CAJ | 학술논문

目的利用蛋白质组学方法筛选铜绿假单胞菌急性肺损伤特异相关蛋白质,为鉴别诊断感染与非感染因素引起的急性肺损伤提供理论依据.方法建立感染(铜绿假单胞菌急性肺损伤)与非感染(包括机械通气急性肺损伤、油酸急性肺损伤)大鼠模型,应用双向凝胶电泳技术分离总蛋白,Image Master双向凝胶电泳软件分析差异表达蛋白质点后,应用基质辅助激光解析电离飞行时间质谱获取肽质量指纹图谱并通过NCBI数据库鉴定差异表达的蛋白质,最后Western blot验证差异表达的蛋白质.结果双向凝胶电泳图谱中找到32个表达量有明显差异的蛋白质点,质谱鉴定出18种蛋白质.其中的过氧化物氧化还原酶Ⅰ(Prx Ⅰ)、钙网蛋白等11种蛋白质在铜绿假单胞菌急性肺损伤组表达上调,并在Western blot验证了PrxⅠ的上调,与凝胶电泳结果一致.谷胱甘肽-S-转移酶α4、甲状腺素蛋白等7种蛋白在铜绿假单胞菌急性肺损伤组表达下调.结论应用比较蛋白组学鉴定出18种差异蛋白,11种在铜绿假单胞菌急性肺损伤组表达上调,7种表达下调,分别参与催化代谢、氧化还原、信号转导等调节.其中PrxⅠ和钙网蛋白可能通过对氧化/抗氧化和免疫应答的调节在细菌感染性急性肺损伤中起着重要作用.本研究为进一步阐明感染相关性急性肺损伤的发病机制提供新的线索.
목적 이용단백질조학방법사선동록가단포균급성폐손상특이상관단백질,위감별진단감염여비감염인소인기적급성폐손상제공이론의거.방법 건립감염(동록가단포균급성폐손상)여비감염(포괄궤계통기급성폐손상、유산급성폐손상)대서모형,응용쌍향응효전영기술분리총단백,Image Master쌍향응효전영연건분석차이표체단백질점후,응용기질보조격광해석전리비행시간질보획취태질량지문도보병통과NCBI수거고감정차이표체적단백질,최후Western blot험증차이표체적단백질.결과 쌍향응효전영도보중조도32개표체량유명현차이적단백질점,질보감정출18충단백질.기중적과양화물양화환원매Ⅰ(Prx Ⅰ)、개망단백등11충단백질재동록가단포균급성폐손상조표체상조,병재Western blot험증료PrxⅠ적상조,여응효전영결과일치.곡광감태-S-전이매α4、갑상선소단백등7충단백재동록가단포균급성폐손상조표체하조.결론 응용비교단백조학감정출18충차이단백,11충재동록가단포균급성폐손상조표체상조,7충표체하조,분별삼여최화대사、양화환원、신호전도등조절.기중PrxⅠ화개망단백가능통과대양화/항양화화면역응답적조절재세균감염성급성폐손상중기착중요작용.본연구위진일보천명감염상관성급성폐손상적발병궤제제공신적선색.
Objective To screen the differential expressed proteins of Pseudomonas aeruginosainduced acute lung injury by proteomics, and to provide the theoretical basis for the identification of acute lung injury caused by bacterial infection and non-infection. Methods The rat models of Pseudomonas aeruginosa -induced acute lung injury and non-infection-induced acute lung injury (including ventilatorinduced acute lung injury and oleic acid-induced acute lung injury) were established. The total proteins were extracted and separated by two-dimensional gel electrophoresis (2-DE). The sites of differential expression were analyzed using Image Master 2D Elite 5. 0 image analysis software,and the peptide mass finger-printing was identified by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS). The biological information of these proteins was searched in the NCBI database and confirmed by Western blot. Results Thirty-two differential expression protein spots were found in 2-DE maps, and eighteen proteins were analyzed and identified by MALDI-TOF-MS. Eleven proteins (including peroxiredoxin Ⅰ . calreticulin etc. ) were upregulated while seven proteins (including glutathione-S-transferaseα4. transthyretin etc. ) were downregulated in Pseudomonas aeruginosa -induced acute lung injury group. The upregulated expression of peroxiredoxin Ⅰ in Pseudomonas aeruginosainduced acute lung injury group was confirmed by Western blot. Conclusions Eighteen different proteins were identified by comparative proteomics, and involved in catalytic metabolism, redox, signal transduction. Eleven proteins were upregulated and seven proteins were downregulated in Pseudomonas aeruginosa-induced acute lung injury group. Peroxiredoxin Ⅰ and calreticulin may play an important role in bacterial infected-acute lung injury by regulating oxidant/antioxidant and immune response.