本文選題：結(jié)核分枝桿菌 + 趨化因子和趨化因子受體�。� 參考：《復旦大學》2005年博士論文

【摘要】：結(jié)核分枝桿菌(Mycobacterium tuberculosis)是已經(jīng)發(fā)現(xiàn)的最為重要的傳染性病原體之一，每年可以引起2—3百萬人感染結(jié)核病。結(jié)核分枝桿菌是典型的胞內(nèi)致病菌，它可以在宿主巨噬細胞內(nèi)存活。巨噬細胞和結(jié)核分枝桿菌的初始相互作用中免疫調(diào)節(jié)因子(包括細胞因子、趨化因子及其它們的受體)，，在細菌能否引起機體發(fā)病中可能起著關(guān)鍵作用。目前，對于結(jié)核桿菌感染中趨化因子和趨化因子受體的了解十分有限。 為了弄清楚趨化因子和趨化因子受體在結(jié)核桿菌感染的巨噬細胞中的變化，我們利用活化的人巨噬細胞系U937作為結(jié)核桿菌感染的體外模型，應(yīng)用基因芯片檢測技術(shù)、半定量PCR、流式細胞術(shù)等手段來研究巨噬細胞在不同感染階段(0小時，6小時，12小時和24小時)的趨化因子和趨化因子受體的表達譜�；蛐酒慕Y(jié)果顯示，結(jié)核分枝桿菌H37Rv感染巨噬細胞6小時和12小時都可以檢測到有變化的基因，感染24小時，明顯上調(diào)的基因有25條，其中顯著上調(diào)的基因有22條。半定量RT—PCR證實了部分基因(IL-8，MIP1-α，RANTES，CXCR4和CCR5)的芯片檢測結(jié)果。 我們在用基因芯片法檢測了結(jié)核桿菌感染巨噬細胞后的全部趨化因子和趨化因子受體的表達變化中，發(fā)現(xiàn)兩個重要的趨化因子受體CCR5和CXCR4是表達上調(diào)的，CCR5和CXCR4是人類艾滋病病毒HIV入侵人體細胞的兩個重要輔助受體，結(jié)合我們在臨床上觀察到的有一部分的艾滋病患者同時患有結(jié)核病，我們推測趨化因子受體CCR5和CXCR4在結(jié)核分枝桿菌感染后的表達上調(diào)可能是結(jié)核�。滩」不嫉牧硪粋�原因。 為了了解CCR5和CXCR4的上調(diào)表達是結(jié)核桿菌感染特異產(chǎn)生的還是炎癥類致病菌都能產(chǎn)生的普遍現(xiàn)象，我們又用另一種致病菌伴放線放線桿菌(Actinobacillus actinomycetemcomitans)去攻擊巨噬細胞，然后從mRNA和蛋白水平觀測CCR5和CXCR4的表達變化，結(jié)果表明伴放線放線桿菌也能刺激這兩個分子的上調(diào)表達。為了進一步揭示CCR5和CXCR4表達變化的分子機理，研究了結(jié)核分枝桿菌感染后巨噬細胞信號轉(zhuǎn)導通路的變化，詳細考察了CCR5和CXCR4的啟動子結(jié)構(gòu)，然后我們定位在MAPK通路和共同的調(diào)節(jié)因子YY1上。運用p38 MAPK的特異性抑制劑SB203580，我們研究其對CCR5，CXCR4和CCR5、CXCR4的共同的負調(diào)控轉(zhuǎn)錄因子YY1的表達影響。最終的實驗結(jié)果表明在我們的模型中 CCR5和CXCR4的上調(diào)主要是p38 MAPK介導的，YY1依賴，這個結(jié)果表明CCR5和CXCR4的上調(diào)和MTB／HIV共感染之間的幾乎沒有關(guān)聯(lián)。
[Abstract]:Mycobacterium tuberculosisis is one of the most important infectious pathogens which can cause 2 to 3 million people to become infected with tuberculosis every year. Mycobacterium tuberculosis is a typical intracellular pathogen that can survive in host macrophages. Immunomodulatory factors (including cytokines, chemokines and their receptors) play a key role in the initial interaction between macrophages and Mycobacterium tuberculosis. At present, the understanding of chemokines and chemokine receptors in Mycobacterium tuberculosis infection is very limited. In order to find out the changes of chemokines and chemokine receptors in macrophages infected by Mycobacterium tuberculosis, we used activated human macrophage cell line U937 as an in vitro model of Mycobacterium tuberculosis infection. Semi-quantitative PCR and flow cytometry were used to study the expression profiles of chemokines and chemokine receptors in macrophages at different stages of infection. The results of microarray showed that the mutated genes could be detected in macrophages infected with Mycobacterium tuberculosis H37Rv for 6 hours and 12 hours. After 24 hours of infection, 25 genes were obviously up-regulated, among which 22 genes were significantly up-regulated. Semi-quantitative RT-PCR confirmed the microarray detection results of IL-8 mil MIP1- 偽 RANTESX CR4 and CCR5. We detected the expression of all chemokines and chemokine receptors in macrophages infected with Mycobacterium tuberculosis by gene chip method. Two important chemokine receptors, CCR5 and CXCR4, were found to be two important coreceptors for the invasion of human cells by HIV / HIV. We speculate that the up-regulated expression of chemokine receptors CCR5 and CXCR4 after Mycobacterium tuberculosis infection may be another cause of TB / AIDS co-infection. To understand whether the upregulation of CCR5 and CXCR4 is specific to Mycobacterium tuberculosis infection or a common form of inflammatory pathogens, we used another pathogen, Actinobacillus actinomycetemcomitans., to attack macrophages. The expression of CCR5 and CXCR4 was observed at the level of mRNA and protein. The results showed that Actinobacillus actinomycetes could also stimulate the up-regulation of the expression of these two molecules. In order to further reveal the molecular mechanism of the expression of CCR5 and CXCR4, the changes of macrophage signal transduction pathway after Mycobacterium tuberculosis infection were studied, and the promoter structures of CCR5 and CXCR4 were investigated in detail. We then localize the MAPK pathway and the common regulatory factor YY1. Using SB203580, a specific inhibitor of p38 MAPK, we studied the effects of SB203580 on the expression of YY1, a common negative regulatory transcription factor, of CCR5CXCR4 and CCR5CXCR4. The final experimental results show that in our model, The up-regulation of CCR5 and CXCR4 was mainly mediated by p38 MAPK, which indicated that there was almost no association between the up-regulation of CCR5 and CXCR4 and MTB/HIV co-infection.
【學位授予單位】：復旦大學
【學位級別】：博士
【學位授予年份】：2005
【分類號】：R378

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