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MyD88抑制剂ST2825对丝状支原体山羊亚种 感染宿主细胞的影响
张俊波,印双红,张红,陆安法
0
(铜仁学院农林工程与规划学院(铜仁市文化科技产业创新研究中心),铜仁 554300;铜仁学院大健康学院,铜仁 554300;铜仁市动物疫病预防控制中心,铜仁 554300)
摘要:
为分析MyD88在丝状支原体山羊亚种(Mmc)感染肺泡上皮细胞中的分子机制。以山羊肺泡上皮细胞为材料,用不同浓度的MyD88抑制剂(ST2825)与细胞孵育,MTT法检测细胞活性,试剂盒检测caspase-3的活性,以及实时定量PCR检测MyD88TNF-a mRNA的表达水平,试剂盒检测H2O2浓度和NO浓度。当ST2825浓度为5、10 和20 μmol·L-1不影响细胞活性,而浓度为40 μmol·L-1时细胞活性显著降低;感染比例(MOI)为10,感染时间为8 、12 和24 h,Mmc可以显著提高MyD88 mRNA的表达水平(P<0.01);可显著提高caspase-3的活性,而ST2825能够显著降低caspase-3(P<0.05);感染时间为24 h,Mmc可显著提高TNF-a mRNA的表达水平、H2O2浓度和NO浓度(P<0.05),而ST2825在10和20 μmol·L-1的浓度时,可显著降低Mmc介导的caspase-3活性、H2O2浓度以及LDH水平(P<0.05),ST2825在5、10和20 μmol·L-1浓度时可显著降低TNF-a mRNA的表达水平和NO浓度(P<0.05)。
关键词:  丝状支原体山羊亚种  MyD88抑制剂  致病机制
DOI:10.13610/j.cnki.1672-352x.20180825.005
基金项目:国家自然科学基金项目(31502067), 中国博士后科学基金项目(2017M613003), 铜仁学院博士科研启动基金项目(trxyDH1504), 贵州省教育厅自然科学研究重点项目(黔教合KY字(2015)411号), 贵州省科技合作计划项目(黔科合LH字[2015]7238; 黔科合LH字[2015]7236号)和贵州省教育厅青年科技人才成长项目[黔教合KY字[2017]312]共同资助。
Effect of MyD88 inhibitor ST2825 on host cells infected with mycoplasma mycoides subspecies capri
ZHANG Junbo,YIN Shuanghong,ZHANG Hong,LU Anfa
(College of Agroforestry Engineering and Planning (Institute of Cultural and Technological Industry Innovation of Tongren), Tongren University, Tongren 554300;College of One Health, Tongren University, Tongren 554300;Animal Disease Prevention and Control Center of Tongren City, Tongren 554300)
Abstract:
To study the molecular mechanism of MyD88 in alveolar epithelial cells infected by filamentous mycoplasma goat subspecies, the goat alveolar epithelial cells were treated with different concentrations of inhibitor ST2825. The cell activity was detected by MTT?Assay?Kit and the caspase-3 activity was measured using Caspase-3?Colorimetric Assay Kit from APExBIO Company. Expression levels?of?TNF-a?mRNA?were analyzed by real-time quantitative PCR,?H2O2 concentration?was measured using?H2O2?Assay Kit,?and NO concentration?was measured using NO Assay Kit. The cell activity was not affected by ST2825?at the concentrations of 5, 10 and 20 μmol·L-1, but it was significantly decreased on the concentration of 40 μmol·L-1. When the infection ratio (MOI) was 10, the expression of?MyD88?mRNA was increased at 8 h, 12 h and 24 h post-infection, and the activity of caspase-3 was also significantly increased by the subspecies of mycelial carcinoma (P<0.01), while the expression of caspase-3 was significantly decreased?by ST2825 (P<0.05).?Expression levels of TNF-a mRNA and the concentrations of H2O2?and NO?were?significantly enhanced by?Mycoplasma at 24 h post-infection
Key words:  mycoplasma mycoides subspecies capri  MyD88 inhibitor  pathogenesis

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