本文選題：TRB3 + MAPK信號(hào)通路　； 參考：《青島大學(xué)》2017年碩士論文

【摘要】：目的1.探討TRB3在C57BL/6小鼠纖維化中的作用。2.外源性調(diào)控TRB3對(duì)博來(lái)霉素誘導(dǎo)的C57BL/6小鼠肺間質(zhì)纖維化的影響,進(jìn)一步在MAPK信號(hào)通路中探討三者之間的關(guān)系。方法1.實(shí)驗(yàn)分組:(1)單純對(duì)照組+生理鹽水(Mock組,對(duì)照組)(2)纖維化組+生理鹽水(fibrosis control組,F組)(3)纖維化+GFP組(F+GFP組)(4)纖維化+Ad-TRB3過(guò)表達(dá)組(F+TRB3組)(5)纖維化+Ad-TRB3-sh RNA干擾組(F+sh-TRB3組)。2.動(dòng)物造模:博來(lái)霉素3.5mg/kg氣管注入,造模次日,進(jìn)行腺病毒尾靜脈注射,分別于第7、14和28天處死實(shí)驗(yàn)小鼠一批,收集BALF、肺組織標(biāo)本供實(shí)驗(yàn)用。3.肺組織HE/Masson染色形態(tài)學(xué)觀察及纖維化評(píng)分,進(jìn)行羥脯氨酸測(cè)定進(jìn)一步明確膠原蛋白含量。ELASIA檢測(cè)肺組織中CollaⅠ/Ⅲ的表達(dá)。免疫組化明確間質(zhì)及上皮相關(guān)蛋白的表達(dá)。對(duì)肺泡灌洗液(BALF)進(jìn)行細(xì)胞分類及計(jì)數(shù)。4.用Western blot方法從蛋白水平檢測(cè)C57BL/6小鼠TRB3、p-ERK1/2、ERK1/2、p-p38MAPK、p38MAPK及EMT相關(guān)蛋白(如E-cadherin、Fibronectin、Vimentin、α-SMA)的表達(dá)情況;用RT-q PCR從m RNA水平檢測(cè)TRB3、pro CollaⅠ/Ⅲ的表達(dá)。結(jié)果1.成功地構(gòu)建出博來(lái)霉素誘導(dǎo)的小鼠肺纖維化模型。2.肺組織HE/Masson染色形態(tài)學(xué)觀察及纖維化評(píng)分,肺組織進(jìn)行羥脯氨酸測(cè),結(jié)果顯示:TRB3過(guò)表達(dá)纖維化程度更重,TRB3表達(dá)下調(diào)纖維化程度明顯減輕。BALF中,TRB3過(guò)表達(dá)表現(xiàn)出細(xì)胞總數(shù)、中性粒細(xì)胞數(shù)明顯增多,巨噬細(xì)胞減少。TRB3表達(dá)下調(diào)組結(jié)果相反。3.Western blot檢測(cè)各組肺組織TRB3、p-ERK1/2、ERK1/2、p-p38MAPK、p38MAPK及EMT相關(guān)蛋白(如E-cadherin、Fibronectin、Vimentin、α-SMA)的表達(dá),RT-q PCR檢測(cè)TRB3、pro CollaⅠ/Ⅲm RNA的表達(dá),結(jié)果顯示:TRB3過(guò)表達(dá)組的p-ERK1/2、ERK1/2、p-p38MAPK、p38MAPK及EMT相關(guān)蛋白及pro CollaⅠ/Ⅲm RNA的表達(dá)上調(diào);而TRB3表達(dá)下調(diào)組結(jié)果相反。結(jié)論1.TRB3過(guò)表達(dá)可增加EMT相關(guān)蛋白及m RNA的表達(dá),TRB3基因表達(dá)下調(diào)后,結(jié)果相反。提示TRB3可能對(duì)EMT進(jìn)行調(diào)控,進(jìn)一步參與肺纖維化的形成。2.在博來(lái)霉素誘導(dǎo)的小鼠肺纖維化可見(jiàn)TRB3及MAPK信號(hào)通路關(guān)鍵蛋白表達(dá)上調(diào),提示TRB3可能通過(guò)MAPK信號(hào)通路參與纖維化過(guò)程。TRB3過(guò)表達(dá)可以加重博來(lái)霉素誘導(dǎo)的肺纖維化,TRB3下調(diào)則可減輕這一纖維化過(guò)程,即通過(guò)外源性調(diào)控TRB3可能會(huì)對(duì)博來(lái)霉素誘導(dǎo)的小鼠肺纖維化產(chǎn)生影響。
[Abstract]:Objective 1.To investigate the role of TRB3 in fibrosis of C57BL/6 mice.Exogenous regulation of TRB3 on pulmonary interstitial fibrosis induced by bleomycin in C57BL/6 mice was conducted to explore the relationship between them in MAPK signaling pathway.Method 1.Animal model: bleomycin (3.5mg/kg) was injected into trachea. The next day, adenovirus was injected into tail vein. The mice were killed on day 714 and 28, respectively.Lung tissue HE/Masson staining morphology and fibrosis score, hydroxyproline determination of collagen content. ELASIA detection of lung tissue Colla 鈪，

本文編號(hào)：1735999