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目的:以荧光标记的二维差异凝胶电泳(2D-DIGE)等蛋白质组学技术分析结直肠癌(CRC)与正常结肠黏膜间的蛋白质表达差异,筛选新的肿瘤标志物。方法:以2D-DIGE及基质辅助激光解析飞行时间质谱(MALDI-TOF-MS)分析和鉴定6对新鲜的CRC与正常大肠黏膜组织的差异表达蛋白;以免疫组化法验证兴趣蛋白在46例CRC与正常大肠黏膜组织中的表达。结果:2D-DIGE分析和MALDI-TOF-MS鉴定结果显示,在CRC组织中表达明显升高的蛋白有泛醌细胞色素C还原酶核心蛋白1(UQCRC1),14-3-3ζ和结蛋白(desmin)等,明显降低的蛋白有碳酸酐酶(CA)I和CA II(均P<0.05)。免疫组化法结果显示,UQCRC1和14-3-3ζ在CRC中的表达强度为2.43±0.81和1.41±1.07,明显高于两者正常黏膜中的表达(1.80±0.96,0.67±0.94)(P<0.001),而desmin在两组织间表达差异无统计学意义(P>0.05);CA I及CA II在CRC中的表达强度为1.67±0.52和1.18±0.84,明显低于两者在正常黏膜中的表达强度(2.93±0.25,2.80±0.50)(P<0.001)。结论:以2D-DIGE为基础的蛋白组学技术结合免疫组化等方法验证是筛选肿瘤标志物的有效手段。UQCRC1,14-3-3ζ,CA I和CA II可能为CRC新的标志物或治疗靶点。
OBJECTIVE: To screen for differences in protein expression between colorectal cancer (CRC) and normal colonic mucosa by proteomic techniques such as 2D-DIGE with fluorescence labeling and to screen novel tumor markers. METHODS: Six pairs of fresh differentially expressed proteins of CRC and normal colorectal mucosa were analyzed and identified by 2D-DIGE and MALDI-TOF-MS. Immunohistochemistry was used to detect the protein of interest in 46 cases CRC and normal colorectal mucosa expression. Results: The results of 2D-DIGE and MALDI-TOF-MS showed that ubiquitin-cytochrome c reductase core 1 (UQCRC1), 14-3-3ζ and desmin desmin) and so on. Significantly decreased proteins were carbonic anhydrase (CA) I and CA II (all P <0.05). The results of immunohistochemistry showed that the expression intensity of UQCRC1 and 14-3-3ζ in CRC was 2.43 ± 0.81 and 1.41 ± 1.07, respectively, which was significantly higher than that in normal mucosa (1.80 ± 0.96,0.67 ± 0.94) (P (P <0.05). The expressions of CA I and CA II in CRC were 1.67 ± 0.52 and 1.18 ± 0.84, which were significantly lower than those in normal mucosa (2.93 ± 0.25, 2.80 ± 0.50) (P <0.001). Conclusion: 2D-DIGE-based proteomics combined with immunohistochemistry is an effective method to screen tumor markers. UQCRC1, 14-3-3ζ, CA I and CA II may be novel markers or therapeutic targets for CRC.