鼻咽癌干细胞miRNA表达特征研究

目的 探讨鼻咽癌干细胞的miRNA表达特征.方法 分离扩增CNE2细胞株鼻咽癌干细胞,流式细胞术检测其干细胞标志物CD133和CD44表达活性,并与鼻咽癌细胞6 10B、5-8F、CNE1、CNE2、HONE1及鼻咽上皮细胞NP69进行对比分析,继行肿瘤相关miRNAs的qRT-PCR检测.同时,在以免疫组化法检测不同临床分期鼻咽癌原发灶组织CD133和CD44表达活性基础上,qRT-PCR法分析其特异性miR 200b表达特征及其与临床分期的相关性.结果 CNE2鼻咽癌干细胞以CD133标志物为突出特征,联合检测CD44具有更高敏感性和特异性;在所检测的肿瘤相关miRNAs中,鼻咽癌干细胞以miR 200b活性低下甚至缺失为基本特征,甚至低于高转移潜能鼻咽癌细胞5 8F.鼻咽癌原发灶组织标本干细胞标志物也以CD133为突出特征,并平行表现与临床分期相关的鼻咽癌干细胞特异性miR 200b活性显著低表达趋势.结论 鼻咽癌干细胞以CD133为特征性标志物,miR-200b活性特异性低表达甚至不表达,可能是其特征性分子靶点。

Expressive characteristics of miRNAs in nasopharyngeal carcinoma stem cells

Objective To investigate the expressive characteristics of miRNAs in nasopharyngeal carcinoma stem cells. Methods Nasopharyngeal carcinoma （NPC） stem cells were＇separated and amplified from NPC cedll line CNE2 at first. Then, flow cytometry （FCM） was carried out to determine the expressive activities of biomarkers CD133 and CD44 on cancer stem cells （CSes） of NPC, in a comparative way with that on NPC cell lines 6 10B, 5-8F, CNE1, CNE2 and HONE1, with nasopharyngeal epithelial cell line NP69 as normal control. Followed was qRT-PCR assay of tumopassociated miRNAs. Meanwhile, on the basis of expressive activity detection of CD133 and CD44 in paraffin imbedded tissue samples of primary locus cases in different clinical staging by immunohistochemistry, qRT PCR assay was used to detect the expressive characteristics of NPC specific miR-200b in freshly taken tissue samples of primary lesion and then we analyzed their correlation with clinical staging. Results Prominent feature of CSC biomarker on NPC stem cells was CD133 expression, with much higher ~ensitivity and specificity when CD44 detection was combined to differentiate CSCs from other ceils. Among the tumor-associated miRNAs detected here, significantly reduced, even missing, expression of miR-200b was the most specific biological phenomenon for NPC stem ceils, still remarkably lower than that of NPC cell line 5-8F with highly metastatic potentiality. It was also true to show such a CSC biomarker expressive feature in tissue samples of untreated NPC with CD133 expression as determined by immunohistochemistry. At the same time, the characteristic expressive model of NPC specific miR-200b was found in a siginificantly redued way in freshly taken untreated NPC tissue samples, showing a tendency specifically correlated with patient＇s clinical staging in a linear cori＇elative way as well. Conclusions NPC stem ceils are in a staus of significantly reduced, even missing, expression of miR-200b, with CD133 as their significantly specific biomarker. The