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中国农学通报

中国农学通报 ›› 2023, Vol. 39 ›› Issue (11): 122-128.doi: 10.11924/j.issn.1000-6850.casb2022-0389

• 畜牧·动物医学·蚕·蜂 • 上一篇    下一篇

脂多糖诱导奶牛子宫内膜上皮细胞炎性损伤的研究

陈佳佳(), 白欣洁()   

  1. 北京农业职业学院，北京 102442
  • 收稿日期:2022-05-09 修回日期:2022-09-06 出版日期:2023-04-15 发布日期:2023-04-10
  • 通讯作者: 白欣洁，女，1987年出生，河北衡水人，副教授，博士，研究方向：基础兽医学。通信地址：102401 北京市房山区稻田南里5号，E-mail：bxj0318@qq.com。
  • 作者简介:

    陈佳佳，女，1989年出生，河南洛阳人，实验师，硕士，研究方向：基础兽医学。通信地址：102401 北京市房山区稻田南里5号，E-mail：。

  • 基金资助:
    北京市特色高水平院校建设项目“打造高水平专业群动物医学专业群技术平台与社会服务建设项目”(11000022T000000444734); 北京市自然科学基金青年项目“紧密连接蛋白在运动马出血性肠炎中的防御机制初探”(6214035)

Study on Inflammatory Injury of Endometrial Epithelial Cells Induced by Lipopolysaccharide in Dairy Cows

CHEN Jiajia(), BAI Xinjie()   

  1. Beijing Vocational College of Agriculture, Beijing 102442
  • Received:2022-05-09 Revised:2022-09-06 Online:2023-04-15 Published:2023-04-10

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摘要：

为了初步探讨大肠杆菌型奶牛子宫内膜炎发生机制，进而为后续筛选出作用于关键信号通路靶点的药物奠定基础。试验通过组织块培养法和传代培养分离纯化得到奶牛子宫上皮细胞(Bovine endometrial epithelial cell，bEEC)。30 µg/mL的脂多糖(Lipopolysaccharide，LPS)刺激bEEC 12 h后，收集细胞用于奶牛子宫内膜上皮细胞体外炎性损伤模型的研究。通过荧光定量RT-PCR检测LPS作用于bEEC 2、6、9、12 h后细胞内IL-1β、IL-8、IL-6、TNF-α mRNA表达，然后通过Western-blot 检测LPS诱导bEEC NF-кB和MAPKs相关蛋白的表达。结果表明：30 μg/mL的LPS作用bEEC细胞2、6、9、12 h可极显著(P<0.01)诱导IL-1β、IL-8、IL-6、TNF-α mRNA表达。LPS可上调MAPKs相关蛋白P38、ERK、JNK的磷酸化水平(P<0.01)，促使NF-κB抑制蛋白IκBα的降解(P<0.05)。说明LPS可激活MAPKs和NF-кB信号通路，诱导大量炎症介质的产生，从而对细胞造成炎性损伤。

关键词: 脂多糖, 奶牛子宫内膜炎, 上皮细胞, 细胞因子, MAPKs

Abstract:

The study aims to investigate the mechanism of endometrial inflammatory in cows with Escherichia coli, and lay a foundation for screening drugs that act on key signaling pathway targets. In this study, bovine endometrial epithelial cells (bEEC) were obtained and purified from cow uterus with primarily cultured tissue explant and trypsin. The cells by 30 μg/mL lipopolysaccharide (LPS) stimulating 12 h was screened out, which was used as the bEEC inflammatory injury model in vitro. IL-1β, IL-8, IL-6 and TNF-α mRNA levels of bEEC induced in 2 h, 6 h, 9 h and 12 h by LPS were detected with RT-PCR, and NF-κB and MAPKs related proteins in LPS-bEEC cells were detected by Western-blot. The results showed that IL-1β, IL-8, IL-6 and TNF-α mRNA were highly expressed in 2 h, 6 h, 9 h and 12 h caused by 30 μg/mL LPS (P<0.01). LPS could up-regulate the phosphorylation levels of P38, ERK and JNK which were related with MAPKs (P<0.01), and promote the degradation of IκBα which was the inhibitory protein of NF-κB (P<0.05). MAPKs and NF-кB signaling pathway of bEEC could be activated by LPS, and then a large number of inflammatory mediators were released, causing inflammation damage of the cells.

Key words: lipopolysaccharide, cow endometritis, epithelial cells, cytokines, MAPKs