基于蛋白组学技术探究危重烧伤患者血小板外囊泡凝血反应的研究
Coagulation reaction of platelet extracellular vesicles in critically burned patients based on proteomics technology
高雷 1李全 1巴特 1曹胜军 1李芳 2侯智慧 1王凌峰2
作者信息
- 1. 内蒙古医科大学第三附属医院烧伤外科,包头 014010
- 2. 内蒙古烧伤医学研究所,包头 014010
- 折叠
摘要
目的 应用蛋白组学技术探讨危重烧伤患者与健康体检者血小板外囊泡(PLT-EVs)中参与凝血反应的差异蛋白.方法 选取内蒙古医科大学第三附属医院健康体检者和重度烧伤患者各6例,抽取静脉血,高速离心法获取PLT-EVs,通过透射电镜、纳米颗粒分析和蛋白质印迹法(Western blot)鉴定.通过蛋白组学技术分析并定量两组PLT-EVs蛋白质,以差异倍数=1.5(上调或下调)筛选差异蛋白,将差异表达蛋白通过基因本体论数据库(GO)和京都基因和基因组数据库(KEGG)进行分析,筛选凝血相关差异前6种蛋白并分析.符合正态分布的计量数据以均数±标准差((-x)±s)表示,采用独立样本t检验比较两组均值,以P＜0.05为差异有统计学意义.结果 电镜下外囊泡呈典型双层膜类圆形颗粒结构,Western blot法显示TSG101、CD63、CD9、CD81阳性,两组外囊泡平均粒径分别为168.0 nm和153.5 nm.定量PLT-EVs中853种蛋白,其中191种蛋白上调,77种蛋白下调,差异蛋白GO和KEGG分析表明:主要涉及凝血功能、炎性反应、免疫反应和机体修复调节等生物过程.差异量前6位参与凝血反应蛋白为纤维酶原、细胞分裂蛋白5、Rho相关蛋白激酶1、Rap1GTP酶激活蛋白2、GRB2相关接头蛋白2和蛋白激酶Cα型.结论 PLT-EVs差异蛋白分析可为危重烧伤患者血小板功能及对凝血反应的调节机制提供依据.
Abstract
Objective To explore the differential proteins related to coagulation reaction in platelet extracellular vesicles(PLT-EVs)between critically burned patients and healthy volunteers by proteomics technology.Methods A total of 6 healthy volunteers and 6 patients with severe burns in the Third Affilia-ted Hospital of Inner Mongolia Medical University were selected,and venous blood was extracted.PLT-EVs were obtained by high-speed centrifugation,and identified by transmission electron microscopy,Nanoparti-cle Tracking Analysis and Western blotting.The proteomic techniques were used to analyze and quantify two groups of PLT-EVs proteins.The differential proteins were screened with a differential multiple of 1.5(up-regulated or down-regulated).Differentially expressed proteins were analyzed through Gene Ontology(GO)and Kyoto Encyclopedia of Genes and Genomes(KEGG)databases.The top 6 proteins related to co-agulation differences were screened and analyzed.The measurement data in line with the normal distribu-tion were expressed as mean±SD,and the independent sample t test was used to compare the mean value of the two groups.P＜0.05 was considered to be statistically significant difference.Results Under the electron microscope,the PLT-EVs showed typical double-layer membrane.The TSG101,CD63,CD9 and CD81 were identified positively by Western blotting.The average particle sizes of the PLT-EVs were 168 nm and 153.5 nm respectively.A total of 853 proteins in PLT-EVs were identified by quantitative analysis,with 191 proteins being up-regulated and 77 proteins being down-regulated.The GO and KEGG analysis showed differential proteins were mainly involved in biological processes,such as coagulation func-tion,inflammatory response,immune response and body repair regulation.The top 6 proteins related to co-agulation reaction were plasminogen,dedicator of cytokinesis protein 5,rho-associated protein kinase 1,rap1 GTPase-activating protein 2,GRB2-related adapter protein 2 and protein kinase C alpha type.Conclusion PLT-EVs differential protein analysis could provide important theoretical basis for the regula-tion mechanism of platelet function and coagulation reaction in critically burned patients.
关键词
烧伤/血小板/外囊泡/蛋白组学/凝血Key words
Burns/Platelet/Extracellular vesicles/Proteomics/Coagulation引用本文复制引用
基金项目
内蒙古自然科学基金(2021MS08066)
内蒙古医学科学院公立医院科研联合基金科技项目(2023GLLH0245)
内蒙古医科大学联合项目(YKD2024LH015)
出版年
2024
NETL
NSTL
万方数据