非典型甲状腺结节细胞学染色标本检测BRAF^(V600E)临床应用价值

Clinical application value of BRAF^(V600E) in the detection of atypical thyroid nodule cytology staining specimens

目的甲状腺结节发病率高,正确判断结节性质对临床诊断和治疗具有重要意义。本研究探讨细胞学染色标本检测BRAFV600E在性质不确定甲状腺结节临床诊断和术前指导的应用价值。方法选取2015-01-01-2016-12-31在济宁医学院附属医院病理科进行细针穿刺的甲状腺结节患者139例,其中54例行甲状腺切除术。139例患者在细针穿刺活检(fine needle aspiration biopsy,FNAB)后提取DNA,采用实时荧光定量聚合酶链反应结合突变阻滞形成系统(real time-amplification refractory mutation system-quantitative PCR,RT-ARMS-qPCR)检测BRAFV600E突变状态(FNAB-BRAFV600E检测法);细针穿刺细胞学(fine needle aspiration cytology,FNAC)诊断后,提取经HE染色的细针穿刺涂片(fine needle aspiration smear,FNAS)上细胞DNA,RT-ARMS-qPCR检测BRAFV600E突变状态(FNAS-BRAFV600E检测法)。提取54例手术患者甲状腺组织石蜡标本中的DNA,RT-ARMS-qPCR检测BRAFV600E突变状态(石蜡-BRAFV600E检测法)。分析3种检测方法的诊断价值,比较技术优势。结果139例患者FNAB和FNAS标本BRAFV600E突变情况完全一致,Kappa=1.00,P<0.001;54例术后石蜡标本和其术前FNAS标本BRAFV600E突变情况完全一致,Kappa=1.00,P<0.001。32例FNAC诊断为良性病变,未进行手术,均无BRAFV600E突变;26例性质不确定患者进行了手术,其中17例术后组织病理学诊断为甲状腺乳头状癌(papillary thyroid carcinoma,PTC),3种方法检测均为BRAFV600E突变型,另外9例诊断为非PTC,3种检测均无BRAFV600E突变;22例诊断为可疑PTC患者,18例进行了手术,术后组织病理学诊断均为PTC,3种检测均为BRAFV600E突变型;17例FNAC诊断为PTC的患者,10例进行了手术,术后组织病理学诊断均为PTC,有1例患者3种检测BRAFV600E状态均为野生型,其余均为突变型,Kappa=0.27,χ~2=67.87,P<0.001。结论 FNAS标本和FNAB标本及术后石蜡标本BRAFV600E突变情况完全一致,对FNAC诊断性质不确定的甲状腺结节患者,FNAS-BRAFV600E检测方法操作更简便,有助于术前明确结节的性质,可以用于术前辅助诊断并指导手术。

OBJECTIVE In view of the high incidence of thyroid nodules and the importance of correctly judging the nature of nodules for clinical diagnosis and treatment,this study investigated the value of BRAF^V600E detection in the clini-cal diagnosis and preoperative guidance of uncertain properties thyroid nodules on fine needle aspiration biopsy(FNAB) specimens.METHODS A total of 139 patients with thyroid nodules who underwent fine needle aspiration at the depart-ment of Pathology,Affiliated Hospital of Jining Medical University were selected from January 1st,2015,to December 31st,2016,and 54 of them underwent surgery.DNA were extracted from all of these 139 patients’FNAB specimens and the BRAF^V600E mutation was detected by real time-amplification refractory mutation system-quantitative PCR(RT-ARMS-qPCR, FNAB-BRAF^V600E assay).HE stained smear DNA(fine needle aspiration smear,FNAS) were extracted after fine needle aspiration cytology(FNAC), diagnoses were finished and the BRAF^V600E mutation was detected (FNAS-BRAF^V600E assay). At the same time,the DNA in the thyroid tissue paraffin specimens of 54 patients was also extracted,in which the RT-ARMS-qPCR was applied to detect the BRAF^V600E mutation status(paraffin-BRAF^V600E assay).Three test methods were used to analyze clinical diagnostic value as well as technical advantages among them.RESULTS The BRAF^V600E mu-tations in FNAB were completely consistent with that of FNASs pecimens in 139 patients (Kappa=1.00,P<0.001);54 patients with postoperative specimens and their preoperative FNAS specimens had identical BRAF^V600E mutations (Kappa=1.00,P<0.001).Totally 32 cases of FNAC were diagnosed as benign lesions without operation and no BRAF^V600E mutation was found;26 cases of patients with uncertain nature nodules underwent surgery,of which 17 cases were diagnosed as papillary thyroid cancer (PTC) histopathologically with observation of BRAF^V600E mutation in all three methods.As for the other 9 cases who were diagnosed as non-PTC,no BRAF^V600E mutations were detected in any of the three methods.Totally 22 patients were diagnosed as suspicious for PTC,and 18 of them subsequently had operations. Paraffin sections further confirmed the diagnosis of PTC,and BRAF^V600E mutation were found in all three detection meth-ods. Among 17 patients who were diagnosed as PTC in FNAC,10 of them underwent surgery followed by the confirmation of the diagnosis of PTC histologically.Only one patient showed wild-type BRAF^V600E status and the mutant type could be found in all the others(Kappa=0.27,χ^2=67.87,P<0.001).CONCLUSIONS FNAS specimens,FNAB specimens and postoperative paraffin specimens are completely consistent in detcting BRAF^V600E mutations but for patients with thyroid nodules with uncertain FNAC diagnostic properties,FNAS-BRAF^V600E detection method is easier to operate and can deter-mine the nature of thyroid nodules before surgery,therefore it can provide preoperative auxiliary diagnosis and guidance of surgery.