应用双向电泳和质谱技术筛选口腔扁平苔藓差异蛋白

Screening of differential expression proteins from human oral lichen planus and normal oral mucosa by two-dimensional difference gel electrophoresis and mass spectrometry

目的:筛选口腔扁平苔藓及正常黏膜组织的差异表达蛋白,为口腔扁平苔藓的发病机制及诊断治疗的研究提供实验室依据。方法:收集口腔扁平苔藓组织及正常口腔黏膜组织,提取组织蛋白,蛋白定量,进行双向电泳,图象分析寻找差异点,基质辅助激光解吸飞行时间质谱(MALDI-TOF-MS)分析鉴定表达差异量较大的蛋白质点,确定所分析的蛋白质类型和功能。结果:(1)人口腔扁平苔藓及正常黏膜组织的双向电泳图谱的平均蛋白质点数分别为1576±67和1608±73,同一组织凝胶在蛋白质点位置上重复性较好。(2)通过比较人口腔扁平苔藓与正常黏膜组织的双向凝胶电泳图谱,得到差异表达蛋白质点数为13个,其中7个点在口腔扁平苔藓中为高表达,6个点在口腔扁平苔藓中为低表达。选择10个表达差异量较大的蛋白质点进行质谱和生物信息学分析,鉴定了其中的4个点,包括有锰超氧化物歧化酶(Mn-SOD),膜联蛋白I,波形蛋白,未知蛋白等。结论:锰超氧化物歧化酶,膜联蛋白I,波形蛋白,未知蛋白可能参与了口腔扁平苔藓的发生和发展,其相关机制尚待进一步阐明。双向电泳-质谱分析技术为口腔扁平苔藓发生发展过程中差异表达蛋白的研究提供了有效的技术手段。

Objective： To find biomarkers for oral lichen planus by comparing differential expressing proteins. Methods：10 cases of oral lichen planus and normal oral mucosa tissues were collected. Total protein was extracted; differential proteome profiles were established and analyzed by two-dimensional polyacrylamide gel electrophoresis （2D-PAGE） and matrix-assisted laser desorption/ionization time of flight mass spectrometry （MALDI-TOF-MS）. Results： （ 1 ） The well-resolved, reproducible 2-DE patterns of oral lichen planus and normal oral mucosa were obtained. The results showed that average protein spots were 1 576 ± 67 and 1 608 ± 73 in oral lichen planus and normal oral mucosa respectively, （2） The 13 differential protein spots were identified by Imaging Master 2D image analysis software between oral lichen planus and normal oral mucosa. There were 7 protein spots in oral lichen planus were higher than those in normal oral mucosa, 6 protein spots in oral lichen planus were lower than those in normal oral mucosa. 10 differential expressing proteins were analyzed by mass spectrometry and bioinformation. 4 of them were well characterized including manganese superoxide dismutase （ Mn-SOD ） , Annexin I, vimentin and unknown proteins. Conclusion： Differential expression proteins might be candidate biomarkers for diagnosis of oral lichen planus ; and proteomic technique is valuable for screening the diagnostic biomarkers.