畜牧兽医学报 ›› 2019, Vol. 50 ›› Issue (4): 781-790.doi: 10.11843/j.issn.0366-6964.2019.04.011

• 营养与饲料 • 上一篇    下一篇

酿酒酵母β-葡聚糖通过膜受体Dectin-1和TLR-2诱导绵羊瘤胃上皮细胞表达SBD-1的机制研究

张曼1,2, 金鑫1,2, 王云鹤1,2, 魏方1,2, 温婧怡1,2, 杨银凤1,2*   

  1. 1. 内蒙古农业大学兽医学院, 呼和浩特 010018;
    2. 农业部动物疾病临床诊疗技术重点试验室, 呼和浩特 010018
  • 收稿日期:2018-10-30 出版日期:2019-04-23 发布日期:2019-04-23
  • 通讯作者: 杨银凤,主要从事反刍动物消化道黏膜免疫研究,E-mail:julie1963@163.com
  • 作者简介:张曼(1990-),女,辽宁沈阳人,博士生,主要从事反刍动物消化道黏膜免疫研究,E-mail:zmimau@163.com
  • 基金资助:

    国家自然科学基金(31560682)

Mechanism of Saccharomyces Cerevisiae β-glucan Inducing SBD-1 Expression in Ovine Rumen Epithelial Cells by Membrane Receptors Dectin-1 and TLR-2

ZHANG Man1,2, JIN Xin1,2, WANG Yunhe1,2, WEI Fang1,2, WEN Jingyi1,2, YANG Yinfeng1,2*   

  1. 1. College of Veterinary Medicine, Inner Mongolia Agricultural University, Hohhot 010018, China;
    2. Key Laboratory of Clinical Diagnosis and Treatment Technology in Animal Disease of Ministry of Agriculture, Hohhot 010018, China
  • Received:2018-10-30 Online:2019-04-23 Published:2019-04-23

摘要:

为了探索酿酒酵母β-葡聚糖诱导绵羊瘤胃上皮细胞(ovine ruminal epithelial cells,ORECs)β-防御素-1(sheep β-defensin-1,SBD-1)表达过程中膜受体Dectin-1和TLR-2可能存在的作用。本研究首先利用免疫组化、RT-PCR、免疫荧光和Western blot等方法检测Dectin-1是否在ORECs内表达,并且采用qPCR和Western blot方法对β-葡聚糖刺激ORECs后细胞膜受体Dectin-1和TLR-2的表达变化进行检测。而后用不同浓度的Dectin-1阻断剂昆布多糖或TLR-2特异性封闭抗体分别预处理ORECs后,采用qPCR和ELISA方法检测SBD-1的表达变化,以确定β-葡聚糖诱导SBD-1表达过程中Dectin-1和TLR-2的参与情况。结果显示:1)绵羊瘤胃组织及ORECs内存在Dectin-1表达,且β-葡聚糖刺激ORECs后Dectin-1和TLR-2的表达水平显著增加(P<0.05);2)不同浓度的昆布多糖和TLR-2封闭抗体均可以极显著降低β-葡聚糖诱导SBD-1的表达(P<0.01),且随着阻断剂和封闭抗体浓度的增加,其抑制SBD-1表达的作用越明显。结果表明,Dectin-1在绵羊瘤胃组织及ORECs内均表达,并且酿酒酵母β-葡聚糖在ORECs中诱导SBD-1的表达是由Dectin-1和TLR-2介导产生的。

关键词: β-葡聚糖, SBD-1, 绵羊瘤胃上皮细胞, Dectin-1, TLR-2

Abstract:

The aim of this study was to explore the possible role of TLR-2 and Dectin-1 in the expression of β-defensin-1 (SBD-1) in ovine ruminal epithelial cells (ORECs) induced by Saccharomyces cerevisiae β-glucan. The immunohistochemistry, RT-PCR, immunofluorescence and Western blot were used to detect the expression of Dectin-1 in ORECs, and qPCR and Western blot were used to detected the expression changes of Dectin-1 and TLR-2 after β-glucan stimulated ORECs. Then, ORECs were pretreated using Dectin-1 blocker laminarin or TLR-2 blocking antibody with different concentrations, and the expression of SBD-1 were detected by qPCR and ELISA to determine the involvement of Dectin-1 and TLR-2 in the induction expression of SBD-1 induced by β-glucan. The results showed that:1) Dectin-1 was expressed in ovine ruminal tissues and ORECs, and the expression levels of Dectin-1 and TLR-2 were significantly increased after β-glucan stimulating ORECs (P<0.05); 2)The laminarin or TLR-2 blocking antibody with different concentrations significantly decreased the expression of SBD-1 induced by β-glucan (P<0.01), and this effect became stronger as the laminarin and blocking antibodies concentration increased. The results indicate that Dectin-1 is expressed in both ovine ruminal tissues and ORECs, and the Saccharomyces cerevisiae β-glucan inducing SBD-1 expression in ORECs is mediated by Dectin-1 and TLR-2.

Key words: β-glucan, SBD-1, ovine ruminal epithelial cells, Dectin-1, TLR-2

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