OTUB1在结肠癌中的表达及与其DCN关系的初步研究

发布时间：2018-03-13 20:31

本文选题：OTUB1　切入点：DCN　出处：《青岛大学》2016年硕士论文　论文类型：学位论文

【摘要】：背景和目的:泛素-蛋白酶体途径是调控细胞内蛋白质降解的重要途径,涉及细胞周期调控、信号通路的转导、细胞DNA修复、炎症反应及肿瘤发展等多个方面。去泛素化酶可逆向调控该途径。饰胶蛋白聚糖(decorin,DCN)是细胞外基质一种成分,能够抑制肿瘤细胞生长、逆转肿瘤细胞表型、抑制肿瘤血管生成。DCN主要通过泛素-蛋白酶体途径进行降解。卵巢肿瘤相关蛋白酶B1(OTU domain-containing ubiquitin aldehyde-binding protein B1,OTUB1)属于去泛素化酶家族的成员,既可以抑制蛋白的泛素化,也可以促进泛素化过程。本研究旨在探讨OTUB1在结肠组织中的表达,并进一步探讨OTUB1与DCN之间的表达趋向。方法:收集结肠癌、腺瘤、正常粘膜组织,采用免疫组织化学法检测OTUB1在结肠组织的表达。应用RT-PCR检测结肠癌组织中DCN的m RNA表达。选定稳定转染OTUB1的结肠癌细胞系lovo和转染空载质粒结肠癌细胞系lovo(对照组)作为研究对象,采用实时荧光定量PCR(Real time quantitative PCR,real time qPCR)和Western blotting,在m RNA和蛋白水平上检测OTUB1与DCN的表达。结果:(1)免疫组化:OTUB1在结肠癌组织中的表达明显高于正常粘膜组织,差异具有统计学意义(P0.05)。(2)RT-PCR结果显示结肠癌组织中扩增出的295bp目的DCN条带明显增亮,差异具有统计学意义(P0.05)。(3)实时荧光定量PCR:转染OTUB1的结肠癌细胞OTUB1与DCN的m RNA表达水平均上调,差异具有统计学意义(P0.05)。(4)Western-blot:与对照组lovo细胞相比较,稳定转染OTUB1的lovo细胞中OTUB1与DCN蛋白均有明显表达,差异均具有统计学意义(P0.05)。结论:OTUB1蛋白在正常结肠粘膜、腺瘤、结肠癌组织中的表达呈递增趋势,OTUB1的过表达抑制了DCN的泛素化降解。
[Abstract]:Background & AIM: the ubiquitin proteasome pathway is an important pathway to regulate the degradation of proteins in cells, which involves cell cycle regulation, signal pathway transduction, and cell DNA repair. Inflammatory response and tumor development are many aspects. Desuginase can reverse regulate this pathway. Decorin DCNC is a component of extracellular matrix, which can inhibit the growth of tumor cells and reverse the phenotype of tumor cells. Inhibition of tumor angiogenesis. DCN is mainly degraded by ubiquitin proteasome pathway. Ovarian tumor-associated protease B1OTU domain-containing ubiquitin aldehyde-binding protein B1OTUB1) is a member of the family of diubiquitin enzymes, which can inhibit the ubiquitin of proteins. The aim of this study was to investigate the expression of OTUB1 in colon tissue and the expression trend between OTUB1 and DCN. Methods: to collect colon cancer, adenoma, normal mucous tissue. Immunohistochemical method was used to detect the expression of OTUB1 in colon tissue. RT-PCR was used to detect m RNA expression of DCN in colon cancer tissue. The colon cancer cell line lovo, which was stably transfected with OTUB1, and the colon cancer cell line, lovo-transfected with empty plasmid, were selected (control group). Group) as the subject of the study, The expression of OTUB1 and DCN at m RNA and protein levels was detected by real-time fluorescence quantitative PCR(Real time quantitative PCRRreal time qPCR) and Western blotting. The results showed that the expression of OTUB1 and DCN in colon cancer tissues was significantly higher than that in normal mucosa tissues. The results of RT-PCR showed that the 295bp DCN band amplified from colon cancer tissue was significantly enhanced, and the difference was statistically significant (P 0.05). The expression of m RNA in OTUB1 transfected colon cancer cell line OTUB1 and DCN were all upregulated. Compared with the control lovo cells, the expression of OTUB1 and DCN protein in lovo cells transfected with OTUB1 was significantly higher than that in the control group (P 0.05, P 0.05). Conclusion the expression of OTUB1 and DCN protein in normal colon mucosa and adenoma was significantly higher than that in control group. The overexpression of OTUB1 in colon cancer tissues inhibited the ubiquitin degradation of DCN.
【学位授予单位】：青岛大学
【学位级别】：硕士
【学位授予年份】：2016
【分类号】：R735.35

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