本文選題：肺纖維化 + 博萊霉素�。� 參考：《吉林大學》2017年博士論文

【摘要】：肺纖維化在病理上表現為I型肺泡上皮細胞持續(xù)性受損、大量炎性細胞在肺組織內浸潤、成纖維細胞增生和分泌細胞外基質,過多分泌的細胞外基質沉積在肺間隔內,最終出現正常肺組織結構異常改變。研究者目前認為失控的氧化應激,氧化/抗氧化平衡失調是誘發(fā)肺組織出現纖維化的分子學基礎,參與肺纖維化發(fā)生、發(fā)展的各個階段。因此,通過上調內源性抗氧化通路可能在治療肺纖維化中起到一定作用。核因子Nrf2能夠通過轉錄活化其下游多種抗氧化劑的表達,增加體內抗氧化劑水平和活性。許多研究表明Nrf2在肺纖維化的發(fā)病中發(fā)揮作用,敲除Nrf2基因可增加博萊霉素誘導肺纖維化的易感性。大量實驗發(fā)現蘿卜硫素(SFN),一種Nrf2激動劑,在多種疾病模型中發(fā)揮間接抗氧化、抗毒性作用。本研究通過體內和體外實驗,目的是探討蘿卜硫素通過Nrf2活化發(fā)揮對博萊霉素所致小鼠肺纖維化的保護作用。為探討SFN對博萊霉素誘導氧化應激所致肺纖維化的作用機制,本實驗采用一次性氣管內注射博萊霉素,建立小鼠肺纖維化體內模型,并于造模后每隔一天給予皮下注射SFN干預處理。分別于第7、28天處死小鼠,收集肺組織標本。應用病理學檢查進行HE染色、Masson染色、TUNEL和免疫組化染色,用于觀察肺組織炎癥及纖維化程度,細胞凋亡和Nrf2的表達情況;利用Western blot法測定促炎癥因子、促凋亡因子、促纖維化因子和氧化物3NT、4HNE的表達水平;使用羥脯氨酸測定試劑盒檢測肺組織中羥脯氨酸含量;分別采用實時定量PCR和Western blot法檢測Nrf2及其下游HO-1、NQO1、SOD1、CAT基因和蛋白水平。此外,我們還在體外培養(yǎng)人肺成纖維細胞,通過血管緊張素II(Ang II)模擬體內氧化應激,通過給予SFN預處理,觀察SFN對Ang II誘導人肺成纖維細胞TGF-β分泌和ROS產生的影響;并應用Nrf2特異性si RNA沉默人肺成纖維細胞的Nrf2基因,從而探討Nrf2在SFN減輕氧化應激所致肺纖維化過程中的作用。SFN干預博萊霉素誘導肺纖維化體內實驗結果顯示:給予SFN后,7、28天時肺泡炎癥改變明顯減輕;7天時因博萊霉素所致肺泡上皮細胞的凋亡減少;28天時SFN干預組肺纖維化程度、評分和羥脯氨酸膠原含量明顯減輕。給予SFN處理,還能降低促炎因子IL-1β、TNF-α、促凋亡因子caspase-3和促纖維化因子TGF-β的表達,減少肺組織氧化標志物3NT、4HNE含量,同時伴隨著Nrf2及其下游基因表達增強。通過進行體外實驗,觀察到SFN能減少因Ang II所致人肺成纖維細胞TGF-β分泌和ROS產生,且發(fā)現這一保護作用是通過上調Nrf2及HO-1表達所致。利用特異性si RNA沉默Nrf2基因可以阻斷SFN保護Ang II誘導成纖維細胞分泌TGF-β的作用。結合以上實驗結果,我們發(fā)現SFN通過活化Nrf2,增加抗氧化酶活性,減輕博萊霉素所致小鼠肺纖維。該研究為SFN用于預防和治療因氧化應激所致肺部炎癥和肺纖維化提供理論研究基礎。
[Abstract]:The pathological manifestations of pulmonary fibrosis are persistent damage of type I alveolar epithelial cells, infiltration of a large number of inflammatory cells in lung tissue, proliferation and secretion of extracellular matrix of fibroblasts, and excessive secretion of extracellular matrix in the interpulmonary septum. Finally, abnormal changes of normal lung tissue structure appeared. Researchers believe that out of control oxidative stress, oxidative / antioxidant imbalance is the molecular basis of pulmonary fibrosis, involved in the occurrence and development of various stages of pulmonary fibrosis. Therefore, up-regulation of endogenous antioxidant pathway may play a role in the treatment of pulmonary fibrosis. Nuclear factor Nrf2 can activate the expression of many antioxidants downstream through transcription and increase the antioxidant level and activity in vivo. Many studies have shown that Nrf2 plays a role in the pathogenesis of pulmonary fibrosis. Knockout of Nrf2 gene can increase the susceptibility of bleomycin induced pulmonary fibrosis. A large number of experiments have found that sulforaphane, a Nrf2 agonist, plays an indirect antioxidant and antitoxic effect in various disease models. The aim of this study was to investigate the protective effect of sulforaphane on bleomycin induced pulmonary fibrosis in mice by Nrf2 activation in vitro and in vivo. In order to investigate the mechanism of SFN on bleomycin induced pulmonary fibrosis induced by oxidative stress, a mouse model of pulmonary fibrosis was established by single intratracheal injection of bleomycin. The model was treated by subcutaneous injection of SFN every other day. The mice were killed on the 7th day and the lung tissues were collected. In order to observe the degree of inflammation and fibrosis, the expression of apoptosis and Nrf2 in lung tissue, the expression of pro-inflammatory factors and pro-apoptotic factors were determined by Western blot method, and the immunohistochemical staining and Tunel staining with HE staining were used to observe the degree of inflammation and fibrosis, the expression of apoptosis and the expression of Nrf2 in lung tissue. The expression levels of fibrogenic factor and oxide 3NTO4HNE, hydroxyproline assay kit were used to detect the content of hydroxyproline in lung tissue, and Nrf2 and its downstream HO-1NQO1SOD1 cat gene and protein were detected by real-time quantitative PCR and Western blot respectively. In addition, we cultured human lung fibroblasts in vitro, simulated oxidative stress by angiotensin II (II(Ang II) in vivo, and observed the effect of SFN on TGF- 尾 secretion and ROS production induced by Ang II by SFN pretreatment. The Nrf2 gene of human lung fibroblasts was silenced by Nrf2 specific si RNA. To explore the role of Nrf2 in alleviating oxidative stress induced pulmonary fibrosis induced by SFN in vivo, the results showed that the alveolar inflammation was significantly reduced at 728 days after SFN administration. Apoptosis of alveolar epithelial cells induced by mycin was reduced at 28 days after treatment with SFN. Scores and hydroxyproline collagen content were significantly reduced. SFN treatment could also reduce the expression of IL-1 尾 -TNF- 偽, caspase-3 and TGF- 尾, and reduce the content of 3NTN ~ 4HNE, which was accompanied by the increased expression of Nrf2 and its downstream genes. Through in vitro experiments, it was observed that SFN could reduce the secretion of TGF- 尾 and the production of ROS in human lung fibroblasts induced by Ang II, and the protective effect was caused by up-regulating the expression of Nrf2 and HO-1. Silencing Nrf2 gene by specific si RNA can block the protective effect of SFN on the secretion of TGF- 尾 in fibroblasts induced by Ang II. Combined with the above results, we found that SFN can increase the activity of antioxidant enzymes by activating Nrf2, and reduce the lung fiber induced by bleomycin in mice. This study provides a theoretical basis for the use of SFN in the prevention and treatment of pulmonary inflammation and pulmonary fibrosis due to oxidative stress.
【學位授予單位】：吉林大學
【學位級別】：博士
【學位授予年份】：2017
【分類號】：R563

【相似文獻】

相關期刊論文 前10條

1 陳曉玲,黃善生,凌亦凌,李文斌,王新良;誘導型一氧化氮合酶活性變化對肺纖維化形成的影響[J];中國病理生理雜志;2000年10期

2 康衛(wèi)國,劉久山;肺纖維化的診斷[J];臨床薈萃;2001年14期

3 李學軍;肺纖維化治療研究進展[J];中國綜合臨床;2001年10期

4 王興勝;肺纖維化治療的研究進展[J];國外醫(yī)學(內科學分冊);2001年09期

5 劉慶華;轉化生長因子-β與肺纖維化[J];國外醫(yī)學(內科學分冊);2002年08期

6 孟瑩,李旭,蔡紹曦;局部組織腎素—血管緊張素系統(tǒng)在肺纖維化中的作用[J];實用醫(yī)學雜志;2002年12期

7 周賢梅,戴令娟,蔡后榮,侯杰;慢性阻塞性肺疾病合并肺纖維化臨床分析[J];臨床薈萃;2003年22期

8 耿開興;肺纖維化60例臨床分析[J];中原醫(yī)刊;2003年12期

9 何平平;張平;;局部腎素-血管緊張素-醛固酮系統(tǒng)與肺纖維化[J];醫(yī)學綜述;2007年09期

10 張永勝;馮一中;顧振綸;;藥物防治肺纖維化的研究進展[J];中國血液流變學雜志;2007年01期

相關會議論文 前10條

1 賈英杰;李小江;孫一予;陳軍;張瑩;黃敏娜;包芳芳;;肺癌放化療導致肺纖維化中醫(yī)藥干預的研究現狀[A];2009年首屆全國中西醫(yī)腫瘤博士及中青年醫(yī)師論壇論文集[C];2009年

2 張紓難;孫瑞華;;建立肺纖維化生存質量評價體系方法初探[A];第五次全國中西醫(yī)結合呼吸病學術交流大會論文匯編[C];2000年

3 黃山英;宋良文;張勇;孫麗;尹紀業(yè);;Ⅳ型膠原和基質金屬蛋白酶在大鼠早期肺纖維化啟動中的作用機制[A];第六屆全國生物醫(yī)學體視學學術會議暨第九屆全軍軍事病理學學術會議、第五屆全軍定量病理學學術會議論文匯編[C];2005年

4 畢黎琦;孫文鑄;郭嘉隆;杜娟;周鳳嬌;;胰島素樣生長因子-Ⅰ和轉化生長因子-β1在肺纖維化形成中的作用研究[A];第十二屆全國風濕病學學術會議論文集[C];2007年

5 王星;李世彬;王詠梅;吳琦;;酒精與大鼠肺纖維化關系的研究[A];中華醫(yī)學會呼吸病學年會——2011（第十二次全國呼吸病學學術會議）論文匯編[C];2011年

6 楊乃女;嚴賴莎;陳少賢;;氟伐他汀對大鼠肺纖維化干預作用的研究[A];中華醫(yī)學會呼吸病學年會——2011（第十二次全國呼吸病學學術會議）論文匯編[C];2011年

7 劉建秋;黃利華;李竹英;;化纖湯防治肺纖維化的實驗研究[A];第七次全國中西醫(yī)結合呼吸病學術交流大會論文匯編（一）[C];2004年

8 楊s，

本文編號：1826877