奶牛乳腺上皮细胞在炎性状态下对基质成纤维细胞生物学特性及功能的影响

发布时间：2018-06-05 23:26

本文选题：乳腺炎 + 乳腺上皮细胞　；参考：《西北农林科技大学》2017年硕士论文

【摘要】：奶牛乳腺炎是由于乳房损伤或病原微生物入侵所引起的机体代谢和生理状态改变的炎症反应,是严重影响奶牛业发展的最常见的疾病之一。在奶牛乳腺发生炎症的早期,乳腺导管中的病原菌会首先侵入腺泡表层的乳腺上皮细胞,进而影响乳腺上皮细胞的生理特性,导致上皮细胞的炎性细胞因子异常分泌。乳腺上皮细胞异常分泌的炎性细胞因子通过内环境影响周围组织中基质细胞的生理特性,使基质细胞初步表现出炎症反应特征。成纤维细胞是构成乳腺组织重要的基质细胞,其生长状态的改变会直接影响乳腺组织正常的结构和功能,因此研究奶牛乳腺上皮细胞在炎性状态下对基质成纤维细胞生物特性和功能的功能影响对于奶牛乳腺炎的防治具有重要的意义。金黄色葡萄球菌和大肠杆菌是引起奶牛乳腺炎主要的两种细菌,脂多糖(lipopolysaccharides,LPS)和脂磷壁酸(lipoteichoic acid,LTA)分别是这两种导致奶牛乳腺炎的细菌主要毒力成分。本试验以荷斯坦奶牛乳腺组织分离的原代乳腺上皮细胞作为研究对象,首先利用LPS和LTA对乳腺上皮细胞进行炎症诱导,确定了LPS和LTA炎症诱导的最适浓度和最适时间,建立了体外诱导的炎性奶牛乳腺上皮细胞模型;然后通过体外间接共培养模式,以LPS和LTA体外诱导的炎性乳腺上皮细胞与正常基质成纤维细胞共培养的方法模拟炎性上皮细胞影响成纤维细胞过程,研究炎性乳腺上皮细胞对基质成纤维细胞活性、增殖、迁移、凋亡、炎性细胞因子分泌等生物学特性的影响,同时对导致这些特性发生变化的分子机制进行探究。结果表明,LPS和LTA诱导的炎性上皮细胞均可促进了基质成纤维细胞的激活,通过MAPK信号通路抑制成纤维细胞的增殖,通过WNT/β-catenin信号通路促进了成纤维细胞的迁移、同时增强成纤维细胞对TNF-α,CCL5,CXCL2,IL6,IL8等炎性相关因子的表达和分泌,诱导细胞外基质蛋白Fibronection和Collagen I的沉积。这些结果表明奶牛乳腺上皮细胞在炎性状态下,通过内环境对基质成纤维细胞生炎症响应。这一结果为进一步研究乳腺基质成纤维细胞在奶牛乳腺炎反应中的作用及奶牛乳腺炎的防治提供了一定的理论依据。
[Abstract]:Cow mastitis is an inflammatory response to metabolic and physiological changes caused by breast injury or invasion of pathogenic microorganisms. It is one of the most common diseases that seriously affect the development of dairy cattle industry. In the early stage of breast inflammation in dairy cows, the pathogenic bacteria in mammary duct will first invade the epithelial cells in the surface of the acinus, and then affect the physiological characteristics of the epithelial cells of the mammary gland, leading to the abnormal secretion of inflammatory cytokines in the epithelial cells. The abnormal secretion of inflammatory cytokines by mammary epithelial cells affects the physiological characteristics of stromal cells in the surrounding tissues through the internal environment and makes the stromal cells exhibit the characteristics of inflammatory reaction. Fibroblasts are the most important stromal cells in mammary gland. The change of their growth state will directly affect the normal structure and function of mammary gland. Therefore, it is of great significance to study the effects of dairy mammary epithelial cells on the biological characteristics and function of stromal fibroblasts in inflammatory state for the prevention and treatment of dairy cow mastitis. Staphylococcus aureus and Escherichia coli are two main bacteria causing dairy cow mastitis, lipopolysaccharide polysaccharide LPSs and lipoteichoic acididate LTAs are the main virulence components of these two bacteria. In this experiment, primary mammary epithelial cells isolated from breast tissue of Holstein dairy cattle were used as the research object. First, LPS and LTA were used to induce the inflammation of breast epithelial cells, and the optimal concentration and time of LPS and LTA inflammation induction were determined. The model of inflammatory dairy cow mammary epithelial cells induced in vitro was established, and the model of indirect co-culture in vitro was established. Inflammatory mammary epithelial cells induced by LPS and LTA were co-cultured with normal matrix fibroblasts in vitro to simulate the effect of inflammatory epithelial cells on the process of fibroblasts and to study the activity and proliferation of inflammatory mammary epithelial cells on stromal fibroblasts. The effects of migration, apoptosis, inflammatory cytokine secretion, and the molecular mechanisms that lead to the changes of these characteristics are also explored. The results showed that both lipopolysaccharide and LTA stimulated the activation of stromal fibroblasts, inhibited the proliferation of fibroblasts through MAPK signaling pathway, and promoted the migration of fibroblasts through WNT/ 尾 -catenin signaling pathway. At the same time, the expression and secretion of inflammatory related factors such as TNF- 伪 CCL5, CXCL2, IL6, IL-8 were enhanced in fibroblasts, and the deposition of extracellular matrix proteins (Fibronection and Collagen I) was induced. These results suggest that dairy mammary epithelial cells in inflammatory state, through the internal environment to matrix fibroblast inflammatory response. The results provide a theoretical basis for the further study of the role of mammary stromal fibroblasts in the response of dairy cow mastitis and the prevention and treatment of cow mastitis.
【学位授予单位】：西北农林科技大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：S858.23

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