细胞HE染色涂片与细胞蜡块检测晚期非小细胞肺癌胸腔积液标本中表皮生长因子受体基因突变  

Cell HE staining smears and paired cell paraffin sections in detection of epithelial growth factor receptor gene of pleural fluid specimens

作　　者：侯芳[1] 齐长海[1] 卢一艳[1] 李方[1] 郝志红[1] HOU Fang;QI Changhai;LU Yiyan;LI Fang;HAO Zhihong(Department of Pathology,Aerospace Center Hospital,Beijing 100049,China)

机构地区：[1]航天中心医院病理科,北京100049

出　　处：《中南大学学报（医学版）》2022年第1期35-44,共10页Journal of Central South University ：Medical Science

摘　　要：目的:伴发胸腔积液的晚期非小细胞肺癌(non-small cell lung cancer,NSCLC)患者已失去手术机会,表皮生长因子受体(epithelial growth factor receptor,EGFR)酪氨酸激酶抑制剂(tyrosine kinase inhibitors,TKIs)是EGFR敏感突变的晚期NSCLC患者的一线用药。但EGFR-TKIs治疗时或治疗后出现的疾病进展以及药物的更新迭代给临床诊治带来了挑战,需要对存档的胸腔积液细胞样本进行EGFR复检或比对研究。本研究采用细胞HE染色涂片与细胞蜡块两种制作方法对NSCLC胸腔积液样本的EGFR突变基因进行检测,分析两种保存方式、保存时间对胸腔积液细胞DNA质量的影响,以期探索出一条保存细胞学资料及充分利用细胞学档案资料的可靠途径。方法:选取2014年10月—2021年4月航天中心医院病理科接收的胸腔积液样本57例,所有样本同时制作细胞HE染色涂片与细胞蜡块。将57例配对的细胞HE染色涂片与细胞蜡块采用扩增阻碍突变系统-聚合酶链反应(amplification refractory mutation system-polymerase chain reaction,ARMS-PCR)技术进行EGFR基因突变检测。DNA检测浓度为2 ng/μL,细胞HE涂片与配对细胞蜡块DNA样本并排进行扩增。结果判读按试剂说明书要求,待测样本的8号孔外控循环阈值(cycle threshold,Ct值)在13~21之间为样本合格,EGFR基因突变Ct值<26时,判断为阳性;26≤Ct值<29为临界阳性,Ct值≥29为阴性。ΔCt值为突变Ct值与外控信号Ct值的差值。比较细胞HE染色涂片与细胞蜡块检测为阳性的ΔCt值。同时将57例患者按时间段分为4组:2014—2015年(n=10),2016—2017年(n=20),2018—2019年(n=17),2020—2021年(n=10)。比较不同时间段的检测结果。结果:57例晚期NSCLC患者中以胸腔积液为首发症状入院者42例,占73.7%;57例中37例发生EGFR突变,突变率64.9%;19del突变和L858R突变为主要的突变类型,分别占37.8%(14/37)和48.6%(18/37);37例突变患者中女性占56.7%(21/37);细胞涂片与配对的Objective:The advanced non-small cell lung cancer(NSCLC)patients with pleural effusion have no opportunity for surgery treatment.Epidermal growth factor receptor(EGFR)tyrosine kinase inhibitors(TKIs)are the first-line drugs for these patients with EGFR-sensitive mutation.However,the disease progression and drug update during or after treatment of EGFR-TKIs bring more challenges and puzzles to clinical diagnosis and treatment,which inevitably requires archived pleural cell samples for EGFR reexamination or comparative study.Understanding the DNA quality of archived pleural fluid samples and effectively using archival data of pleural fluid cells are of great significance for tracing the origin of cases and basic medical research.This study aims to evaluate the consistency of EGFR mutant gene expression between the 2 methods,and to explore a reliable way for preserving cytological data and making full use of cytological archival data via cell HE staining smear and cell paraffin section.Methods:A total of 57 pleural fluid cytology cases in the Department of Pathology of China Aerospace Center Hospital from October 2014 to April 2021 were selected.Tumor cells were detected by cell HE staining smears and immunohistochemical staining for TTF-1 and Napsin A in the paired cell paraffin sections.There were more than 200 tumor cells in cell HE staining smear and the proportion of tumor cells were≥70% in matched cell paraffin sections.Patients with 2 cell smears(one for cell data retention and the other for DNA extraction)were selected as the research subjects,and 57 pleural fluid samples were enrolled.EGFR gene mutation was detected by amplification refractory mutation systempolymerase chain reaction in 57 paired cell HE staining smears and cell paraffin sections.DNA concentration was 2 ng/μL.Cell HE smear was amplified side-by-side with DNA samples from paired cell paraffin sections.Result determination was according to the requirements of the reagent instructions.The external control cycle threshold(Ct)value of the No.

关 键 词：表皮生长因子受体 非小细胞肺癌 胸腔积液 细胞HE染色涂片 细胞蜡块 

分 类 号：R734.2[医药卫生—肿瘤]