基于液质联用的胃癌血清中O-糖组学分析

发布时间：2019-03-09 20:35

【摘要】：目的：运用糖组学分析方法,比较胃恶性肿瘤患者和正常对照血清样本中O-glycan的表达情况,建立血清O-glycan数据库,找出差异性表达的O-glycan。从而找到胃恶性肿瘤相关的特征性O-glycan,为胃恶性肿瘤的诊断提供新的靶点,评估其诊断价值。并为其他肿瘤血清样本中O-glycan的研究提供分析基础。方法：1)利用非还原性方法释放157例胃恶性肿瘤患者和144例正常对照血清样本中O-glycan,并用PMP标记；2)通过SPE柱层析方法纯化O-glycan; 3) O-glycan在LC-MS和LC-MS/MS的正离子模式下进行定性和定量分析；4)对胃恶性肿瘤患者和正常对照血清样本中O-glycan进行提取离子流,并进行相对定量；5)利用偏最小二乘法(PLS-DA)、t检验、受试者工作特征曲线(ROC)分析所有O-glycan对应的提取离子流强度；6)分析O-glycan与临床上使用的肿瘤标志物之间的相关性；7)利用免疫组化的方法分析在胃恶性肿瘤组织和对应的癌旁组织中MUC1以及Tn-MUC1的表达。结果：1) O-glycan Corel、Core2、ST-antigen和Core2合成的复杂糖链(m/z733.33、m/z 809.42)在胃恶性肿瘤血清中表达上调(P0.0001); 2) Core3、m/z 529.75、diST-antigen在胃恶性肿瘤血清中显著下调(P0.001)；3)通过LC-MS/MS分析清楚上述离子的结构信息；4)通过Pearson相关性分析,得出血清中O-glycan与血清中分泌型CEA和MUC1相关性显著；5)通过免疫组化得出胃恶性肿瘤组织中MCU1显著上调及糖基化异常。结论：在本研究中通过非还原性解离和PMP标记,并用LC-MS和LC-MS/MS能够实现对血清中O-glycan的定性和定量分析；并且经过统计分析,证明了157例胃恶性肿瘤患者和144例正常对照血清中O-glycan的差异,且血清中的O-glycan和糖蛋白(MUC1、CEA)相关性显著；并且这些糖蛋白在胃恶性肿瘤组织中的糖基化修饰也发生异常。
[Abstract]:Aim: to compare the expression of O-glycan in serum samples of patients with gastric malignant tumor and normal controls by means of glycomorphological analysis, and to establish a serum O-glycan database to find out the differentially expressed oglycan. It provides a new target for the diagnosis of gastric malignant tumor and evaluates its diagnostic value. It also provides an analytical basis for the study of O-glycan in serum samples of other tumors. Methods: 1) Oglycan was released from serum samples of 157 patients with gastric malignant tumor and 144 normal controls by non-reductive method and labeled with PMP. 2) purification of O-glycan by SPE column chromatography; 3) qualitative and quantitative analysis of O-glycan in positive ion mode of LC-MS and LC-MS/MS; 4) O-glycan was extracted from the serum samples of patients with gastric malignant tumor and normal controls, and the relative quantitative analysis was carried out. 5) partial least squares (PLS-DA), t) test was used to analyze the extracted ion current intensity of all O-glycan using the receiver operating characteristic curve (ROC), 6) the correlation between O-glycan and clinical tumor markers was analyzed. 7) Immunohistochemistry was used to analyze the expression of MUC1 and Tn-MUC1 in gastric malignant tumor tissues and corresponding paracancerous tissues. Results: 1) the expression of complex glycosaminoglycan synthesized by O-glycan Corel,Core2,ST-antigen and Core2 was up-regulated in serum of gastric malignant tumor (P0.0001); 2) Core3,m/z 529.75, diST antigen was significantly down-regulated in serum of gastric malignant tumor (P0.001); 3) the structure information of the above-mentioned ions was clear by LC-MS/MS analysis, 4) the correlation between O-glycan in serum and secretory CEA and MUC1 in serum was significant by Pearson correlation analysis. 5) the results of immunohistochemistry showed that MCU1 was up-regulated and glycosylation was abnormal in gastric malignant tumors. Conclusion: in this study, the qualitative and quantitative analysis of O-glycan in serum can be achieved by non-reductive dissociation and PMP labeling, together with LC-MS and LC-MS/MS. After statistical analysis, the difference of serum O-glycan between 157 patients with gastric malignant tumor and 144 normal controls was proved, and the correlation between serum O-glycan and glycoprotein (MUC1,CEA) was significant. And the glycosylation modification of these glycoproteins in gastric malignant tumor tissue is also abnormal.
【学位授予单位】：苏州大学
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：R735.2

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