无乳链球菌感染对奶牛乳腺上皮细胞乳蛋白合成影响的分子机制研究

Streptococcus agalactiae is the main pathogen of subclinical breast inflammation induced by dairy cows. Its infection is significantly associated with dairy cow immune function and milk quality, especially milk protein synthesis. The project is based on a large sample analysis to determine the correlation between Streptococcus agalactiae and bovine milk cells (SCC), total bacterial count and milk quality in dairy cows, and to reveal the milk-like milk (STLR2 and sTLR4, proinflammatory cytokines IL-6, etc.) were detected by LCMS / MS and gel electrophoresis techniques. The specific biomarkers in milk samples and blood samples were screened and analyzed. To investigate the effect of Streptococcus agalactiae on the physical and chemical indexes of mammary gland and the lactation physiology of mammary gland epithelial cells. The mammary epithelial cell model was constructed and the protein phosphorylation group was used to study the effects of streptococcus The changes of signal molecules associated with the synthesis of milk protein caused by the process of inflammation of mammary gland epithelial cells were confirmed by RT-PCR and Western blot analysis. The relationship between the key pathways of inflammation and the pathways of protein synthesis was revealed. Molecular Mechanism of Streptococcus Aeruginis on Milk Protein Synthesis in Dairy Cows.

无乳链球菌作为诱发奶牛亚临床乳腺炎症的主要致病菌，它的感染与奶牛免疫机能和牛乳品质尤其是乳蛋白合成显著相关。本项目拟以大样本分析检测奶牛原奶中无乳链球菌与牛乳体细胞数（SCC）、总细菌计数以及牛乳品质等的相关性为基础，揭示无乳链球菌检出乳样乳成分的品质特征；利用LCMS/MS、凝胶电泳等技术对乳样和血样中的特定生物标记物进行筛选、检测并对血中特定细胞因子（sTLR2和sTLR4，促炎细胞因子IL-6等）的mRNA表达变化进行鉴定，研究和揭示无乳链球菌感染对奶牛血液理化指标和乳腺上皮细胞泌乳生理的作用机理；构建乳腺上皮细胞模型，采用蛋白质磷酸化组学技术研究无乳链球菌感染后奶牛乳腺上皮细胞炎症发生过程所导致的与乳蛋白合成相关的信号分子的变化，并通过RT-PCR、Western印迹分析验证其表达变化，揭示炎症发生关键通路与乳蛋白合成通路之间的关系以及无乳链球菌影响奶牛乳腺乳蛋白合成的分子机制。

本项目立足于奶牛养殖行业的市场需求，对当前制约奶牛业健康发展的奶牛乳房炎进行机制研究与探索。无乳链球菌不仅是引起奶牛乳腺炎的主要致病菌，同时也是重要的人畜共患性致病菌，因此具有重要的公共卫生意义。本研究以无乳链球菌为切入点，通过临床乳房炎血液与乳汁大样本采集分析，鉴定出特定的生物标记物，并探讨潜在的生物标志物与乳汁微生物及代谢物之间的关系，明确了无乳链球菌型乳腺炎相关免疫与炎性指标的生物标记物，为进一步检测该类型乳房炎提供了重要数据支撑；通过体外培养奶牛乳腺上皮细胞系，建立了无乳链球菌与乳腺上皮细胞共感染的体外损伤模型，解答了无乳链球菌型奶牛乳腺上皮细胞损伤及其对泌乳信号转导通路的影响，深入探究了无乳链球菌型乳腺炎乳腺上皮细胞模型的蛋白质组学变化规律。同时，本研究通过苦参碱、大豆异黄酮等植物提取物对无乳链球菌体外抑菌活性相关研究，更好的阐明了无乳链球菌的毒力因子及其调控机制，为治疗和预防无乳链球菌型乳腺炎，及筛选无乳链球菌的疫苗靶点指明了重点研发的方向和目标。随着细胞水平及分子水平研究的深入、基因组学、转录组学和蛋白质组学等方面技术的发展，逐渐加深了对无乳链球菌的全面认识。本项目为开发无乳链球菌型乳腺炎的诊断试剂盒提供有利的数据支撑，同时对治疗和预防无乳链球菌型乳房炎具有重要意义。特别是高科技的发展及中医药理论的全新解读，对无乳链球菌的致病机制和防控手段的研究将愈加完备，这对于加快推进绿色健康养殖，保障我国人民的乳品安全具有重要意义，可产生较大的经济、社会和生态效益。