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目的:分析比较人大肠癌不同转移潜能细胞株的差异表达蛋白质图谱,筛选并探讨肿瘤转移相关蛋白与大肠癌发生发展转移的关系。方法:利用双向凝胶电泳(2-DE)联合基质辅助激光解析离子化-飞行时间-质谱(MALDI- TOF-MS)技术,分析人大肠癌高转移性细胞株SW620和低转移性细胞株SW480差异表达蛋白质图谱,查询数据库筛选可能的大肠癌转移相关蛋白质并进行免疫组化和Western免疫印迹验证。结果:MalenieⅢ软件分析结果显示2-DE图谱重复性、匹配性较好。两种细胞株有25个蛋白点差异表达(SW620 14个,SW480 11个),经质谱分析及数据库查询鉴定有9个蛋白可能与转移相关。其中AnnexinⅠ、β-半乳糖苷结合蛋白1、钙调蛋白、主要组织相容性复合物Ⅱ类抗原、cofilin、超氧化物歧化酶和Rab相关GTP结合蛋白仅在SW480中表达,可能对大肠癌转移起抑制作用;而人硫氧还蛋白、泛素偶联酶仅在SW620中表达,可能对大肠癌转移起促进作用。选择其中感兴趣的AnnexinⅠ进行验证,其在细胞株的表达结果与2-DE一致。AnnexinⅠ在大肠正常组织不表达,在大肠癌无转移组和转移组中表达上调,三组表达差异具有显著性(H=19.801,P =0.000)。表达与大肠癌淋巴结转移呈正相关性(Z=-2.554,P=0.011),但表达与大肠癌不同组织学类型(H=0.678,P =0.839)和不同病理分级(H=0.470,P=0.747)均无相关性。大肠癌转移组中原发癌和淋巴结转移癌表达差异无显著性(Z=0.737,P=0.461)。结论:大肠癌转移是多种蛋白质功能共同作用的结果,不同转移潜能细胞株SW620和SW480蛋白质图谱明显差异表达。其中AnnexinⅠ蛋白可作为反映大肠癌生物学行为和判断预后的重要指征。
OBJECTIVE: To analyze and compare differentially expressed protein profiles of different metastatic potential human colorectal cancer cell lines, and to screen and explore the relationship between tumor metastasis-associated proteins and the development and metastasis of colorectal cancer. Methods: Two-dimensional gel electrophoresis (2-DE) combined with matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) was used to analyze the expression of SW620 in human colorectal cancer cell line SW620 and SW480 Differentially expressed protein profiles were screened by database for possible colorectal cancer metastasis-associated proteins and verified by immunohistochemistry and Western immunoblotting. Results: Malenie Ⅲ software analysis showed 2-DE pattern repeatability, better matching. There were 25 protein spots differentially expressed in the two cell lines (14 SW620 and 11 SW480). Nine proteins identified by mass spectrometry and database query may be related to metastasis. Annexin Ⅰ, β-galactoside binding protein 1, calmodulin, major histocompatibility complex class Ⅱ antigen, cofilin, superoxide dismutase and Rab-related GTP-binding protein were only expressed in SW480, Transfer inhibition; and human thioredoxin, ubiquitin-coupled enzyme is only expressed in SW620 may promote the metastasis of colorectal cancer. The Annexin I of interest was selected for verification, and its expression in cell lines was consistent with that of 2-DE. Annexin I was not expressed in the normal tissues of the large intestine, but was up-regulated in the non-metastatic and metastatic groups of colorectal cancer. There was a significant difference between the three groups (H = 19.801, P = 0.000). The expression was positively correlated with the lymph node metastasis of colorectal cancer (Z = -2.554, P = 0.011), but not with histological type (H = 0.678, P = 0.839) ) Are not relevant. There was no significant difference in the expression of primary cancer and lymph node metastasis between the metastasis group and the metastasis group (Z = 0.737, P = 0.461). CONCLUSION: Colorectal cancer metastasis is the result of a combination of multiple protein functions. The protein profiles of SW620 and SW480 in different metastatic potential cell lines are significantly different. Annexin Ⅰ protein can be used as an important indicator to reflect the biological behavior of colorectal cancer and determine the prognosis.