正常脊髓室管膜与脊髓室管膜瘤的差异蛋白组学研究

Identification of discrepant proteins between normal ependyma and ependymoma of spinal cord by using proteomics technology

目的 应用双向凝胶电泳（2-DE）和基质辅助激光解吸离子电离飞行时间质谱（MAL-DI-TOF-MS）的蛋白组学方法,观察正常脊髓室管膜与脊髓室管膜瘤的差异表达蛋白质.方法 收集7例脊髓室管膜瘤标本作为肿瘤组,4例尸解得到的脊髓室管膜标本作为对照组.提取各组标本的蛋白质,定量后进行IEF和SDS-PAGE双向电泳分离,得到的2-DE图谱经染色后,通过软件找到差异蛋白点并且使用MALDI-TOF-MS技术进行鉴定,部分差异蛋白质进行Western blot验证.结果 成功鉴定出50个有效的差异蛋白质点（其中肿瘤组较对照组上调24个,下调13个,仅在肿瘤组中表达10个,仅在对照组表达3个）.Western blot实验证实了差异蛋白波形蛋白（Vimentin）、膜联蛋白Ⅰ（Annexin Ⅰ）和HSPA8在肿瘤组中的高表达.结论 应用2-DE串联MALDI-TOF-MS技术能够建立具有高分辨率及可重复性的正常脊髓室管膜与脊髓室管膜瘤2-DE图谱,并能有效鉴定差异蛋白质.其中的Vimentin、Annexin Ⅰ、HSPA8等蛋白可能与脊髓室管膜瘤的发病机制有关.

Objective To evaluate the differentially expressed proteins between ependymoma and normal ependyma of spinal cord by using two-dimension electrophoresis （2-DE） combined with MALDI-TOF-MS proteomics technology. Methods Seven spinal ependymoma tissues were obtained through operations and four nomal ependyma tissues were obtained through autopsy. The total proteins in the samples were extracted, quantitated, and subjected to 2-DE, quick Coomassie brilliant blue staining, discrepant protein spots selection, spot cutting, protein enzymolysis and MALDI-TOF-MS. The differentially expressed proteins were obtained and some of them were verified by Western blotting. Results Fifty effective discrepant protein spots were selected. Of the total 50 protein spots, 24 protein spots were up-regulated and 13 were down-regulated in tumor group as compared with the control group. Total effective discrepant protein spots were carried out by MALDI-TOF MS and MASCOT database searching Western blotting test verified that vimentin, annexin Ⅰ and HSPA8 were up-regulated in tumor group as compared with the control group.Conclusion 2-DE combined with MALDI-TOF-MS proteomics technology can select discrepant proteins between spinal ependymoma and normal ependyma tissues. Bioinformatical analysis indicated that vimentin, annexin Ⅰ and HSPA8 was probably related to origin of spinal ependymoma.