论文部分内容阅读
为筛选鼻咽癌(nasopharyngeal carcinoma,NPC)的分子标志物,采用激光捕获显微切割技术(laser capture microdissection,LCM)分别从NPC组织和正常鼻咽上皮组织(normal nasopharyngal epithelial tissue,NNET)中切割纯化NPC细胞和正常鼻咽上皮细胞(normal nasopharyngal epithelial cells,NNEC),应用二维凝胶电泳(two-dimensional electrophoresis,2-DE)分离LCM纯化细胞的蛋白质,图像分析识别差异表达的蛋白质点,基质辅助激光解吸电离飞行时间质谱(MALDI-TOF-MS)和电喷雾电离串联质谱(ESI-Q-TOF-MS)鉴定差异蛋白质点,Western blot检测差异蛋白cytokeratin8(CK8)在LCM纯化的NPC细胞和NNEC以及具有不同分化程度或转移潜能的4株NPC细胞中的表达,免疫组织化学检测CK8在63例NPC、28例NNET及20例颈淋巴结转移NPC组织中的表达水平.建立了LCM纯化的NPC细胞和NNEC的2-DE图谱,质谱鉴定了29个差异蛋白质,其中15个蛋白质只在NPC表达或表达明显增高,14个蛋白质在NPC中表达下调或缺失;Western blot结果显示,CK8的表达水平在NPC中较NNET明显下调,并与NPC细胞株的分化程度和转移潜能有关;免疫组织化学结果显示,CK8在NPC组织中的表达较NNET明显下调,在颈淋巴结转移NPC中的表达较原发NPC明显上调.研究结果提示,CK8与NPC分化及淋巴结转移相关,有望成为预测NPC转移和区别NPC分化程度的分子标志物.
To screen the molecular markers of nasopharyngeal carcinoma (NPC), they were cut from normal nasopharyngeal epithelial tissue (NNET) and normal NPC using laser capture microdissection (LCM) Purified NPC cells and normal nasopharyngeal epithelial cells (NNECs) were isolated and purified by two-dimensional electrophoresis (2-DE). The differentially expressed proteins were identified by image analysis. Differential protein spots were identified by MALDI-TOF-MS and ESI-Q-TOF-MS. Western blot was used to detect the expression of cytokeratin 8 (CK8) in LCM-purified NPC cells And NNEC, as well as in four NPC cells with different differentiation degree or metastatic potential.The expression of CK8 in 63 NPC, 28 NNET and 20 NPC patients was detected by immunohistochemistry.The LCM purified NPC cells and NNEC 2-DE map, mass spectrometry identified 29 differential proteins, of which 15 proteins were only expressed or significantly increased in NPC, 14 Western blot results showed that the expression of CK8 was significantly downregulated in NPC compared with NNET and correlated with the differentiation degree and metastatic potential of NPC cell lines.Immunohistochemistry showed that CK8 was positively expressed in NPC tissues The expression of CK8 was significantly up-regulated in primary NPC compared with that in primary NPC (P <0.05) .The results suggest that CK8 is associated with NPC differentiation and lymph node metastasis, which is expected to become a molecular marker for predicting NPC metastasis and differentiating NPC.