血浆外泌体miR-452作为胃癌诊断的生物标志物

The efficacy of miR-452 in plasma exosomes as a biomarker for the diagnosis of gastric cancer

目的筛选胃癌血浆外泌体中差异表达的miRNA,分析其对胃癌诊断的敏感性与特异性及与胃癌患者的疾病特点之间的相关性。方法收集南京医科大学第一附属医院消化内科病理确诊为胃癌的住院患者与消化内科及体检中心的无癌健康对照人群的血液样本,离心后试剂盒抽提血浆外泌体及其RNA,通过透射电子显微镜观察、蛋白质印迹分析(Western blotting)鉴定标志蛋白及纳米粒径分析(nanoparticle tracking analysis, NTA)对血浆外泌体进行鉴定,定量逆转录聚合酶链反应(qRT-PCR)鉴定与胃癌发病相关的miRNA。结果共纳入胃癌患者80例和健康对照80名。成功对2例血浆外泌体(GC1和NC1)进行鉴定以证实其相关特性。在初始筛选阶段,对12例胃癌和12名健康对照者的血浆外泌体RNA进行qRT-PCR验证并依据其各自的2^-△Ct值,分析受试者的诊断ROC曲线下面积(AUC)。与健康对照者相比,胃癌的血浆外泌体miR-452-5p表达量显著升高(P=0.002),miR-452-5p的AUC值为0.798(95%CI:0.552~0.883)。而后纳入新的68对样本对筛选结果进一步验证,miR-452-5p的表达趋势与初始筛选一致,且诊断AUC值为0.733(95%CI:0.647~0.818)。分析已验证的miR-452-5p表达水平与胃癌病理特征无相关性(P>0.05)。结论血浆外泌miR-452-5p可能为检测胃癌的新型潜在生物标志物,然而其对疾病的病理特性的影响仍需更大样本量的实验进一步研究。

Objective To screen the differentially expressed miRNA in exosomes of gastric cancer, and to analyze the effects for the diagnosis of gastric cancer and its correlation with the disease characteristics of gastric cancer. Methods Blood samples were collected from hospitalized patients and healthy controls in the Department of Gastroenterology, the First Affiliated Hospital of Nanjing Medical University. After centrifugation, plasma exosomal RNA was extracted and the plasma exosomes was identified by Transmission electron microscopy, Western blotting, and nanoparticle tracking analysis(NTA), quantitative reverse transcriptase polymerase chain reaction(qRT-PCR) was used to identify the miRNAs. Results 80 patients with gastric cancer and 80 cases of healthy controls were included. In screening stage, qRT-PCR was performed on 12 gastric cancers and 12 healthy controls plasma exosomal RNAs and the area under the ROC curve(AUC) of the subjects was analyzed according to the 2^-△Ct of qRT-PCR. Compared with healthy controls group, miR-452-5p was significantly increased(P=0.002) in gastric cancer group, and the AUC value of miR-452-5p was 0.798(95% CI: 0.552-0.883). Then new 68 pairs were further validated and the results showed that the expression of miR-452-5p was consistent with the screening stage, and the AUC was 0.733(95% CI: 0.647-0.818). However, the relationship between the expression of miR-452-5p and the clinicopathological variables of gastric cancer was not statistically related(P>0.05). Conclusion Plasma exosomes miR-452-5p may be a potential biomarker for the detection of gastric cancer. However, its exact effect on the pathological characteristics of the disease still requires further experimental studies.