肝特异性循环exosomes的miRNA谱：一种潜在的HCC筛选标志

Current literatures suggested that Hepatocellular carcinoma (HCC) tissue miRNA profile could emerge as biomarker for early diagnosis of the cancer. Because of high degree of technical difficulty of toumor tissue biopsy and large area defect of wound, many groups have been intrested in finding biomarkers of this disease from serum miRNA profile. However, the origin of serum miRNAs is the key problem of these reseraches which have never been resolved. Therefor, they have not found the high identity serum miRNA profile as HCC specificity biomarker. Recently, a novel class of miRNAs in serum exosomes have been proven as stabilze and feasible biomarker of cancers. And these exosomes have characteristic of definite tissue origin and could be specific collected. Nevertheless, there is still no report about the miRNA profile, which is from serum exosomes with liver origin, used to be the biomarker of HCC. Consenqently, we will use the method of magnetic activated exosomes sorting to specific capture liver origin exosomes from HCC patients serum. Between these exosomes and the same patient origin of tumor cells, we will discover the identical and diagnostic specificity miRNA profile by miRNA microarray and large sample test of qRT-PCR. Then we will verify the miRNA profile can be the HCC biomarker. The result of this study will provide a novel and noninvasive screening tool for HCC early diagnosis, and conduce to enchance the HCC detection rate. Thus it will furtherly improve the prognosis of patients.

研究证明HCC组织miRNA表达谱可望成为HCC标志物用于肝癌早期诊断。因存在癌组织测定技术难度高和创面大的问题，人们开始在外周血中寻找有诊断价值的miRNA谱。但由于未解决血清中miRNAs来源这一关键问题，至今未找到同一性好、特异性高的肝癌血清miRNAs标志物。新近研究显示，循环exosomes有组织溯源性和可特异性富集性，其中的miRNA谱用于疾病的诊断是稳定、可行的。但目前将肝特异性循环exosomes中miRNA谱用于鉴定HCC还未见报道。综上，我们拟采用免疫捕获技术特异收集HCC患者血清中肝源性exosomes，运用miRNA芯片技术筛选和大样本QRT-PCR鉴定，找到这些exosomes中和同源癌细胞中表达一致、有诊断特异性的miRNA谱，并证明其可做为HCC的新生物标志物。该研究将为HCC的早期诊断提供新的、无创性筛查的工具，有助于提高HCC早期检出率，改善癌症患者预后。

大量研究认为循环miRNA对癌症的诊断很有前景。我们研究发现循环miRNA的抽提，逆转录和扩增步骤会直接影响血清miRNA的定量分析。现有的报道推荐采用肿瘤组相对于正常组的相对定量分析靶miRNA。但我们的实验和其他的实验结论也都证明，没有选择正确的内参基因，研究结论就会出现偏差。因此，我们展开了HCC 患者、HBV感染者和正常人群中循环exosome的内参miRNA的研究，以及HCC患者肝癌切除手术前后血清exosome中miRNA内参的研究。实验中逆转录实时荧光定量PCR的检测，我们严格按照MIQE标准执行。同时为了确保研究的可连续性和标本符合统计学要求，我们进一步研究了溶血标本对结果分析的干扰作用。研究结论发表了SCI论文2篇，1篇中文核心期刊,申请专利2项。