SELDI技术分析体外培养不同肝细胞株差异蛋白的表达

目的: 运用表面增强激光解吸/离子化/飞行时间质谱(SELDI-TOF-MS) 技术,检测体外培养的肝癌细胞株(HepG2),转染乙肝病毒的肝癌细胞株(HepG2.2.15)与正常肝细胞株(LO2)蛋白质的差异表达,筛选肝细胞癌的标志蛋白,为进一步研究肝癌发病的蛋白质组学机制奠定基础. 方法: 常规培养上述3种细胞,细胞状态良好时收集细胞,裂解细胞后采用 SELDI-TOF-MS技术用WCX2芯片检测HepG2, HepG2.2.15, LO2细胞内蛋白质组学的差异表达. 结果: WCX2芯片共捕获91个蛋白,在Mr 5000～15 000区段,与正常肝细胞株比较,有7个蛋白质在两种肝癌细胞中出现明显变化,其中2个蛋白在肝癌细胞中表达量增高,5个蛋白在肝癌细胞中表达量降低;另外两种肝癌细胞株也有其各自的标志蛋白,9个蛋白分子在HepG2表达量增高,10个蛋白分子在HepG2.2.15表达量增高. 结论: SELDI蛋白芯片技术检测可作为检测肝癌早期生物标记的方法,它简便,敏感性高,重复性好,本文发现的这些组织特异性蛋白生物标记对肝癌的早期诊断有潜在的应用价值,对我们从血清或组织标本中筛选和鉴定不同型别肝癌细胞之间的标志蛋白有重要意义,从而为从蛋白质水平研究肝癌的发病机制及新的治疗靶位的寻找奠定了一定的基础.

Analysis of different protein expressions in different liver cell strains in vitro using SELDI

AIM: To examine the differential expressions of proteins in hepatoma cell line (HepG2),hepatoma cell line transfected by HBV (HepG2.2.15)compared with normal liver cell line(LO2) by using surface-enhanced laser desorption/ionization time-of-flight mass spectrometry (SELDI-TOF-MS),so as to establish a foundation for further studying on the mechanism of hepatoma at the protein level. METHODS: The 3 types of cells above mentioned were cultured as general and collected when they were in good conditions. After cell disruption,SELDI-TOF-MS was employed to detect the differential expressions of proteomes of HepG2,HepG 2.2.15 and LO2. RESULTS: Ninety-one proteins were captured by WCX2 array. Compared with normal liver cell lines,in the segments from M_r 5000 to 15 000,proteins in the HepG2 and HepG2.2.15 showed apparent changes,in which 2 proteins' expressions were increased in carcinoma cells,the other 5 decreased. Nine proteins in HepG2 were increased and 10 proteins in HepG 2.2.15 were increased. CONCLUSION: The SELDI ProteinChip technology can be a desired method in detecting the biological markers in the earlier period of hepatoma. This method is of convenience,high sensitivity,and good reproducibility. The biological tags of tissue-specific proteins we found have a potential applying value in the early diagnosis of hepatoma. These tags are also meaningful in screening and identifying signal proteins from serum and tissue specimen in different types of hepatoma. Certain foundation has been established in studying the etiopathogenesis of hepatoma at the level of proteins,as well as the searching of new therapeutic target sites.