小鼠肺爆震伤蛋白质组学研究

来源 :中华急诊医学杂志 | 被引量 : 0次 | 上传用户:xujuenrong
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目的:通过检测不同时间点肺爆震伤小鼠肺蛋白质组学变化,探究肺爆震伤损伤机制。方法:60只健康雄性C57BL/6小鼠,随机(随机数字法)分为对照组、爆震后12 h组、24 h组、48 h组、72 h组及1周组(每组10只)。在北部战区总医院动物实验室进行实验,利用自主研发的精准爆震装置建立小鼠肺爆震伤模型;通过大体观察和HE染色评价肺组织损伤情况;基于液相色谱-串联质谱联用(liquid chromatography-tandem mass spectrometry,LC-MS/MS)的蛋白质组学技术对小鼠肺组织蛋白进行定量分析,筛选出差异表达蛋白,在此基础上利用生物信息学工具分析蛋白质组学变化。结果:肺爆震伤后,小鼠肺组织表面可观察到散在出血点,且随时间延长逐渐增多,24 h出血最为严重。HE染色可见爆震后12 h和24 h的肺泡正常组织结构消失,肺泡腔内弥漫性出血,大量炎细胞浸润,肺间质渗出液增多,肺泡壁增厚,肺泡腔萎陷融合。LC-MS/MS共鉴定到6 861个蛋白质,定量到608个差异表达蛋白,其中在12、24、48、72 h及1周分别有227个、140个、202个、258个和71个差异蛋白。基因本体论(gene ontology, GO)分析得到包括细胞粘附、细胞外基质组织及胶原原纤维组织等130个生物进程子类,外泌体、细胞外基质及细胞质等66个细胞组分子类,以及细胞外基质结构组成、肌动蛋白结合及抗氧化活性等43个分子功能子类。KEGG分析共得到细胞外基质受体相互作用、黏着斑及PI3K-Akt信号通路等24个子类。结论:小鼠肺爆震后早期不同时间点差异表达蛋白组合亦不同,损伤机制复杂,特别是12~24 h肺损伤最为严重,炎症反应显著。“,”Objective:To establish the lung blast injury model in mice, detect the proteomic changes of lung in mice at different time points, and explore the mechanism of lung blast injury.Methods:A total of 60 healthy male C57BL/6 mice were randomly (random number) divided into the control group, 12-h group after thorax blast, 24-h group, 48-h group, 72-h group and 1-week group (n n=10 each group). Experiments were carried out in the animal laboratory of the General Hospital of the Northern Theater Command. The model of lung blast injury in mice was established by using a self-developed precision blast device, and the lung tissue injury situation was evaluated by gross observation and HE staining. The proteins in mouse lung tissue were quantitatively analyzed based on LC-MS/MS proteomic technology, and the differentially expressed proteins were screened. On this basis, bioinformatics tool was used to analyze proteomic changes.n Results:After lung blast injury, scattered bleeding spots could be observed on the surface of lung tissue of mice, and the bleeding points were gradually increased with time, showing a patchy distribution, and the symptoms were the most severe at 24 h. The results of HE staining showed that the normal tissue structure of alveoli disappeared at 12 and 24 h under light microscopy with diffuse bleeding in the alveolar cavity, infiltration of a large number of inflammatory cells, increased interstitial exudate, thickened alveolar wall, and collapsed and merged alveolar cavity. A total of 6 861 proteins were identified by LC-MS/MS in lung tissue samples of mice after thorax blast, and 608 differentially expressed proteins were quantified, of which 227, 140, 202, 258 and 71 differential proteins were at 12 h, 24 h, 48 h, 72 h, and 1 week, respectively. According to GO analysis, 130 biological process subtypes including cell adhesion, extracellular matrix tissue and collagen fibril tissue were obtained. Besides, 66 cellular component involving extracellular exosomes, extracellular matrix and cytoplasm were obtained. And 43 molecular functional subclasses such as extracellular matrix structure composition, actin binding and antioxidant activity were obtained. KEGG analysis yielded 24 pathways including ECM-receptor interactions, focal adhesions and PI3K-Akt signaling pathway across the endothelium.Conclusions:Differentially expressed protein combinations are also different at different time points in the early stage after lung blast in mice, and the injury mechanism is complicated. The lung blast injury is the most serious at 12-24 h after blast and produces significant inflammatory response.
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