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目的:探讨新的方法筛选发现前列腺癌的分子干预靶点。方法:收集2007年2月至2008年4月间我院收治手术切除前列腺癌组织和良性前列腺增生组织及其配对的患者血清标本3例。提取组织总蛋白,双向电泳分离并转膜,正常人血清封闭,患者血清作为一抗杂交,行Western印迹检测,前后比较差异表达蛋白,MALDI-TOF-MS质谱分析,数据库查询鉴定后进行RT-PCR法和Western印迹验证。结果:该方法改进可以显著提高展示两组差异表达蛋白。鉴定发现差异表达的蛋白点包括自吞噬蛋白1(Beclin1),谷胱苷肽S转移酶P(GSTP1-1),二氢二醇脱氢酶2(DDH2),葡萄糖依赖性胰岛素释放肽受体(GIPR),磷脂酰乙醇胺结合蛋白(PEBP),烯醇酶(ENO1),锰-超氧化物歧化酶(MnSOD),磷酸甘油酸变位酶1(PGAM1)、肽基脯氨基顺反异构酶(PPIA)等,其中Beclin1mRNA和蛋白在前列腺癌组织中的表达显著性下调。结论:采用改进的血清免疫印迹技术为筛选前列腺癌发生发展的分子提供了一种新的方法和思路;Beclin1调控的自吞噬机制可能发挥了关键性的作用。
Objective: To explore a new method to screen molecular targets of prostate cancer. Methods: Three cases of sera from patients with prostate cancer and benign prostatic hyperplasia (BPH) and their paired patients undergoing surgical resection in our hospital from February 2007 to April 2008 were collected. Tissue proteins were extracted and separated by two-dimensional electrophoresis and then transferred into the membrane. The normal human serum was blocked. The serum of the patients was used as the primary antibody to hybridize. Western blotting was used to detect the protein in the tissues. MALDI-TOF-MS was used to analyze the protein. PCR and Western blotting. Results: This method can significantly improve the display of two groups of differentially expressed proteins. Identified differentially expressed protein spots include Beclin1, GSTP1-1, DDH2, glucose-dependent insulin-releasing peptide receptor (GIPR), phosphatidylethanolamine binding protein (PEBP), enolase (ENO1), manganese-superoxide dismutase (MnSOD), phosphoglycerate mutase 1 (PGAM1) Enzyme (PPIA) and so on. The expression of Beclin1 mRNA and protein in prostate cancer tissues was significantly down-regulated. CONCLUSION: The improved serum immunoblotting technique provides a new method and idea for the screening of molecules that are responsible for the development of prostate cancer. Beclin1-mediated autophagy may play a key role.