SELDI技术分析肾母细胞瘤体外和体内分泌蛋白质的表达

摘要

目的 运用表面增强激光解析电离飞行时间质谱(SELDI-TOF-MS)技术分析肾母细胞瘤细胞株G401细胞培养液与正常胚肾细胞株CCC-HKE-1细胞培养液的差异蛋白质,分析G401利用血清中的蛋白质,并比较G401培养液与患儿血清存在的相似蛋白质.方法 培养含10%胎牛血清及无血清的G401细胞和含10%胎牛血清的胚肾细胞株CCC-HEK-1细胞,收集以上三种培养液及G401裂解液,收集肾母细胞瘤患儿术前血清.将以上样本应用于WCX2(弱阳离子交换)芯片,用SELDI-TOF-MS技术检测、阅读和分析各组间差异及相似蛋白质.结果 G401和CCC-HEK-1培养液间存在显著的蛋白质差异表达;G401分泌的蛋白质质荷比有:3996、4084、4144、4215、4232、5980、6069、6094、6855、8553Da;G401利用血清中的蛋白质质荷比有:3830、3917、4144、4460、4795、4932、5688、5981、6023、6050、6075、6094、6188、6203、6857、7064、7308、7964 Da;质荷比为4082、4143Da的蛋白质在肿瘤细胞培养液和患儿血清中都检测到.结论 SELDI-TOF-MS技术应用于体外培养的肾母细胞瘤细胞和患儿血清方法简便、敏感性高、重复性好;从而为从蛋白质水平研究肾母细胞瘤的发病机制和治疗靶位的寻找奠定基础.%Objective To analyze the differentially expressed protein between cell culture medium of Wilms' tumor cell line G401 and normal embryonic kidney cell line CCC-HEK-1,and compare the similar protein between cell culture medium of (3401 and patients" serum by SELDI-TOF-MS technique.Methods G401 and CCC-HEK-1 with 10% FCS,G401 without FCS and the lysate of G401 were collected.The serum of Wilms tumor patients before operation was gathered in WCX2 proteinchip.The difference and similar proteins were detected and analyzed by SELDI-TOF-MS technique.Results Significantly different proteins were detected between cell culture medium G401 and CCCHEK-1.G401 secretory proteins with molecular weight (MW,Da) were 3996,4084,4144,4215,4232,5980,6069,6094,6855,and 8553.G401 consumed serum proteins with MW (Da) were 3830,3917,4144,4460,4795,4932,5688,5981,6023,6050,6075,6094,6188,6203,6857,7064,7308 and 7964.Protein with MW (Da) of 4082 and 4143 were expressed in both G401 cell culture medium and the serum of Wilms tumor cases.Conclusions The advantages of SELDI-TOF-MS technique are simple,high sensibility and repeatable in vitro and in vivo.It's useful to investigate the pathogenesis and therapeutic targets of Wilms' tumors.