不同转移潜能肝癌细胞株的差异蛋白质组的二维液相色谱分析  

Proteome analysis of hepatocellular carcinoma cell lines with different metastasis potentials by two-dimensional liquid chromatography

作　　者：彭艳[1] 张霞[2] 王葵[3] 蒋平[1] 焦炳华[1] 倪健 

机构地区：[1]第二军医大学基础医学部生物化学与分子生物学教研室,上海市200433 [2]上海交通大学药学院,上海市200240 [3]第二军医大学东方肝胆外科医院,上海市200438 [4]上海晨健抗体组药物有限公司,201203

出　　处：《世界华人消化杂志》2007年第13期1482-1487,共6页World Chinese Journal of Digestology

基　　金：上海市科学技术委员会资助课题;No.054319938~~

摘　　要：目的:对不同转移潜能肝癌细胞株MHCC97- H(高转移)和MHCC97-L(低转移)差异表达的蛋白质进行二维液相色谱分离和MALDI-TOF质谱鉴定.方法:将肝癌细胞株MHCC97-H和MHCC97- L细胞裂解样品按蛋白质PI进行一维的色谱聚焦分离,然后每个PI组分再按疏水性经二维反相无孔硅胶HPLC分离,利用ProteoVue软件将UV光吸收图谱转换成PI对疏水性的胶图,再利用DeltaVue软件比较升高或降低的差异蛋白.收集差异蛋白峰进行胰酶酶解,然后进行MALDI-TOF质谱鉴定.结果:2D图谱显示共有72个差异蛋白条带,共鉴定出了9个差异蛋白.分别为M2型丙酮酸激酶、ATP合成酶α亚单位、热休克蛋白60、Toll样受体9、含黄素单加氧酶、钙网硬蛋白前体、锰超氧化物岐化酶、nm23-H1、G-蛋白偶连受体激酶5;其中4个蛋白在高转移细胞株MHCC97-H中表达升高,5个蛋白在低转移细胞株MHCC97-L表达升高.结论:这些差异蛋白可能在肝癌的转移中起关键作用.AIM： To analyze the proteome of hepatocellular carcinoma cell lines, MHCC97-H （high metastasis） and MHCC97-L （low metastasis）, with different metastasis potentials by twodimentional liquid phase chromatographic fractionation followed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry （MALDI-TOF/MS）. METHODS： Cell lysates of MHCC97-H and MHCC97-L were fractionated according to PI using chromatofocusing with analytical columns in the first dimension, followed by separation of the proteins in each PI fraction using nonporous reversed-phase high performance liquid chromatography （HPLC）. A two-dimensional map of the protein content for each cell line based upon pI versus hydrophobicity as detected by UV absorption was generated and a differential display map indicating the presence of up-or down-regulated proteins was exhibited using ProteoVue and DltaVue software. The different proteins were TOF/MS was digested by trypsin and MALDITOF/MS was performed. RESULTS： Image analysis of two-dimensional map revealed 72 differentially expressed proteins between MHCC97-H and MHCC97-L cells, and 9 protein peaks （including M2 pyruvate kinase, α-subunit of ATP synthase, heat shock protein 60, Toll-like receptor 9, flavoprotein oxygenase, pro-calreticulin, manganese superoxide dismutase, nm23-H1, and G-protein-coupled receptor kinase 5） were further identified by tryptic digestion, peptide mass fingerprinting and mass spectrometry. Four proteins were up-regulated while 5 were down-regulated in MHCC97-H cells. CONCLUSION： The identified differentially expressed proteins may play important roles in the metastasis of hepatocellular carcinoma.

关 键 词：蛋白质组 肝细胞癌 肿瘤转移 二维液相 色谱分离 MALDI-TOF质谱 

分 类 号：R735.7[医药卫生—肿瘤]