血浆MicroRNA用于急性主动脉夹层早期诊断和预后判断的临床基础研究

Acute aortic dissection (AAD) is a severe cardiovascular disease with high mortality and morbidity , which may be rapidly fatal without early diagnosis and appropriate management .A timely and accurate diagnosis is the key to achieve a positive outcome. MicroRNAs (miRNAs) are small regulatory RNAs that have shown promise as tissue-basedmarkers for cardiovascular disease classi?cation and prognostication. Our previous studies show that miRNAs（miR-23a，miR-150 etc.） have specific expression in the intima of aortic dissection . We hypothesized that miRNAs could be an ideal class of blood-based biomarkers for AAD detection. In this study, microarray was performed on plasma of controls and AAD patients. Completing the statistical analysis for significance and prediction, we selected candidate miRNAs for a second large study by RT-qPCR in controls and whom were diagnosed with AAD . Additionally, we combined specific miRNA with different stages and subtypes of ADD to complete the integrative analysis. Finally, we explore the histology of specific miRNA through animal experiments, and assess the stability of miRNA. Given this, we speculate that miRNAs could have value in the setting of detecting AAD for which clinically validated blood biomarkers are lacking. Biochemical assay of AAD is noninvasive, inexpensive, rapid and easy to perform and may be a potential tool for early detection and diagnosis of suspected AAD. These biomarkers provide important clues in the diagnosis, differential diagnosis and prognosis of AAD.

急性主动脉夹层（AAD）破裂死亡率极高，早期诊断并施以合理的治疗可大幅提高患者生存率。循环血中微小RNA（miRNA）是一类新近研究发现具有分子诊断标志物优点的内源性小分子核苷酸序列。本项目组前期研究表明夹层主动脉中膜存在miRNA的特异改变，并通过高通量技术初步筛选出一组AAD患者循环血中差异miRNA（miR-23a，miR-150等）；那么循环血中miRNA是否可以作为AAD的生物标记物呢？本研究拟：①针对芯片筛选结果进行芯片显著性分析和预测分析以获得与AAD密切相关的特定miRNA；②针对特定miRNA进行大样本的定量验证,分析其对AAD早期诊断的特异性及敏感性；③特定miRNA与AAD患者病程转归、预后及分型等临床资料相结合进行综合分析；④通过动物实验探索特定miRNA的组织学基础，并对其进行稳定性评价。以期为AAD的早期诊断和预防、病情监测寻找临床可用的实验室检查指标。

目的：急性主动脉夹层（acute aortic dissection，AAD）破裂死亡率极高，是灾难性的临床急症之一；早期诊断并施以合理的治疗是挽救患者生命的关键。虽然诊疗技术在不断进步，其临床误诊率仍然高达39%。本研究以急性主动脉夹层患者血浆中miRNA为研究对象，为急性主动脉夹层的早期诊断和病情监测寻找临床可用的实验室检查指标。方法：本研究纳入受试者共103例，其中主动脉夹层患者63例和对照组40例（包括：健康志愿者，n=17；急性心肌梗死，n=11；主动脉动脉瘤，n=10；肺梗死]，n=2）。首先，利用高通量筛选技术对24例标本（急性主动脉夹层组，n=8；对照组，n=16）对1205种人类miRNA和142种人病毒miRNA进行筛选；然后，在全部受试者中利用实时荧光定量逆转录链式聚合酶扩增技术对筛选得到的候选miRNA进行定量验证；最后，我们利用接收者操作特征（receiver operating characteristic, ROC）曲线并计算曲线下面积（area under the receiver operating characteristic curve, AUC）等统计手段结合参与受试者的临床资料评价特定miRNA用于急性主动脉夹层诊断的可靠性。结果：通过高通量筛选共获得16种候选miRNA，并在整个样本中进行qRT-PCR定量验证；经验证，血浆中miR-15a在急性主动脉夹层组（n=37）与对照组（n=40）比较P=0.008，差异倍数（fold change，FC）=2.795；ROC分析显示miR-15a对急性主动脉夹层的检测敏感度为75.7%，特异性为82.5%，AUC为0.761。进一步研究提示，37例急性主动脉夹层患者血浆中let-7b, miR-15a, miR-23a and hcmv-miR-US33-5p水平显著高于14例非夹层的胸痛患者(P=0.000, 0.000, 0.026和0.011; FC=8.462, 8.938, 17.851和21.975)。ROC分析提示诊断准确率分别是0.887, 0.855, 0.925和0.815。结论：研究发现在急性主动脉夹层患者血浆中miR-15a和miR-23a的表达水平显著增高，并持续在较高水平直至患者获得合理的治疗；对急性主动脉夹层的临床诊断和病程监控具有重要意义。