LST-R1细胞差异表达蛋白的初步筛选和鉴定

Screening and Identifying Differentially Expressed Proteins in LST-R1 Cells

背景与目的:大肠侧向发育型肿瘤(colorectal laterally spreading tumor,CLST)生长方式为沿大肠粘膜浅表侧向扩散而少向肠壁深层侵袭,该特点与普通大肠癌的易深层侵袭高转移的特性迥异,因此CLST可作为大肠癌侵袭转移研究的对照病变模型。本研究应用蛋白质组学方法分析和鉴定CLST细胞株LST-R1和结肠肿瘤SW480细胞株及LoVo细胞株差异表达蛋白。方法:以2-DE分离三种细胞株总蛋白质,银染显色,进行差异蛋白质分析。采用MOLDI-TOF质谱对40个差异蛋白质斑点进行肽质谱指纹图(peptides mass fingerprinting,PMF)分析,最后在蛋白质数据库中搜索匹配的蛋白质。结果:使用pH4～7IPG胶条测得2-DE图谱蛋白质斑点数分别为1285±51、1184±47和1124±54;蛋白质上样量为150μg时分别获得989、935和893个蛋白质斑点。综合分析LST-R1、SW480和LoVo细胞株2-DE图谱,多数蛋白质斑点的分布和表达量相似,这表明CLST也表达一般大肠肿瘤所具有的蛋白质谱。LST-R1细胞和SW480细胞在pH4～7IPG胶条2-DE图谱间存在(96±7)个蛋白质差异点,其中(50±6)个点仅在LST-R1细胞株中表达或表达明显增强,(47±5)个点仅在SW480细胞中表达或表达明显增强;而在LST-R1细胞和LoVo细胞的2-DE图谱间存在(108±10)蛋白质差异点,其中(56±8)个点仅在LST-R1细胞株中表达或表达明显增强,(52±11)个点仅在LoVo细胞中表达或表达明显增强;三种细胞之间共鉴定出19个差异表达蛋白质。结论:筛选出19个CLST细胞株LST-R1相关蛋白质,这些蛋白可能在CLST病变浅表扩散形成和大肠癌粘附侵袭过程中发挥一定的作用。

BACKGROUND ＆ OBJECTIVE= Colorectal laterally spreading tumor （CLST） rarely invades deeply but always laterally spreads along the colorectal mucosa, therefore, CLST could be used as a comparative model in studying the invasion and metastasis of colorectal cancer （CRC）. This study was to identify differentially expressed proteins between a CLST cell line LST-R1 and two colorectal carcinoma cell lines SW480 and LoVo using proteomic technology. METHODS.. Total proteins of LST-R1, SW480 and LoVo cells were isolated by two-dimensional electrophoresis （2-DE）. Differentially expressed protein spots were analyzed with Melanie 3 software. The peptide mass fingerprints （PMFs） of differently expressed proteins were analyzed by MALDI-TOF mass spectrum. Subsequently matched proteins were searched through protein databases. RESULTS： Using pH4-7 IPG gels with 250 ug protein loading, the numbers of protein spots in 2-DE maps were 1285±51 in LST-R1 cells, 1184±47 in SW480 cells, and 1124±54 in LoVo cells; when with 150 ug protein loading, the numbers were 989, 935 and 893, respectively. The distribution and levels of these proteins in 2-DE maps of LST-R1, SW480 and LoVo cells were analogical which indicated CLST also expresses the protein profile of common colorectal tumors. In 2-DE maps, 96 ＋7 differential protein spots were detected between LST-R1 cells and SW480 cells with 50±6 only expressed or obviously over-expressed in LST-R1 cells and 47±5 in SW480 cells; 108±10 differential protein spots were detected between LST-R1 cells and LoVo cells with 56±8 only expressed or obviously over-expressed in LST-R1 cells and 52±11 in LoVo cells. Nineteen differentially expressed proteins were identified among LST-R1, SW480 and LoVo cells. CONCLUSION.. Nineteen differentially expressed proteins are possibly involved in laterally spreading of CLST and adhesion and invasion of CRC.