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2025年5月10日 星期六
  2018, Vol. 26 Issue (12): 2132-2141    
  研究论文与报告 本期目录 | 过刊浏览 | 高级检索 |
金黄色葡萄球菌和大肠杆菌感染BMECs对细胞因子和受体表达及细胞凋亡的影响
赵凌1,潘阳阳2,何翃闳3,李秦4,丁天翊1,张慧珠1,樊江峰2,崔燕2,余四九2
1. 甘肃省牛羊胚胎工程技术研究中心
2. 甘肃农业大学
3. 甘肃农业大学动物医学院
4.
Effect on the Expression of Cytokine, Receptor and Apoptosis in BMECs by Staphylococcus aureus and Escherichia coli Infected
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摘要 奶牛(Bos taurus)乳腺炎的产生使奶牛业的发展受到制约,奶牛健康遭到破坏的同时,乳制品的质量也随之进行下降。病原微生物是乳腺炎产生的主要源头,细胞的凋亡由病原微生物引起,而且能够干扰病症的发展过程。本研究旨在探究金黄色葡萄球菌(Staphylococcus aureus)和大肠杆菌(Escherichia coli)感染奶牛乳腺上皮细胞(bovine mammary epithelial cells, BMECs)对细胞因子和受体的表达及细胞凋亡的影响。本实验通过体外培养BMECs并进行免疫化学细胞鉴定,选取纯化的第5代BMECs,分别加入0、101、102、103、104 CFU/mL (CFU: 菌落形成单位, colony-forming units)浓度的S. aureus或E. coli,经过公式计算,以最佳感染量0.6 mL感染细胞,实时荧光定量PCR (quantitative real-time PCR, qRT-PCR)对细胞中白细胞介素1β (interleukin-1, IL-1β)和受体IL1RⅠ及肿瘤坏死因子(tumor necrosis factor, TNF-α)的基因表达量进行检测,通过末端脱氧核苷酸转移酶介导的dUTP缺口末端标记测定法(terminal dexynucleotidyl transferase (TdT)-mediated dUTP nick end labeling, TUNEL)对细胞凋亡进行检测。结果表明,本实验所培养的细胞不表达波形蛋白,表达角蛋白18和β-酪蛋白,故培养的是BMECs。分别以0、101、102、103、104 CFU/mL的S. aureus处理细胞12 h,IL1RⅠ、IL-1β、TNF-α的表达量分别在浓度为102、104、103 CFU/mL时最高。分别以0、101、102、103、104 CFU/mL的E. coli处理12 h,IL1RⅠ、IL-1β、TNF-α的表达量分别在浓度101、103、104 CFU/mL时最高。利用浓度为101、102、103、104 CFU/mL的S. aureus或E. coli对细胞进行处理,在浓度为104 CFU/mL时凋亡率最高。本研究结果表明,不同浓度菌液感染BMECs,细胞因子表达呈上升趋势,其受体表达呈下降趋势,并诱导细胞出现凋亡现象。本实验为研究因细菌感染而导致乳腺炎的致病机制提供了科学依据。
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赵凌
潘阳阳
何翃闳
李秦
丁天翊
张慧珠
樊江峰
崔燕
余四九
关键词 奶牛金黄色葡萄球菌大肠杆菌乳腺上皮细胞qRT-PCR    
Abstract:The occurrence of cow (Bos taurus) mastitis has restricted the development of dairy industry. It damages the cow health and reduces the quality of dairy products.The main cause of mastitis is pathogenic microorganism. Pathogenic microorganisms could induce cell apoptosis and thus interfere the development of the disease process. The purpose of this study was to explore the expression of cytokines and receptors, and the level of cell apoptosis in bovine mammary epithelial cells (BMECs) infected by Staphylococcus aureus and Escherichia coli. This study was intended to explore the S. aureus and E. coli (with fabrication: Oli) infected BMECs on the expression of cytokine. The BMECs were cultured in vitro and identified by immunohistochemistry, then the purified fifth generation of BMECs were selected and infected S. aureus or E. coli with the concentration of 0, 101, 102, 103, 104 CFU/mL, respectively. By the formula computing, the best infected quantity of S. aureus and E.coil was 0.6 mL; and the number of cell apoptosis was detected by one-step TUNEL (terminal dexynucleotidyl transferase (TdT)-mediated dUTP nick end labeling). The expression of interleukin-1β (IL-1β), IL1RⅠ and TNF-α were detected by means of qRT-PCR detection. The results showed that cells expressed not only keratin 18 also beta casein. In addition, the BMECs didn't express vimentin. The cultured cells were identified as BMECs. The highest mRNA expression of IL1RⅠ, IL-1β and TNF-α were at the S.aureus concentrations of 102 CFU/mL, 104 CFU/mL and 103 CFU/mL, respectively. The highest mRNA expression of IL1RⅠ, IL-1β and TNF-α were at the E. coli concentrations of 101 CFU/mL, 103 CFU/mL and 104 CFU/mL, respectively. The apoptosis rate was the highest at 104 CFU/mL with a concentration of cells treated by S.aureus or E. coli. The result indicated that different concentrations of bacterial infecting BMEC could increase the expression of cell factor, while decreased the expression of the receptor, which led to apoptosis of BMEC. This consequence could provide scientific basis for the pathogenesis of mastitis caused by bacterial infection .
Key wordsBovine    Staphyloccus aureus    Escherichia coli    Mammary epithelial cells    qRT-PCR
收稿日期: 2018-03-09      出版日期: 2018-11-20
ZTFLH:  S823.8+9  
基金资助:高寒环境相关调节因子对牦牛卵母细胞成熟及附植前不同来源胚胎发育调控的作用机制;奶牛场常见病防控技术示范推广;牦牛泌乳调控相关基因筛选与功能分析
通讯作者: 余四九     E-mail: sjyu@163.com
引用本文:   
赵凌 潘阳阳 何翃闳 李秦 丁天翊 张慧珠 樊江峰 崔燕 余四九. 金黄色葡萄球菌和大肠杆菌感染BMECs对细胞因子和受体表达及细胞凋亡的影响[J]. , 2018, 26(12): 2132-2141.
链接本文:  
http://journal05.magtech.org.cn/Jwk_ny/CN/     或     http://journal05.magtech.org.cn/Jwk_ny/CN/Y2018/V26/I12/2132
 
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