, QIN Tao, REN Zhe. Structural Characterization and Immunomodulatory Activities of Polysaccharide Extracted from Lagotis brevituba Maxim[J]. Acta Veterinaria et Zootechnica Sinica, 2021, 52(11): 3270-3281.

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