Mining the Gastric Cancer Secretome: Identification of GRN as a Potential Diagnostic Marker for Early Gastric Cancer
Mining the Gastric Cancer Secretome: Identification of GRN as a Potential Diagnostic Marker for Early Gastric Cancer
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DOI：
10.1021/pr201014h
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发表时间：
2012-03-01
影响因子：
4.4
通讯作者：
Chung, Maxey C. M.
中科院分区：
文献类型：
--
作者：
Loei, Hendrick;Tan, Hwee Tong;Chung, Maxey C. M.
胃癌是全球癌症死亡的第二大原因，目前临床上还没有与胃癌诊断和预后相关的生物标志物。在本研究中，我们采用基于2D-LC-MS/MS的方法，结合iTRAQ标记，研究了胃癌细胞系AGS和MKN7的分泌组。通过在条件培养基和全细胞裂解物之间进行比较分析，我们的工作流程使我们能够区分条件培养基中真正分泌的蛋白质和细胞内污染物。与AGS和MKN7细胞的全细胞裂解物相比，90种蛋白质在条件培养基中具有更高的丰度。利用信号肽和非经典分泌预测工具以及在线外泌体数据库，我们证明了这90种蛋白质中高达92.2%可以通过经典或非经典分泌途径输出细胞。然后，我们对AGS和MKN7之间的分泌组进行了定量比较，鉴定出43种差异表达的分泌蛋白。其中，免疫组化发现GRN在胃肿瘤组织中频繁表达，而在正常胃上皮中不表达。夹心ELISA法也显示胃癌患者血清GRN水平升高，尤其是早期胃癌患者。受试者工作特征（ROC）曲线分析证实血清GRN可为胃癌患者提供诊断鉴别
Gastric cancer is the second leading cause of cancer deaths worldwide, and currently, there are no clinically relevant biomarkers for gastric cancer diagnosis or prognosis. In this study, we applied a 2D-LC-MS/MS based approach, in combination with iTRAQ labeling, to study the secretomes of the gastric cancer cell lines AGS and MKN7. By performing a comparative analysis between the conditioned media and the whole cell lysates, our workflow allowed us to differentiate the bona fide secreted proteins from the intracellular contaminants within the conditioned media. Ninety proteins were found to have higher abundance in the conditioned media as compared to the whole cell lysates of AGS and MKN7 cells. Using a signal peptide and nonclassical secretion prediction tool and an online exosome database, we demonstrated that up to 92.2% of these 90 proteins can be exported out of the cells by classical or nonclassical secretory pathways. We then performed quantitative comparisons of the secretomes between AGS and MKN7, identifying 43 differentially expressed secreted proteins. Among them, GRN was found to be frequently expressed in gastric tumor tissues, but not in normal gastric epithelia by immunohistochemistry. Sandwich ELISA assay also showed elevation of serum GRN levels in gastric cancer patients, particularly those with early gastric cancer. Receiver operating characteristic (ROC) curves analysis confirmed that serum GRN can provide diagnostic discriminations for gastric cancer patients