海藻糖低温保存血小板miRNA差异表达分析  

Differential expression profiles of platelet miRNAs after trehalose cryopreservation

作　　者：黄梅莓[1] 王世春[1] 张强[1] 宋敏[1] 吴建君[1] 陈敏[1] 陈寒梅[1] 李晨晨[1] 张丹[1] 刁荣华[1] 赵树铭[1] 

机构地区：[1]第三军医大学西南医院输血科,中国人民解放军重庆血站,重庆400038

出　　处：《第三军医大学学报》2016年第18期2066-2070,共5页Journal of Third Military Medical University

基　　金：国家自然科学基金面上项目(81270649)~~

摘　　要：目的探讨海藻糖低温保存血小板过程中miRNA的变化。方法收集3份来自相同血型无偿献血员的单采血小板,少量混匀后,平均分为2组。对照组:新鲜血小板组(22℃,100%血浆储存的新鲜血小板2 h);实验组:海藻糖低温保存血小板组(10℃,70%海藻糖保存液储存5 d);采用高通量测序方法对两组样品进行了Small RNA测序,进行生物信息分析,主要包括已知miRNA鉴定、miRNA差异表达分析及富集分析。结果 2组样品经Small RNA高通量测序,平均每个样品获得了11兆净读值(Clean reads),经过生物信息分析,分别获得了821个已知miRNA。通过差异表达分析,在已知的miRNA中共有460个差异表达miRNA,其中194个miRNA表达上调,266个miRNA表达下调,其中显著差异表达已知miRNA共44个,分别在钙离子通路、MAPK通路、凋亡路径、轴突导向等储存损伤相关路径中富集程度较高。相对于对照组,实验组中hsa-miR-4449、hsa-miR-1296表达显著上调(P<0.01,且差异倍数>2),hsa-miR-665表达被抑制。结论血小板经过海藻糖低温保存后的miRNA呈现出特征性表达变化,提示miRNA活动参与了低温时海藻糖对血小板储存损伤的保护作用。Objective To determine the changes of platelet microRNAs duing trehalose cryopreservation. Methods Three bags of apheresis platelets derived from 3 healthy volunteer blood donors of same blood type were firstly mixed well, and subsequently distributed into the control group （22 ℃, with 100% plasma for 2 h） and the experimental group [ 10 ℃, with 70% trehalose preservation solution （TPS） for 5 d 1. High throughput small RNA sequencing was applied to the 2 groups of platelets, and then the obtained bioinformatic data were analyzed, including known miRNAs identification, differential expression analysis and KEGG pathway analysis. Results High throughput small RNA sequencing obtained 11 million clean reads data for each sample, and bioinformatic analysis identified 821 known miRNAs. In the known miRNAs, a total of 460 miRNAs were differentially expressed, among which 194 were up-regulated and 266 down-regulated. Among these miRNAs, 44 known microRNAs had significant differences in expression level, and they were respectively enriched in the signaling pathways relative to storage impairment, such as calcium signaling pathway, MAPK pathway, apoptotic pathway and axon guidance. Compared to the control group, the expression levels of hsa-miR-4449 and has-miR-1296 were up-regulated significantly in experiment group （P 〈0.01, with a fold change of 〉 2）, but the expression of hsa-miR-665 was suppressed. Conclusion After low temperature storage in TPS, platelets have distinctive profiles of differences in miRNAs expression, which suggested that miRNAs are involved in the protection of trehalose to platelet impairment in low temDerature.

关 键 词：血小板 血小板储存损伤 MIRNA 高通量测序 

分 类 号：R331.143[医药卫生—人体生理学] R394.2[医药卫生—基础医学]