本文關(guān)鍵詞： emodin 核因子-κB 抑制蛋白-κB I-κB激酶 RNA干擾 脂多糖 誘生性一氧化氮合酶 腫瘤壞死因子-α 白介素-10 巨噬細(xì)胞 急性胰腺炎 急性肺損傷 一氧化氮 中西醫(yī)結(jié)合　出處：《大連醫(yī)科大學(xué)》2006年博士論文　論文類(lèi)型：學(xué)位論文

【摘要】：目的： 探討核因子-κB炎癥信號(hào)通路在全身炎癥反應(yīng)時(shí)肺泡巨噬細(xì)胞過(guò)度活化中的作用，觀察NF-κBp65／p50的活性和核易位情況、蛋白激酶IKK的活性變化、炎性細(xì)胞因子的表達(dá)水平，并深入探究蛋白激酶IKKαγ對(duì)上述炎癥信號(hào)通路中多個(gè)環(huán)節(jié)的協(xié)同調(diào)節(jié)效應(yīng)及Emodin治療作用的分子生物學(xué)機(jī)制；同時(shí)，進(jìn)一步揭示一氧化氮(NO)在急性重癥胰腺炎急性肺損傷大鼠發(fā)病中的雙重功能及該作用與肺泡巨噬細(xì)胞NF-κB活性變化的密切聯(lián)系，為臨床治療急性肺損傷等多種全身炎癥反應(yīng)性疾病、尋求更為合適的藥物作用靶點(diǎn)提供新的實(shí)驗(yàn)佐證。 方法： 實(shí)驗(yàn)一：以小鼠RAW264.7巨噬細(xì)胞系為研究對(duì)象，應(yīng)用逆轉(zhuǎn)錄PCR、蛋白雜交印跡、免疫細(xì)胞化學(xué)染色、免疫熒光染色等方法，觀察emodin對(duì)由LPS誘導(dǎo)產(chǎn)生的炎癥反應(yīng)的調(diào)節(jié)作用。共分為7組，每組5個(gè)培養(yǎng)皿：Group 1：RAW264.7巨噬細(xì)胞正常培養(yǎng)組；Group 2、3、4：給予LPS(101μg／ml)刺激，分三個(gè)時(shí)間點(diǎn)30min、2hr、5hr；Group 5、6：分為三個(gè)時(shí)間點(diǎn)，在給予LPS(10μg／ml)刺激的基礎(chǔ)上，，同時(shí)加入emodin(20gμg／ml)進(jìn)行干預(yù)，于不同作用時(shí)間點(diǎn)收集的樣本。 實(shí)驗(yàn)二：由NF-κB介導(dǎo)的過(guò)度炎癥反應(yīng)在許多炎性疾病的發(fā)生發(fā)展過(guò)程中發(fā)揮著舉足輕重的作用，是全身炎癥反應(yīng)時(shí)復(fù)雜病理變化的中心環(huán)節(jié)，同時(shí)也是多種因素、多層面、多環(huán)節(jié)協(xié)同作用的結(jié)果。為進(jìn)一步闡明上述協(xié)同作用的分子生物學(xué)機(jī)制，應(yīng)用RNA干擾技術(shù)將IκB蛋白激酶α及γ基因沉默，然后觀察RAW264.7小鼠巨噬細(xì)胞經(jīng)LPS刺激后，NF-κB的活化以及多種NF-κB依賴性的
[Abstract]:Objective:. To investigate the role of nuclear factor- 魏 B inflammatory signaling pathway in the excessive activation of alveolar macrophages during systemic inflammatory reaction, to observe the activity and nuclear translocation of NF- 魏 Bp65/p50, the changes of protein kinase IKK activity and the expression of inflammatory cytokines. The co-regulation effect of protein kinase IKK 偽 緯 on many parts of the above inflammatory signaling pathway and the molecular biological mechanism of Emodin therapy were also investigated. To further explore the dual function of nitric oxide (no) in the pathogenesis of acute lung injury in rats with severe acute pancreatitis and its close relationship with the change of NF- 魏 B activity in alveolar macrophages. To provide new experimental evidence for clinical treatment of various systemic inflammatory and reactive diseases such as acute lung injury and seeking more suitable drug targets. Methods:. Experiment 1: mouse RAW264.7 macrophage cell lines were studied by reverse transcription-PCR, Western blot, immunocytochemical staining, immunofluorescence staining and so on. To observe the effect of emodin on the inflammatory response induced by LPS, we divided into 7 groups, 5: 1: RAW264.7 macrophage normal culture group (n = 5: 1: RAW264.7) were stimulated by LPS(101 渭 g / ml, and they were divided into three groups at 30 min. On the basis of the stimulation of LPS(10 渭 g / ml, emodin(20g 渭 g / ml was added at the same time, and the samples were collected at different time points. Experiment two: the excessive inflammatory reaction mediated by NF- 魏 B plays an important role in the occurrence and development of many inflammatory diseases. It is the central link of complex pathological changes in systemic inflammatory reaction, and it is also a variety of factors and layers. In order to further elucidate the molecular mechanism of the synergistic effect, I 魏 B protein kinase 偽 and 緯 genes were silenced by RNA interference technique. Then, the activation of NF- 魏 B and various NF- 魏 B-dependent macrophages stimulated by LPS in RAW264.7 mice were observed.
【學(xué)位授予單位】：大連醫(yī)科大學(xué)
【學(xué)位級(jí)別】：博士
【學(xué)位授予年份】：2006
【分類(lèi)號(hào)】：R363;R285

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