亚甲蓝光化学法病毒灭活血浆外泌体微小RNA差异表达谱分析  

Differential expression profile of exosomal miRNA in virus inactivated plasma treated with methylene blue photochemistry

作　　者：陈倩[1] 冯增强 谭琳[2] 饶彩霞[1] 张炯炯[1] 赵静[1] 唐秋萍[1] CHEN Qian;FENG Zengqiang;TAN Lin;RAO Caixia;ZHANG Jiongjiong;ZHAO Jing;TANG Qiuping(Hainan Blood Center,Haikou 570311,Hainan Province,China;不详)

机构地区：[1]海南省血液中心,海南海口570311 [2]中国热带农业科学院海口实验站,海南海口571199

出　　处：《中国生物制品学杂志》2024年第7期817-823,共7页Chinese Journal of Biologicals

基　　金：海南省自然科学基金面上项目(820MS159)。

摘　　要：目的 探讨亚甲蓝光化学法(methylene blue photochemistry,MB-P)病毒灭活对血浆外泌体微小RNA(microRNA,miRNA)表达的影响,以期为MB-P病毒灭活血浆的质量控制提供新的参考指标。方法 收集2021年7月—2022年4月采集的健康无偿献血者全血11人份。离心获得血浆,同一人份新鲜血浆分为2份,分别制备为新鲜冰冻血浆(fresh frozen plasma,FFP)和MB-P病毒灭活血浆。采用差速离心法提取血浆中的外泌体,通过透射电子显微镜(transmission electron microscope,TEM)和纳米颗粒跟踪分析(nanoparticle tracking analysis,NTA)仪对提取的外泌体进行鉴定,并利用芯片技术检测外泌体miRNA的表达谱。采用qRT-PCR法对差异表达的4个miRNA进行验证,生物信息学方法预测差异表达miRNA的靶基因,并对差异表达基因进行GO功能富集分析和KEGG信号通路分析。结果 提取的两组囊泡状物质的形态特征及直径大小均符合外泌体特征。与对照组比较,MB-P组血浆外泌体中存在14个差异表达的miRNA,其中6个miRNA表达上调,8个miRNA表达下调。qRT-PCR验证结果与芯片表达趋势一致。差异表达的miRNA靶基因涉及DNA结合、离子结合、催化活性等功能,并参与核酸代谢、生物合成、转录调控等多种生物学过程;富集显著的功能通路与病毒感染性疾病、肿瘤、PI3K-Akt信号通路、MAPK信号通路等密切相关。结论 MB-P病毒灭活血浆和FFP外泌体miRNA存在差异性表达,血浆外泌体miRNA有望成为MB-P病毒灭活血浆质量评价的潜在参考指标。Objective To explore the effect of methylene blue photochemistry(MB-P)viral inactivation treatment on the expression of microRNA(miRNA)in plasma exosomes,in order to provide a new reference for the quality control of MB-P virus inactivated plasma.Methods Whole blood samples of 11 healthy volunteers were collected from July 2021 to April2022.Fresh plasma from the same person was prepared into two parts,fresh frozen plasma(FFP)and MB-P virus inactivated plasma,respectively.The plasma exosomes were isolated by differential centrifugation,and identified by transmission electron microscopy(TEM)and nanoparticle tracking analysis(NTA).Then the expression profiles of miRNA were detected by microarray technique.Furthermore,four differentially expressed miRNA were verified by qRT-PCR,the target genes of differentially expressed miRNA were predicted by bioinformatics methods,and GO function enrichment analysis and KEGG pathway enrichment analysis were performed on the differentially expressed genes.Results The morphological characteristics and diameters of the extracted vesicles of the two groups were consistent with the characteristics of exosomes.Compared with the control group,there were 14 differentially expressed miRNA in plasma exosomes of MB-P group,of which the expression of six miRNA was up-regulated and eight miRNA was down-regulated.The results of qRT-PCR were generally consistent with the expression trend of microarray.The target genes of differentially expressed miRNA were mainly involved in DNA binding,ion binding,catalytic activity and other functions,and participated in a variety of biological processes such as nucleic acid metabolism,biosynthesis,and transcription regulation.In addition,significantly enriched functional pathways were closely related to viral infectious diseases,tumors,PI3K-Akt signaling pathway,MAPK signaling pathway and so on.Conclusion The expression of exosome miRNA in MB-P virus inactivated plasma was different from that in FFP.The plasma exosome miRNA may be used as a potential referen

关 键 词：亚甲蓝光化学法 新鲜冰冻血浆 病原体灭活 外泌体 微小RNA 

分 类 号：R457.1[医药卫生—治疗学]