【摘要】：E-選擇素是表達(dá)在內(nèi)皮細(xì)胞表面的一種重要的粘附分子。在組織受到損傷或有炎癥反應(yīng)時，它的表達(dá)上調(diào)。升高的E-選擇素水平對血流中的白細(xì)胞有強(qiáng)烈的吸引作用，聚集到損傷或炎癥部位的白細(xì)胞釋放各種氧自由基以及炎癥因子，對入侵的病原體起到了很好的殺傷作用。但是異常表達(dá)的E-選擇素以及過度聚集和激活的白細(xì)胞往往也會對正常組織產(chǎn)生嚴(yán)重?fù)p傷。比如由LPS引起的急性肺損傷的發(fā)病就涉及這個過程。在本工作中我們提出假說——如果可以阻斷E-選擇素這種重要的免疫粘附分子，可以對急性肺損傷起到顯著的保護(hù)作用。 我們使用LPS作為炎癥刺激劑在小鼠上復(fù)制急性肺損傷的模型，實驗對象是E-選擇素基因敲除鼠以及野生型對照小鼠。E-選擇素基因敲除小鼠在造模后的狀態(tài)明顯好于野生型對照組。12小時后，用紅外照相機(jī)記錄各組小鼠的皮膚表面溫度。隨后，，取小鼠血樣，肺組織樣本，肺泡灌洗液等樣本做進(jìn)一步檢測：E-選擇素和髓過氧化物酶(MPO)蛋白在肺組織中的表達(dá)，肺組織丙二醛(MDA)含量，血清炎癥因子水平，肺組織干濕比(wet/dry),病理學(xué)染色等等。結(jié)果發(fā)現(xiàn)LPS造模后，E-選擇素基因敲除小鼠的血清炎癥因子TNF-α、IL-1β明顯低于野生型對照小鼠，肺泡灌洗液里的白細(xì)胞聚集也明顯較少，MPO蛋白的表達(dá)更少，MDA含量更低，體溫更接近正常。另外生存率實驗表明，LPS作用后的E-選擇素基因敲除小鼠的生存率明顯優(yōu)于野生型小鼠的造模對照。 隨后我們希望可以通過體外篩選可以找到有效的E-選擇素抑制劑。LPS刺激人臍靜脈內(nèi)皮細(xì)胞(HUVEC)可以明顯上調(diào)E-選擇素的表達(dá)，但是100uM的西咪替丁在不影響細(xì)胞活力的情況下，對E選擇素的表達(dá)有顯著的抑制作用。 進(jìn)一步的實驗發(fā)現(xiàn)，小鼠口服西咪替丁(200mg/kg)，可以明顯改善LPS誘導(dǎo)的急性肺損傷。在造模后的12小時，紅外成像系統(tǒng)表明小鼠的體溫更接近正常，肺水腫(干濕比)指數(shù)明顯低于模型對照組，血清炎癥因子TNF-α更低，生存率也明顯延長。 我們得出結(jié)論：阻斷E選擇素可以緩解LPS誘導(dǎo)的急性肺損傷，西咪替丁的保護(hù)作用很可能與它抑制E-選擇素的效應(yīng)有關(guān)。
[Abstract]:E-selectin is an important adhesion molecule expressed on endothelial cell surface. Its expression is up-regulated when the tissue is damaged or has an inflammatory response. The elevated level of E- selectin has a strong attraction to the white blood cells in the blood stream. The white blood cells gathered in the injured or inflammatory sites release various oxygen free radicals and inflammatory factors, which can kill the invading pathogens. However, abnormal expression of E-selectin and excessive aggregation and activation of white blood cells can also cause severe damage to normal tissues. For example, acute lung injury caused by LPS is involved in this process. In this work, we hypothesized that if E- selectin, an important immune adhesion molecule, could be blocked, it could play a significant protective role in acute lung injury. We used LPS as an inflammatory stimulant to model acute lung injury in mice. Eselectin gene knockout mice and wild type control mice were studied. The status of Eselectin gene knockout mice was significantly better than that of wild type control mice after 12 hours. The skin surface temperature of each group was recorded by infrared camera. Then, the blood samples, lung tissue samples and alveolar lavage fluid samples of mice were taken for further detection: the expression of E- selectin and myeloperoxidase (MPO) protein in lung tissue, the content of malondialdehyde (MDA) in lung tissue, the level of serum inflammatory factor, and so on. Lung tissue dry-wet ratio (wet/dry), pathological staining and so on. The results showed that the levels of serum inflammatory cytokines TNF- 偽 and IL-1 尾 in E- selectin gene knockout mice were significantly lower than those in wild type control mice, and the leukocyte aggregation in alveolar lavage fluid was significantly lower, and the expression of MPO protein was less in E- selectin gene knockout mice. MDA levels are lower and body temperature is closer to normal. In addition, the survival rate of E- selectin gene knockout mice treated with LPS was significantly better than that of wild-type mice. Then we hope that we can find an effective inhibitor of E- selectin through in vitro screening. (HUVEC) stimulated by LPS can significantly up-regulate the expression of E- selectin in human umbilical vein endothelial cells. But cimetidine of 100uM inhibited the expression of E-selectin without affecting cell viability. Further experiments showed that cimetidine (200mg/kg) could significantly improve acute lung injury induced by LPS in mice. After 12 hours, the infrared imaging system showed that the body temperature of the mice was closer to normal, the index of pulmonary edema (dry-wet ratio) was significantly lower than that of the model control group, the serum inflammatory factor TNF- 偽 was lower, and the survival rate was longer. We conclude that blocking E-selectin can alleviate acute lung injury induced by LPS and the protective effect of cimetidine may be related to its inhibitory effect on Eselectin.
【學(xué)位授予單位】：第二軍醫(yī)大學(xué)
【學(xué)位級別】：博士
【學(xué)位授予年份】：2012
【分類號】：R563.8

【共引文獻(xiàn)】

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1 楊小紅 ,宋明英 ,羅軍敏;創(chuàng)傷患者早期血清可溶性E-選擇素水平及其臨床意義[J];貴州醫(yī)藥;2002年08期

相關(guān)博士學(xué)位論文 前2條

1 陳秀凱;感染性休克致急性腎損傷的血流動力學(xué)及機(jī)制研究[D];北京協(xié)和醫(yī)學(xué)院;2011年

2 張銀中;細(xì)胞因子及異丙酚對中性粒細(xì)胞凋亡的影響[D];中國協(xié)和醫(yī)科大學(xué);2003年

相關(guān)碩士學(xué)位論文 前3條

1 陳志明;大鼠皮膚切創(chuàng)愈合過程中細(xì)胞間粘附分子及P選擇素表達(dá)的研究[D];中國醫(yī)科大學(xué);2003年

2 姚嶺松;E-選擇素在新生大鼠內(nèi)毒素誘導(dǎo)肺損傷的表達(dá)及地塞米松對其表達(dá)的影響[D];蘇州大學(xué);2007年

3 陳宏貞;按摩對骨骼肌損傷修復(fù)過程中bFGF表達(dá)的影響[D];武漢體育學(xué)院;2009年

本文編號：2404692