荧光原位杂交与逆转录聚合酶链反应在尤文肉瘤/原始神经外胚层肿瘤分子诊断中的应用

Application of fluorescence in-situ hybridization and reverse transcription-polymerase chain reaction in molecular diagnosis of Ewing′s sarcoma and primitive neuroectodermal tumor

目的探讨运用荧光原位杂交(FISH)与逆转录聚合酶链反应(RT-PCR)检测尤文肉瘤/原始神经外胚层肿瘤(ES/PNET)石蜡包埋组织特异性染色体易位的临床应用价值。方法收集ES/PNET10例,分离石蜡包埋组织中的肿瘤细胞,利用Vysis公司的EWSR1双色易位分离探针,间期核FISH检测EWS基因的易位。运用RT-PCR检测t(11;22)(q24;q12)和t(21;22)(q22;q12)形成的融合转录子EWS-FLI1和EWS-ERG。结果10例ES/PNET中FISH观察9例有EWS基因的易位。RT-PCR8例检测出EWS-FLI1的表达,没有检测出EWS-ERG的表达。结论利用FISH与RT-PCR检测ES/PNET石蜡包埋组织特异性染色体易位可作为可靠的分子诊断指标;两者相比,FISH敏感性和稳定性要优于RT-PCR。

Objective To detect tumor specific chromosome translocations and associated fusion transcripts in paraffin-embedded tissue by interphase fluorescence in-situ hybridization（FISH） and reverse transcription polymerase chain reaction（ RT-PCR）, respectively, and to evaluate their diagnostic values for Ewing＇s sarcoma/primitive neuroectodermal tumor （ES/PNET）. Methods Nuclei of the tumor cells and total RNA were extracted from 10 cases of ES/PNET. Interphase FISH was utilized to analyze the EWS gene translocation with a dual color, break apart probe （Vysis company）. RT-PCR was used to detect t（ 11 ;22） （q24;q12） and t（21 ;22） （q22;q12） fusion transcripts. Results Among 10 cases of ES/PNET, the EWS- FLI1 fusion transcript was detected in 8 by RT-PCR. EWS-ERG fusion transcript was not detected in any of the cases. EWS gene translocation was found in 9 of 10 cases by FISH. Condusions Interphase FISH and RT-PCR can be reliably applied to paraffin-embedded tissues for molecular diagnosis of ES/PNET. Between the two approaches, interohase FISH provides a more sensitive and stable result.