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为分析结肠上皮癌变进程中的差异表达蛋白质,筛选结肠癌诊断候选标志物,先采用激光捕获显微切割技术(LCM)纯化人正常结肠组织和结肠癌组织,再采用同位素标记对蛋白质进行相对和绝对定量(i TRAQ),然后结合强阳离子柱分离和反相液质联用(2D-LC-MS/MS)鉴定差异表达蛋白质。试验共鉴定了1 042个非冗余蛋白,筛选出与人结肠癌变相关的差异蛋白162个,其中在结肠癌中表达上调的蛋白质68个,表达下调的蛋白质94个,功能分析表明,这些差异蛋白质涉及内质网蛋白质加工、糖酵解/糖原异生、黏着斑细胞连接、调节微丝组装和细胞外基质受体相互作用等分子功能。进一步采用Western blotting及免疫组织化学技术(IHC)验证了其中差异蛋白S100A9的表达,证实了定量蛋白质组学结果的可靠性。研究表明:差异表达蛋白质与结肠癌变相关,并有可能成为结肠癌诊断的候选分子标志物,为进一步深入研究这些蛋白质在结肠癌发生和发展进程中的分子机制,为结肠癌的发病机制研究提供新线索。
To analyze differentially expressed proteins in the course of carcinogenesis of colorectal epithelium, candidate diagnostic markers for colon cancer were selected. Purified human normal colorectal and colon cancer tissues were obtained by laser capture microdissection (LCM) Absolutely quantified (i TRAQ) was then used to identify differentially expressed proteins in combination with strong cation column separation and reverse phase LC-MS (MS-MS / MS). A total of 1 042 non-redundant proteins were identified and 162 differentially expressed proteins were detected in human colon cancer. Among them, 68 proteins were up-regulated in colon cancer and 94 proteins were down-regulated. Functional analysis showed that these differences Proteins are involved in the molecular functions of endoplasmic reticulum protein processing, glycolysis / gluconeogenesis, focal cell attachment, regulating assembly of microfilaments, and extracellular matrix receptor interactions. Western blotting and immunohistochemistry (IHC) were used to verify the expression of S100A9, which confirmed the reliability of quantitative proteomics results. Studies have shown that differentially expressed proteins are correlated with colon cancers and may be potential molecular markers for the diagnosis of colon cancer. To further investigate the molecular mechanisms of these proteins in the development and progression of colon carcinomas, we provide evidence for the pathogenesis of colon cancer New clues.