人类肝癌细胞HepG2、Bel-7402和Li-7植入裸鼠体内后微量蛋白表达分析

Expression of microproteins in nude mice injected with hepatoma cell lines of HepG2,Bel-7402 and Li-7

目的探讨Hep G2、Bel-7402和Li-7不同肝癌细胞系在裸鼠体内生长过程中血清蛋白谱的表达情况,寻求与肝癌早期诊断相关的微量蛋白标志物。方法将40只BALB/c A-nu裸鼠随机均分为4组,分别为Hep G2、Li-7和Bel-7402组以及对照组,对Hep G2、Bel-7402和Li-7三种癌细胞分别进行培养和传代,制备浓度为1.5×107/m L癌细胞悬液,在每组裸鼠皮下分别注射相应的癌细胞悬液0.2 m L,对照组注射RPMI1640细胞培养基,观察裸鼠的生长情况,于注射后20、40和60 d通过摘眼球法采集裸鼠血液样本,用表面增强激光解吸电离飞行时间质谱仪(SELDI-TOF-MS)检测微量蛋白谱,采用Ciphergen Protein Chip 3.0和Ciphergen Biomaker Wizard 3.1软件和统计学分析软件SPSS16.0对所得的数据进行分析,筛选出差异性蛋白。结果 Hep G2组将Hep G2细胞注射裸鼠后,在裸鼠血清中出现29个差异蛋白,规律变化明显的有8个,其中上调的有5个,下调的有3个。Li-7组将Li-7细胞注射裸鼠后,出现12个差异表达蛋白,其中表达增强的有7个,表达减弱的有5个。Bel-7402组将Bel-7402细胞注射裸鼠后,血清中出现5个差异蛋白,其中表达增强的有4个,表达减弱的有1个。结论不同类型的肝癌细胞在裸鼠体内生长过程中会产生不同的微量差异蛋白,这些蛋白经鉴定、检测开发有望成为临床不同细胞类型肝癌早期诊断的蛋白标志物。

Objective To investigate the serum proteinogram of hepatoma cell lines（ Hep G2,Bel-7402 and Li-7）which grow in the nude mice in order to find the microprotein markers which are associated with early diagnosis of hepatoma. Methods We classified 40 BALB/c A-nu nude mice into 4 groups randomly and marked them as Hep G2 group,Li-7group,Bel-7402 group and control group,respectively. Three types of hepatoma cell lines（ Hep G2,Bel-7402 and Li-7）were cultured and the cells were prepared into cell suspension（ 1. 5 × 107/ml）,respectively. We injected the cell suspension into the relevant groups with 0. 2 m L in every mouse and the control group was injected with RPMI1640 medium.These nude mice were observed for 2 months and we got their blood on the 20 th,40th,60 th day by eyeball extirpating. We detected the proteinogram using surface enhanced laser desorption/ionization time of flight mass spectrometry（ SELDI-TOFMS） and then,we analyzed the data by Ciphergen Protein Chip 3. 0 and Ciphergen Biomaker Wizard 3. 1 and SPSS 16. 0 to screen the differentiated proteins. Results After the mice in Hep G2 group were injected with cell line Hep G2,there were29 differentiated proteins. Among them,8 proteins changed significantly and 5 proteins were up-regulated and 3 were downregulated. After the mice in Li-7 group were injected with cell line Li-7,there were 12 differentiated proteins. Among them,7 proteins enhanced,and 5 proteins weakened. After the mice in Bel-7402 group were injected with cell line Bel-7402,there were 5 differentiated proteins. Among them,4 proteins enhanced and 1 protein weakened. Conclusions Different subtypes of hepatoma cell lines injecting into nude mice can express different microproteins in the serum of mice.These proteins could be identified and detected to be promising markers of different subtypes of hepatoma for the early diagnosis.