卡萨巴赫-梅里特现象患儿外周血循环micro RNA表达谱分析

Analysis of plasma micro RNA expression profiles in children with Kasabach-Merritt phenomenon

目的对卡萨巴赫-梅里特现象(Kasabach-Merritt phenomenon,KMP)患儿外周血循环微RNAs(micro RNAs, miRNAs)表达谱并进行差异分析。方法选取2016年10月至2017年3月广州市妇女儿童医疗中心介入血管瘤科收治的确诊为KMP的患儿3例(研究组)和年龄、性别相匹配的门诊健康体检的健康儿童3例(对照组)。两组患儿入院后进行药物治疗前均抽取外周静脉血2 ml,提取外周血血浆中总RNA。应用Illumina高通量测序技术进行miRNAs测序分析,采用生物信息学方法对差异表达miRNAs进行靶基因预测。结果研究组和对照组血浆RNA的纯度范围在1.37~1.83,浓度范围在88.04~286.17 ng/μl;血浆中small RNAs文库的摩尔浓度范围在30.3~48.2 nmol/L,片段长度范围在145~147 bp。研究组和对照组分别获得54 297 754与62 794 433条高质量的small RNA序列,均呈正态分布在22 nt序列两侧,其中20~24 nt序列所占比例分别为95.1%和87.0%。研究组和对照组的表达谱显示组间差异表达的miRNAs共22个。其中表达上调的有12个,以miR-423-3p表达上调最多,达7.3倍;表达下调的有10个,以let-7f-5p表达下调最多,达0.13倍。生物信息学预测差异表达的miRNAs的靶基因主要包括生物学过程、细胞过程、代谢过程、信号转导调控、细胞蛋白修饰、细胞生物合成调控等相关基因。结论KMP患儿外周血循环miRNAs差异表达谱的建立为探讨循环miRNAs作为生物标记物在KMP发病机制中的作用研究提供了科学依据。

Objective To analysis the plasma differential microRNA expression profiles in children with Kasabach-Merritt phenomenon (KMP). Methods Three children diagnosed with KMP and 3 age and gender-matched healthy outpatient children were chosen as objective and control groups. Two milliliter peripheral blood was extracted from both groups pre-treatment and total RNA isolated from plasma. The sequence of miRNAs was analyzed by Illumina high throughput sequencing. And the target genes of differentially expressed miRNAs were predicted by bioinformatics. Results The purity and concentration range of plasma RNA were 1.37-1.83 and 88.04-286.17 ng/μl in KMP and control groups respectively. The concentration range of small RNAs library and range of fragment length were 30.3-48.2 nmol/L and 145-147 bp in KMP and control groups respectively. The total small high-quality RNA sequences acquired from KMP and control groups were 54297754 and 62794433 respectively. All sequences showed normal distribution on both sides of 22 nt and sequence lengths with 20-24 nt accounted for 95.1% and 87.0% in KMP and control groups respectively. As compared with control group, 22 differentially expressed miRNAs including 12 up-regulated and 10 down-regulated miRNAs were identified in KMP group. Among the differentially expressed miRNAs, the up-regulating extent of miR-423-3P reached 7.3 folds and down-regulation of let-7f-5p 0.13 fold. Most target genes were involved in biological/cellular process, metabolism, regulation of signal transduction, cellular protein modification and regulation of biosynthetic process. Conclusions Establishing differential expression profiles for plasma miRNAs provides scientific rationales for elucidating the role of circulating miRNAs as biomarkers in the pathogenesis of KMP.