急性心肌梗死相关循环微小RNA快速检测新方法的研究

Missed diagnosis and misdiagnosis are major problems in the clinical diagnosis and treatment of acute myocardial infarction (AMI). In recent years, studies have shown that several circulating microRNAs (miRNA) as a new kind of biomarkers can be used for the auxiliary diagnosis of AMI, and effectively reduce the rate of missed diagnosis and misdiagnosis. However, it is regrettable that the existing miRNA detection methods cannot meet both the necessary conditions for AMI diagnosis at the same time - a short detection time (within 1 hour) and a high sensitivity (10-14 to 10-13 mol / liter). This project proposes the effective integration of the three aspects of "rapid capture of miRNA by click chemistry", "signal amplification of electrocatalytic deposition of polydopamine" and "acquisition of electrochemical detection signals" to develop a novel method for the electrochemical detection of miRNA with favorable timeliness and sensitivity. Biosensing mechanism, kinetic process and etc. will be deeply studied; the key performance of the detection will be optimized; animal model and clinical samples will be used for the actual detection; and bioinformatics method will be adopted to establish a comprehensive diagnostic index. Through this project, it is expected to develop a novel miRNA detection method that can meet the diagnostic needs of AMI, and thus contribute to the application of miRNA in the clinical diagnosis of AMI.

漏诊及误诊是急性心肌梗死（AMI）临床诊治的重大问题。近年来的研究表明，几种循环微小RNA（miRNA）作为新型标志物可用于AMI的辅助诊断，并有效地降低漏诊率和误诊率。但遗憾的是，现有miRNA检测方法难以同时满足AMI诊断所需的两个必要条件——检测时间短（1小时内）和灵敏度高（10-14~10-13摩尔/升）。本项目提出通过“点击化学介导的miRNA界面快速捕获”，“电催化聚多巴胺的信号放大”，以及“电化学检测信号的获取”三个方面的有效整合，从而发展可以兼顾检测的时效性和灵敏性的miRNA电化学检测新方法。对其中涉及的生物传感机理、动力学过程等进行深入的研究，对检测的关键性能进行重点优化，运用动物模型和临床样本进行实际检测，运用生物信息学方法建立诊断综合指标。通过本项目的开展，有望发展出可满足AMI诊断需求的miRNA检测新方法，从而为推动miRNA在AMI临床诊断中的应用做出贡献。

本项目针对急性心肌梗死（AMI）临床诊断中有可能出现的漏诊误诊，提出以几种循环微小RNA（miRNA）作为新型标志物辅助AMI的诊断。项目开展过程中，通过miRNA界面快速捕获、聚多巴胺辅助的信号放大以及三种检测手段（比色法、荧光法和电化学方法）对miRNA实现分级检测，发展出可以兼顾时效性和灵敏度的miRNA快速检测方法。以急性心肌梗死相关的miR-208a-3p为例，采用比色法可以在100 pM至500 nM的浓度范围内实现线性检测，检测限为22.87 pM；采用荧光法的线性检测范围为10 pM至100 nM，检测限为3.37pM；电化学（阳极溶出伏安法）方法的线性检测范围为1 pM至10 nM，检测限为0.3 pM，较比色法低76倍。其中，比色法和荧光法检测时间均在10分钟左右，相当快速，而电化学方法则有更高的灵敏度。项目对AMI相关的其他几种miRNA，如miR-1和miR-499等同样进行了检测，取得了类似的结果。本项目为急性心肌梗死的临床筛查提供了潜在的快速检测方案。