慢性鼻-鼻窦炎鼻息肉与正常鼻黏膜差异蛋白质的检测及其意义

Determination of differentially expressed proteins and it′s significance among chronic sinusitis, nasal polyps and normal nasal mucosa

目的 采用蛋白质组学技术筛选慢性鼻-鼻窦炎、鼻息肉和正常鼻黏膜组织之间差异表达的蛋白质，初步鉴定出慢性鼻-鼻窦炎和鼻息肉的候选蛋白质标记物。方法 应用固相pH4～7胶条行双向凝胶电泳，胶体考马斯亮蓝染色后，扫描2-DE胶，应用PDquest图像分析软件比较慢性鼻-鼻窦炎、鼻息肉和正常鼻黏膜组织2-DE图谱，得到三组之间差异表达的蛋白质点。经MALDI—TOF—MS鉴定后获得这些差异表达蛋白质的肽质量指纹图谱，用Mascot软件查询NCBInr和SWISS—PROT数据库，得出被测蛋白质的鉴定结果。结果 所获双向凝胶电泳图谱分辨率高、重复性好。测得鼻息肉、慢性鼻-鼻窦炎及正常鼻黏膜组织的蛋白质平均斑点数分别为1020±40、1112±10和1008±25；平均匹配率分别为（93±2）％、（95±1）％和（90±3）％。三组之间共计有13个明显差异表达的蛋白质点。初步筛选出角蛋白8和阿朴脂蛋白AI作为鼻息肉的候选标志物，以及PLUNC蛋白、超氧化物歧化酶、自然杀伤细胞促进因子B、垂体腺苷酸环化酶激活肽为慢性鼻-鼻窦炎的候选标志物。结论 用蛋白质组学技术能高通量筛选慢性鼻-鼻窦炎、鼻息肉和正常鼻黏膜间存在的差异表达蛋白质，这将为慢性鼻-鼻窦炎、鼻息肉分类、分型分期和预后判断标准寻找新的客观参考指标。

Objective To investigate the differentially expressed proteins among chronic sinusitis, nasal polyps and normal nasal mucosa by means of proteomic technology, and select the candidate biomarkers of chronic sinusitis and nasal polyps. Methods Proteins extracted from chronic sinusitis, nasal polyps and normal nasal mucosa were separated and the differentially expressed proteins were identified by series of proteomic tools, including immobilized pH4-7 gradient two-dimensional sodium dodecyl sulfate polyacrylamide gel electrophoresis, modified coomassie brilliant blue staining, images scanning by the Image Scanner apparatus, PDQuest analysis software, peptide mass fingerprinting based on matrix-assisted laser desorption ionization time of flight mass spectrometry （MALDI-TOF-MS） by in-gel digestion extract, and Mascot searching in NCBInr and SWISS-PROT databases. Results The 2-DE patterns with high resolution and reproducibility were obtained. The protein spots separated and visualized in chronic sinusitis, nasal polyps and normal nasal mucosa gel were 1020 ± 40, 1112 ± 10 and 1008 ± 25, respectively. And the match rates were （ 93 ± 2） % , （ 95 ± 1 ） % 和 （ 90± 3 ） % respectively. Thirteen differentially expressed spots were found from chronic sinusitis, nasal polyps and normal nasal mucosa gel. We selected and recommend Keratin 8 and APOA1 proteins as candidate biomarkers of nasal polyps, and PLUNC protein, PACAP protein, NKEF-B and SOD as candidate biomarkers of chronic sinusitis. Conclusions The differentially expressed proteins among chronic sinusitis, nasal polyps and normal nasal mucosa can be efficiently and relatively reliably identified via the techniques of proteomics. These techniques will play a very important role in the researches for new objective indicators possibly employed in the future classifying, staging and prognosis.