转导EB病毒转录的核内小RNA促进OSC-19细胞恶性化

Malignant transformation of a human oral squamous cell carcinoma cell strain OSC-19 by EBV encoded small RNAs

目的 :通过转导EB病毒 (EBV)转录的核内小RNA (EBERs)探讨其对舌癌细胞株OSC 19细胞生长特性的影响。方法 :EBERs基因编码区序列是利用聚合酶链反应 (PCR)技术从EB病毒阳性的鼻咽癌 (NPC)细胞株NPC KT细胞中扩增出。为了在细胞内大量表达EBERs,利用分子生物学技术制作了EBERs片段的 4次重复排列 4EBERs和 10次重复排列 10EBERs,分别亚克隆到 pCEP4载体质粒并转染OSC 19细胞。利用North ernBlot法确定细胞内EBERs的表达后 ,对EBERs转染后的OSC 19细胞在培养中的生长特性、软琼脂中的细胞克隆的形成进行了观察。结果 :在培养中大量表达EBERs的OSC 19细胞之间丧失了细胞间的相互连接 ,细胞呈纺锤状 ,并在软琼脂中形成大量细胞克隆。而非表达EBERs的OSC 19细胞没有发现细胞生长特性的改变 ,在软琼脂中没有形成大量细胞克隆。结论 :EBERs的大量表达可以导致OSC 19细胞的分散活性和基质非依赖性 ,提示EBERs在上皮细胞株OSC 19细胞中有致癌作用 ;大量表达EBERs的OSC 19细胞为进一步研究EBERs的作用提供了一个有用的上皮细胞模型。

Objective:To observe the growth characteristics of the EBERs-transfected OSC (oral squamous carcinoma)-19 cells, through generated overexpression of EBERs in OSC-19 cells.Method:EBERs fragment were amplified by PCR from EBV-positive NPC-KT cell DNA. Using molecular biologic technique, we constructed the EBERs-expressing plasmids,which contained , 4 or 10 tandem repeats of the EBERs. The OSC-19 cells were transfected with those plasmids, respectively. We demenstrated the overexpression of EBERs in EBERs-transfected OSC-19 cells by Northern Blot analysis, and compared the growth characteristics in culture and colony formation in soft agar of Hyg r- and EBERs-transfected OSC-19 cells.Result:10EBERs-transfected OSC-19 cells lost cell-to-cell contacts and acquired long spindle-shape morphology in culture and formed colonies in soft agar. However, parental and Hyg r-transfected OSC-19 cells scarcely did.Conclusion:In this study, we established 10EBERs-transfected OSC-19 cells, which overexpressing the EBERs and showed that it has a scattering activity in culture and a role in anchorage-independent growth in soft agar.These results indicat that EBERs has a role of malignant transformation in OSC-19 cells. However, the 10EBERs-transfected OSC-19 cell provides a useful epithelial model for studying the function of EBERs.