CAJ | 학술논문

目的建立实时荧光定量聚合酶链反应(RFQ-PCR)检测非小细胞肺癌(NSCLC)患者痰中增殖诱导配体(APRIL)mRNA含量的方法,探讨痰脱落细胞APRIL mRNA表达在NSCLC诊断中的意义.方法对2007年8月至2008年5月南通大学附属医院71例NSCLC患者和62例肺部良性病变患者痰中APRIL mRNA进行检测,同时取65名健康成人痰做对照.用RFQ-PCR技术实时检测PCR产物的荧光强度,由软件自动计算出待测样本中靶基因mRNA的准确含量,以靶基因和内参β2-微球蛋白(β2-M)mRNA含量的比值作为评价其表达水平的指标,并与细胞学检查结果进行比较.计量资料采用t检验,所得结果用x±s表示;对计数资料采用X2检验.结果 RFQ-PCR法检测APRIL mRNA含最的线性范围为38～3.8×106copies/ul,批内和批间变异系数(CV)分别为8.5%和13.6%.NSCLC组痰中APRIL mRNA表达水平显著高于肺部良性病变组和健康对照组(t值为10.50和11.32,P<0.01);良性病变组与健康组比较差异无统计学意义(t=0.379,P>0.05).以健康组APRIL mRNA的x±2s为cut-off值,肺癌组APRIL mRNA表达的阳性率为81.7%(58/71),明显高于肺部良性病变组的3.2%(2/62)和健康对照组的1.5%(1/65).NSCLC组痰中APRIL mRNA的表达与性别、年龄、吸烟史、TNM分期及淋巴结转移尤关(t值分别为1.700、1.014、1.484、1.298及1.186,均P>0.05),与病理分型和肿瘤部位有关(t值为1.650和1.873,均P<0.05).RFQ-PCR阳性率为82%(58/71),高于细胞形态学阳性率的14%(10/71),差异有统计学意义(X2=67.68,P<0.01).结论 RFQ-PCR检测痰中APRIL mRNA含量具有较高的敏感度和特异度,有助于NSCLC的临床诊断.
목적 건립실시형광정량취합매련반응(RFQ-PCR)검측비소세포폐암(NSCLC)환자담중증식유도배체(APRIL)mRNA함량적방법,탐토담탈락세포APRIL mRNA표체재NSCLC진단중적의의.방법 대2007년8월지2008년5월남통대학부속의원71례NSCLC환자화62례폐부량성병변환자담중APRIL mRNA진행검측,동시취65명건강성인담주대조.용RFQ-PCR기술실시검측PCR산물적형광강도,유연건자동계산출대측양본중파기인mRNA적준학함량,이파기인화내삼β2-미구단백(β2-M)mRNA함량적비치작위평개기표체수평적지표,병여세포학검사결과진행비교.계량자료채용t검험,소득결과용x±s표시;대계수자료채용X2검험.결과 RFQ-PCR법검측APRIL mRNA함최적선성범위위38～3.8×106copies/ul,비내화비간변이계수(CV)분별위8.5%화13.6%.NSCLC조담중APRIL mRNA표체수평현저고우폐부량성병변조화건강대조조(t치위10.50화11.32,P<0.01);량성병변조여건강조비교차이무통계학의의(t=0.379,P>0.05).이건강조APRIL mRNA적x±2s위cut-off치,폐암조APRIL mRNA표체적양성솔위81.7%(58/71),명현고우폐부량성병변조적3.2%(2/62)화건강대조조적1.5%(1/65).NSCLC조담중APRIL mRNA적표체여성별、년령、흡연사、TNM분기급림파결전이우관(t치분별위1.700、1.014、1.484、1.298급1.186,균P>0.05),여병리분형화종류부위유관(t치위1.650화1.873,균P<0.05).RFQ-PCR양성솔위82%(58/71),고우세포형태학양성솔적14%(10/71),차이유통계학의의(X2=67.68,P<0.01).결론 RFQ-PCR검측담중APRIL mRNA함량구유교고적민감도화특이도,유조우NSCLC적림상진단.
Objective To establish a real-time fluorescence quantitative polymerase chain reaction (RFQ-PCR) method for quantifying a proliferation-inducing ligand (APRIL) Mrna in sputum samples from patients with non-small cell lung cancer (NSCLC), and to evaluate its role in the diagnosis of NSCLC. Methods Seventy-one cases of NSCLC and 62 cases of benign pulmonary disease were enrolled in this study from August 2007 to May 2008 in Affiliated Hospital of Nantong University, Jiangsu. Sixty-five healthy volunteers served as the control. The fluorescence of the PCR products was detected continuously during the amplification by RFQ-PCR. According to the standard curves created by plasmid DNA, the expression level of target genes in clinical samples was determined using software. The results were presented as the ratios of target genes to β2-microglobulin (β2-M ) Mrna, and compared with those obtained by conventional cytological method. Results The detection range of the assay was from 38 copies/ul to 3.8× 106 copies/ul. The coefficients of variation values of both intra-experimental and inter-experimental reproducibility were 8.5% and 13.6% ,respectively. The expression of APRIL Mrna in tumor sputum was higher than that in benign pulmonary disease and healthy volunteers (t=10.50, 11.32, P<0.01). The positive rate for APRIL Mrna expression was 81.7% (58 of 71) in sputum samples of NSCLC, 3.2% (2/62) in benign pulmonary disease and 1.5% (1/65) in healthy volunteers when cut-off values for positivity were set at the x±2s of Mrna expression in health volunteers. The level of APRIL Mrna of NSCLC was not related to sex, age, smoking status, TNM stage and lymph node metastasis (P>0.05, respectively), but was related to pathology subtype and the location of tumors (P<0.05, respectively). The APRIL Mrna assay (82%) produced a higher detection rate than conventional cytological method (14%) (X2= 67.68, P<0.01). Conclusion Measurement of the expression of APRIL Mrna in sputum by RFQPCR showed high sensitivity and specificity, which maybe useful in diagnosing NSCLC.