侵袭性与非侵袭性垂体腺瘤的蛋白质差异表达谱的建立

摘要

目的:建立分辨率高和重复性好的侵袭性与非侵袭性垂体腺瘤的双向凝胶电泳图谱,并识别鉴定出其差异表达的蛋白质.方法:利用固相pH梯度双向凝胶电泳分离侵袭性与非侵袭性垂体腺瘤的总蛋白质,凝胶经银染显色后,运用ImageMaster 2D图像分析软件进行比较分析、识别差异表达的蛋白质,应用基质辅助激光解吸电离飞行时间质谱(MALDI-TOF-MS)及数据库搜索鉴定部分差异蛋白质点.结果:得到了分辨率较高、重复性较好的侵袭性与非侵袭性垂体腺瘤的双向凝胶电泳图谱,将同一侵袭性垂体腺瘤组织和非侵袭性肿瘤组织分别进行了3次双向凝胶电泳,3块凝胶的蛋白质点数分别为1 080±24和1 035±28,匹配点数分别为975±45和91 8±56,匹配率分别达90.3%和88.7%.同一组织的3块胶在蛋白质点位置上有较好的重复性,不同胶间蛋白质点在等电聚焦(IEF)方向的偏差是(1.563±0.259)mm,在十二烷基磺酸钠聚丙烯酰胺凝胶电泳(SDS-PAGE)方向上的偏差为(1.088±0.206)mm.比较分析侵袭性垂体腺瘤和非侵袭性垂体腺瘤凝的双向凝胶电泳图谱,发现两者间的99个表达有明显差异的蛋白质点.初步对其中30个差异蛋白质点进行了肽质量指纹图分析,鉴定出一些与细胞周期调控、信号转导等有关的蛋白质.结论:建立了分辨率较高且重复性较好的侵袭性与非侵袭性垂体腺瘤的双向凝胶电泳图谱,并识别鉴定出一些侵袭性与非侵袭性垂体腺瘤之间差异表达的蛋白质,为进一步筛选侵袭性垂体腺瘤的蛋白质表达数据库提供了基础.%Objective To establish high resolution, reproducible 2-dimensional electrophoresis (2-DE) profiles of invasive and non-invasive pituitary adenoma tissues and to identify differentially ex-pressed proteins between the invasive and non-invasive tissues. Methods The proteome from invasive and non-invasive pituitary adenomas tissues was dissected and analyzed by: (1) immobilized pH gradient two-dimensional polyacrylamide gel electrophoresis, (2) silver staining, (3) imageMaster 2-D software analysis, (4) peptide mass fingerprint based (PMS) on matrix-assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF-MS), and (5) database comparison. Results High-resolution 2-D patterns of invasive and non-invasive pituitary adenoma tissues were successfully produced and re-peated 3 times for each sample. An average of 1080±24 and 1035±28 spots were detected for invasive and non-invasive pituitary adenoma tissues, respectively. Additionally, 975±45 and 918±56 spots were found to have an average matching rate of 90.3% and 88.7% for invasive and non-invasive tissues, re-spectively. The spot positional deviation was (1.563±0.259) mm for IEF and (1.088±0.206) mm for SDS-PAGE. A total of 99 spots of differential expression were matched between the invasive and non-in-vasive pituitary adenoma tissues. Thirty differential proteins, some of which were involved in the regula-tion of cells cycle and signal transduction, were initially characterized by PMS. Conclusion The acquisi-tion of well-resolved and reproducible 2-D patterns of invasive and non-invasive pituitary adenoma tissues and the identification of differentially expressed proteins provides a proteome database for invasive pituita-ry adenomas.