【摘要】：乳腺炎是奶牛常見的疾病之一,是由于機(jī)體代謝和生理狀態(tài)改變、乳房損傷并感染病原微生物等引起的乳腺炎癥反應(yīng),對奶牛業(yè)的危害巨大。除乳腺上皮細(xì)胞外,乳腺間質(zhì)成纖維細(xì)胞(Mammary stromal fibroblast,MSF)是構(gòu)成乳腺組織的重要組成部分,在小鼠和人類乳腺的正常發(fā)育及乳腺相關(guān)疾病中發(fā)揮著重要的作用。為了研究MSF是否參與奶牛乳腺的炎癥反應(yīng)過程,本研究分別從正常泌乳期奶牛和患臨床型乳腺炎奶牛的乳腺組織分離并純化出正常乳腺間質(zhì)成纖維細(xì)胞(Normal MSF,N-MSF)和炎癥相關(guān)乳腺間質(zhì)成纖維細(xì)胞(Inflammation-related MSF,In-MSF)進(jìn)行體外培養(yǎng),并通過顯微鏡、CCK-8細(xì)胞增殖實驗、Annexin V/PI流式細(xì)胞凋亡法、熒光定量PCR和ELISA等方法對兩者進(jìn)行了形態(tài)學(xué)、細(xì)胞增殖、細(xì)胞凋亡、炎性相關(guān)基因表達(dá)等生物學(xué)特性的對比。結(jié)果發(fā)現(xiàn),N-MSF與In-MSF在形態(tài)上和細(xì)胞凋亡率無明顯差異;但I(xiàn)n-MSF的增殖速率明顯高于N-MSF(P0.05);與N-MSF相比較,TNF-a、IL6和IL8三種炎性相關(guān)細(xì)胞因子在In-MSF的mRNA表達(dá)水平均顯著增高(P0.05);同時In-MSF分泌的TNF-a、IL6和IL8蛋白水平均顯著的高于N-MSF(P0.01,P0.05)。結(jié)果表明,乳腺間質(zhì)成纖維細(xì)胞在奶牛發(fā)生乳腺炎后其生物學(xué)特性發(fā)生了一系列的變化,這些變化可能進(jìn)一步影響乳腺炎的發(fā)生與發(fā)展,這一發(fā)現(xiàn)將為奶牛乳腺炎的診治和預(yù)防提供新的思路。
[Abstract]:Mastitis is one of the common diseases in dairy cows, which is caused by changes in metabolism and physiological state, breast injury and infection of pathogenic microorganisms, which is harmful to dairy industry. In addition to mammary epithelial cells, (Mammary stromal fibroblast is an important component of mammary gland tissue, which plays an important role in the normal development of mouse and human mammary glands and breast related diseases. In order to study whether MSF is involved in the inflammatory reaction of dairy cow mammary gland, In this study, normal breast stromal fibroblasts (Normal MSFN-MSF) and inflammatory associated breast stromal fibroblasts (Inflammation-related MSF-In-MSF) were isolated and purified from breast tissues of normal lactating cows and clinical mastitis cows. The biological characteristics such as morphology, cell proliferation, apoptosis and inflammatory related gene expression were compared by flow cytometry with Annexin V/PI, fluorescence quantitative PCR and ELISA. The results showed that there was no significant difference in morphology and apoptosis between N-MSF and In-MSF. However, the proliferation rate of In-MSF was significantly higher than that of N-MSF (P0.05); compared with N-MSF, the levels of mRNA expression of TNF-afi-IL6 and IL8 were significantly increased in In-MSF (P0.05), and the levels of TNF-afi-IL-6 and IL8 protein secreted by In-MSF were significantly higher than those of N-MSF (P0.01p0.05). The results showed that there were a series of changes in the biological characteristics of breast stromal fibroblasts after mastitis, which may further affect the occurrence and development of mastitis. This discovery will provide new ideas for the diagnosis, treatment and prevention of dairy cow mastitis.
【學(xué)位授予單位】：西北農(nóng)林科技大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2015
【分類號】：S858.23

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