本文選題：晚期氧化蛋白產(chǎn)物 + 人胚肺成纖維細(xì)胞�。� 參考：《遵義醫(yī)學(xué)院》2017年碩士論文

【摘要】：目的:研究晚期氧化蛋白產(chǎn)物(7)advanced oxidation protein products,AOPP(8)對人胚肺成纖維細(xì)胞(human embryonic lung fibroblasts MRC-5,MRC-5)活性氧自由基(7)reactive oxygen species,ROS(8)及細(xì)胞外基質(zhì)纖維連接蛋白(fibronectin,FN)及I型膠原(collagen I,Col I)的作用。方法:1.將MRC-5細(xì)胞分為人血清白蛋白(human serum albium,HSA)組,AO-PP時間效應(yīng)組(7)0、12、24、48及72小時(8)和濃度效應(yīng)組(7)0、50、100、200及400μg/ml(8),NADPH氧化酶抑制劑夾竹桃麻素(7)apocynin,APO(8)組(7)AOPP+APO(8)。2.用上述不同濃度的AOPP與MRC-5細(xì)胞共同培養(yǎng)不同的時間,MTT法檢測細(xì)胞的活力。3.采用ELISA法檢測FN、Col I及轉(zhuǎn)化生長因子(7)transforming growth factor-β1,TGF-β1(8)在細(xì)胞上清中的濃度,通過RT-PCR檢測各組細(xì)胞上述3個指標(biāo)m RNA的表達(dá)。4.根據(jù)MTT實驗結(jié)果選取AOPP作用最佳的時間點和濃度點刺激MRC-5細(xì)胞;流式細(xì)胞儀和熒光顯微鏡檢測活性氧自由基ROS的表達(dá)。5.NADPH氧化酶抑制劑APO與MRC-5細(xì)胞預(yù)孵育1小時,加入200μg/ml AOPP刺激48小時,觀察細(xì)胞內(nèi)ROS的變化,從蛋白和基因水平檢測FN、Col I及TGF-β1的表達(dá)。結(jié)果:1.不同濃度的AOPP(50、100、200、400μg/ml)作用于MRC-5細(xì)胞12、24、48及72小時,發(fā)現(xiàn)AOPP對MRC-5細(xì)胞具有顯著的增殖作用(P0.05(8),且在濃度為200μg/ml及時間48h達(dá)到峰值。2.FN、Col I及TGF-β1從蛋白分泌程度和基因表達(dá)水平均上調(diào)(P0.05(8),且在濃度200μg/ml及時間48h時達(dá)到峰值;未經(jīng)修飾的人血清白蛋白對上述相關(guān)纖維指標(biāo)的表達(dá)無明顯影響(P0.05)。3.MRC-5細(xì)胞與200μg/ml AOPP作用48小時,與對照組相比,細(xì)胞內(nèi)ROS水平顯著升高(P0.05)。4.MRC-5細(xì)胞與APO預(yù)孵育1小時,加入200μg/ml AOPP刺激48小時,細(xì)胞上清中FN、Col I及TGF-β1蛋白分泌減少且m RNA表達(dá)下調(diào)P0.05㖞,MRC-5細(xì)胞中ROS水平下降(P0.05),未經(jīng)修飾的人血清白蛋白對ROS的表達(dá)無明顯影響P0.05㖞。結(jié)論:AOPP能夠促進(jìn)體外MRC-5細(xì)胞的增殖,引起細(xì)胞外基質(zhì)FN、Col I表達(dá)上調(diào)及肺纖維化相關(guān)生長因子TGF-β1的表達(dá)增加,初步推測可能與NADPH氧化酶介導(dǎo)產(chǎn)生的ROS有關(guān)。
[Abstract]:Objective: to study the effects of advanced oxidation protein products of advanced oxidative protein (AOPP8) on human embryonic lung fibroblasts MRC-5 MRC-5 (reactive oxygen species-ROS8), extracellular matrix fibronectin (FNN) and collagen I (I) in human embryonic lung fibroblasts. Method 1: 1. MRC-5 cells were divided into two groups: human serum albumin (HSA) group (serum albium) group (AO-PP time effect group) and concentration response group (HSA group), and the concentration response group (HSA group) was divided into two groups: AO-PP group (n = 7) and concentration response group (n = 7) (n = 10), 7apocynin in AO-PP group (n = 8) and a concentration response group (n = 7), in which 7apocynin in AO-PP group (n = 8) and a concentration response group (n = 7) were divided into two groups. The activity of MRC-5 cells was detected by MTT assay with different concentrations of AOPP and MRC-5 cells at different time points. ELISA assay was used to detect the concentration of ELISA Col I and transforming growth factor- 尾 1 (TGF- 尾 1 + 8) in the cell supernatant. The expression of m RNA was detected by RT-PCR. According to the results of MTT experiment, the best time point and concentration point of AOPP were selected to stimulate MRC-5 cells, and the expression of free radical ROS was detected by flow cytometry and fluorescence microscope. 5. APO, inhibitor of NADPH oxidase, was preincubated with MRC-5 cells for 1 hour. After stimulation with 200 渭 g/ml AOPP for 48 hours, the changes of ROS were observed, and the expression of FNNCol I and TGF- 尾 1 were detected at the level of protein and gene. The result is 1: 1. Different concentrations of AOPP(50100200400 渭 g / ml were applied to MRC-5 cells for 48 and 72 hours. It was found that AOPP had a significant proliferative effect on MRC-5 cells, and reached a peak value of 200 渭 g/ml and 48h. 2. FNCol I and TGF- 尾 1 up-regulated P0.05Col I and TGF- 尾 1 from the level of protein secretion and gene expression, and reached the peak at the concentration of 200 渭 g/ml and 48h. There was no significant effect of unmodified human serum albumin on the expression of the above mentioned fiber markers. 3. MRC-5 cells were exposed to 200 渭 g/ml AOPP for 48 hours. Compared with the control group, the intracellular ROS level was significantly increased. 4. MRC-5 cells were preincubated with APO for 1 hour. After stimulation with 200 渭 g/ml AOPP for 48 h, the secretion of FNNCol I and TGF- 尾 1 protein in supernatant was decreased, and the expression of m RNA was down-regulated. The level of ROS in P0.05 g/ml AOPP MRC-5 cells was down-regulated. Unmodified human serum albumin had no significant effect on the expression of ROS. ConclusionOPP can promote the proliferation of MRC-5 cells in vitro, induce the up-regulation of extracellular matrix FNCol I expression and increase the expression of pulmonary fibrosis related growth factor TGF- 尾 1, which may be related to the ROS production mediated by NADPH oxidase.
【學(xué)位授予單位】：遵義醫(yī)學(xué)院
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2017
【分類號】：R563

【相似文獻(xiàn)】

相關(guān)期刊論文 前10條

1 武志強;A study to the fibroblast-populated collagen lattices[J];Chinese Journal of Traumatology;2000年03期

2 ;Effect of anti-fibrosis compound on collagen expression of hepatic cells in experimental liver fibrosis of rats[J];World Journal of Gastroenterology;2000年06期

3 ;Effect of Gangliosides on the Adhesion of Neuroblastom Cells to Collagen[J];中國當(dāng)代兒科雜志;2001年04期

4 凌翔,陳衛(wèi)民,劉勝洪,王罡;Expression of TGF-β in Region of Bone Defect Repaired by Collagen/Nano-beta-Tricalcium Phosphate Composite Artificial Bone[J];華中科技大學(xué)學(xué)報(醫(yī)學(xué)英德文版);2003年03期

5 趙英仁,朱風(fēng)群,和水祥,萇新明,喬文;PRIMARY STUDY ON MECHANISM OF COLLAGEN SYNTHESIS INHIBITION IN CULTURED HUMAN FETAL HEPATOCYTES PRETREATED BY SALVIA MILTIORRHIZA BUNGE[J];Academic Journal of Xi'an Jiaotong University;2003年02期

6 朱少華,朱金龍,肖堅,任亮,劉良,周亦武;Long-time Fulvic Acid Supplementation Modulates Hydroxylysyl Glycosylation of Collagen in Mice[J];華中科技大學(xué)學(xué)報(醫(yī)學(xué)英德文版);2004年05期

7 笪邦紅,曹陽,魏厚仁,陳志新,水迎波,李忠玉;Antagonistic Effects of Tranilast on Proliferation and Collagen Synthesis Induced by TGF-β_2 in Cultured Human Trabecular Meshwork Cells[J];華中科技大學(xué)學(xué)報(醫(yī)學(xué)英德文版);2004年05期

8 劉剛,胡蘊玉,趙建寧,吳蘇稼,熊卓,呂榮;Effect of type I collagen on the adhesion, proliferation, and osteoblastic gene expression of bone marrow-derived mesenchymal stem cells[J];Chinese Journal of Traumatology;2004年06期

9 劉新暉;Cellular compatibilty of the nano-hap/collagen and bone marrow stem cell[J];China Medical Abstracts;2005年03期

10 肖強兵;陳安民;郭風(fēng)勁;;Effects of Icariin on Expression of OPN mRNA and Type ⅠCollagen in Rat Osteoblasts in Vitro[J];華中科技大學(xué)學(xué)報(醫(yī)學(xué)英德文版);2005年06期

相關(guān)會議論文 前10條

1 沈家弘;蕭哲志;許準(zhǔn)榕;;Caffeic acid phenethyl ester——一種新的collagen拮抗劑抑制血小板凝集反應(yīng)的機轉(zhuǎn)探討[A];第六屆海峽兩岸心血管科學(xué)研討會論文摘要集[C];2007年

2 朱杰;郭連紅;王國棟;;Binding Force between Anti-collagen I Antibody and Collagen Fiber Observed with Atomic Force Microscopy[A];第一屆全國生物物理化學(xué)會議暨生物物理化學(xué)發(fā)展戰(zhàn)略研討會論文摘要集[C];2010年

3 ;Nutritional effect of marine collagen supplementation in postoperative patients[A];中國食品科學(xué)技術(shù)學(xué)會第八屆年會暨第六屆東西方食品業(yè)高層論壇論文摘要集[C];2011年

4 ;Isolation and Characterization of Pepsin-Soluble Collagen from the Skin of Peru Squid(Dosidicus Gigas)[A];漁業(yè)科技創(chuàng)新與發(fā)展方式轉(zhuǎn)變——2011年中國水產(chǎn)學(xué)會學(xué)術(shù)年會論文摘要集[C];2011年

5 Dilare Adi;Yitong Ma;Xiang Xie;Zhenyan Fu;Fen Liu;Bangdang Chen;;A novel polymorphism of the COL4A1 gene is associated with coronary artery disease in Uygur population in China[A];中華醫(yī)學(xué)會第十五次全國心血管病學(xué)大會論文匯編[C];2013年

6 Dong Shiwu;Guo Hongfeng;Kang Fei;Yang Bo;Yan Yanfei;Miao Ying;;rFN/Cad-11-modified collagen typeⅡbiomimetic interface promotes the adhesion and chondrogenic differentiation of mesenchymal stem cells[A];中國解剖學(xué)會2013年年會論文文摘匯編[C];2013年

7 LIU Yang;REN Li;WANG Yingjun;;A Novel Antibacterial Collagen Scaffold Using for Corneal Tissue Engineering[A];2013廣東材料發(fā)展論壇——戰(zhàn)略性新興產(chǎn)業(yè)發(fā)展與新材料科技創(chuàng)新研討會論文摘要集[C];2013年

8 Hiroyasu Uzui;;Profibrotic influence of high glucose on human cardiac fibroblast functions[A];第十三次全國心血管病學(xué)術(shù)會議論文集[C];2011年

9 ;Studies of therapeutic effect and mechanism of total saponin from D.nipponica Mak in CIA rat induced by typeⅡcollagen[A];中國藥理學(xué)會第十次全國學(xué)術(shù)會議�？痆C];2009年

10 ;Influence of TMP on Adhesion Formation during Tendon Healing[A];第五屆全國青年體育科學(xué)學(xué)術(shù)會議、第二屆中國體育博士高層論壇論文集[C];2008年

相關(guān)博士學(xué)位論文 前2條

1 Mohammad Hashemipour;[D];北京中醫(yī)藥大學(xué);2010年

2 徐珍娥;炎癥加重脂質(zhì)誘導(dǎo)的腎臟損傷及阿托伐他汀干預(yù)作用的實驗研究[D];重慶醫(yī)科大學(xué);2010年

相關(guān)碩士學(xué)位論文 前10條

1 平櫓;芡實對糖尿病腎病大鼠腎臟組織MMP-9、TIMP-1及Collagen Ⅳ表達(dá)的影響[D];山西醫(yī)科大學(xué);2015年

2 田原;越橘花青素對人鞏膜成纖維細(xì)胞表達(dá)MMP-2、CollagenⅠ蛋白的影響[D];暨南大學(xué);2016年

3 卜召鋒;依達(dá)拉奉對UUO大鼠模型MDA、p-ERK1/2、NF-κB p65及Collagen Ⅲ表達(dá)的影響[D];河北醫(yī)科大學(xué);2016年

4 寧志龍;利用分子動力學(xué)模擬研究兩復(fù)合物系統(tǒng)：VWF-A3與Collagen、VWF-A1與VWF-A3[D];華南理工大學(xué);2016年

5 張幫艷;AOPP對MRC-5細(xì)胞增殖及FN、Collagen I表達(dá)的影響[D];遵義醫(yī)學(xué)院;2017年

6 蘇sセ，

本文編號：1819605