血浆SHOX2/RASSF1A/PTGER4基因甲基化检测对肺结节的诊断价值
The Diagnosis Value of Plasma SHOX2/RASSF1A/PTGER4 Gene Methylation Detection in the Pulmonary Nodules

【目的】探究血浆SHOX2/RASSF1A/PTGER4基因甲基化检测对肺结节性质判定及诊断的价值。【方法】抽取静脉血8~10 ml进行血浆分离，分离后的血浆进行DNA的提取与制备，其中包括裂解、结合、洗涤、亚硫酸盐转化、脱洗等步骤。进行PCR检测，分别设置SHOX2、RASSF1A、PTGER4甲基化的阈值，检测游离DNA中基因甲基化状态。采用首创的P值(阳性指数，Positive Index)计算公式法分析样本检测数据，依据阳性判断值判定样本阳阴性。【结果】84例胸部CT提示肺结节的患者，进行血浆游离DNA甲基化检测，其中阳性和阴性分别有18例和66例，阳性与阴性结果与患者的性别、吸烟、结节大小、多发与否无关；与年龄、结节的密度及病理类型有关，差异有统计学意义(P < 0.05)。通过金标准病理诊断肺癌有8例，其中6例甲基化检测为阳性，阳性率为75%；恶性结节与良性结节的甲基化阳性指数数值分别为2.00 ± 1.38和1.10 ± 0.92，差异有统计学意义(P < 0.05)。SHOX2/RASSF1A/PTGER4基因甲基化诊断肺癌的曲线下面积(AUC)、灵敏度和特异度分别为0.754、75%和84.2%。SHOX2/RASSF1A/PTGER甲基化与病理诊断评定结果存在一致性，Kappa = 0.842。【结论】SHOX2/RASSF1A/PTGER4基因甲基化检测可以成为肺结节性质早期判断的一种可靠且灵敏的新型生物学标志物，为肺癌的早期诊治提供依据。
[Objective] To investigate the value of plasma SHOX2/RASSF1A/PTGER4 gene methylation detection in the identification and diagnosis of pulmonary nodules. [Method] Venous blood 8 - 10 ml was collected for plasma separation, and the separated plasma was used for DNA extraction and preparation, which included the steps of lysis, binding, washing, sulfite conversion, desorption and other steps. DNA methylation levels of SHOX2, RASSF1A and PTGER4 were detected by real-time fluorescence quantitative PCR. According to the set threshold, the analysis was performed using dedicated interpretation software. The original P value (Positive Index) calculation formula method was used to analyze the sample test data, and the positive and negative samples were determined according to the positive judgment value. [Result] In 84 patients with pulmonary nodules indicated by chest CT, plasma free DNA methylation of SHOX2/RASSF1A/PTGER4 gene was detected. A total of 84 patients with pulmonary nodules indicated by chest CT were selected for plasma SHOX2/RASSF1A/PTGER4 gene methylation detection, including 18 cases of positive result and 66 cases of negative result. The positive and negative results were not related to gender, smoking, nodule size, or multiple nodules. It was related to age, density of nodules and pathological type. There were 8 patients who were pathologically diagnosed as lung cancer, of which 6 were positive by SHOX2/RASSF1A/PTGER methylation test, and the positive rate was 75%. The methylation values of malignant and benign nodules were 2.00 ± 1.38 and 1.10 ± 0.92, respectively. The area under the curve (AUC), sensitivity and specificity of plasma SHOX2/RASSF1A/PTGER4 gene methylation in diagnosis of lung cancer were 0.754, 75% and 84.2%, respectively. SHOX2/RASSF1A/PTGER methylation was consistent with pathological diagnosis, Kappa = 0.842. [Conclusion] Detection of SHOX2/RASSF1A/PTGER4 gene methylation in plasma