长链非编码RNA linc-UBC1在结直肠癌中的表达及其功能研究

发布时间：2018-04-18 17:27

  本文选题：长链非编码RNA + linc-UBC1　； 参考：《南方医科大学》2017年硕士论文

【摘要】：目的现已有大量的研究证明了长链非编码RNA(lncRNA)对恶性肿瘤发生发展的调控作用。据研究报道长链非编码RNA linc-UBC1可通过结合PRC2而影响膀胱癌预后。本研究拟通过检测linc-UBC1在结直肠癌(CRC)中的表达,探索linc-UBC1对CRC细胞生物学功能的影响,初步揭示linc-UBC1对CRC的作用,为今后CRC的诊断和治疗提供新的思路。方法1.收集96例结直肠癌患者的手术标本(包含癌组织和对应癌旁组织)和患者的临床病理资料,结直肠癌细胞系(SW480、SW620、HCT116和HT29)以及正常结肠上皮细胞NCM460由南方医科大学捐赠。2.通过实时荧光定量聚合酶链反应技术(RT-qPCR)检测结直肠癌组织和对应癌旁组织linc-UBC1的表达量,并分析其与临床病理资料的关系,采用Kaplan—Meier法分析linc-UBC1与生存资料的关系。3.通过RT-qPCR检测4种结直肠癌细胞系及正常结肠上皮细胞NCM460中linc-UBC1的表达量,选取linc-UBC1表达量最高的结直肠癌细胞系SW620,通过脂质体转染技术干扰SW620细胞系中的linc-UBC1基因,并使用RT-qPCR检测其干扰效率。4.运用CCK8、流式细胞术及Transwell实验检测linc-UBC1基因干扰后的SW620细胞系的增殖能力、凋亡率及周期、迁移及侵袭能力的变化。5.运用western blot实验检测linc-UBC1基因干扰后的SW620细胞系的凋亡相关蛋白(cleaved caspase-3、cleaved caspase-9 和 Bcl-2)表达量的变化。结果1.linc-UBC1在结直肠癌组织中的相对表达量为6.55±6.60,远高于癌旁组织的相对表达量1.67±1.54,两者比较差异有统计学意义(t=8.32,P0.001)。2.通过分析linc-UBC1的表达量与临床病理的关系得出:linc-UBC1的表达量与结直肠癌的TNM分期、肿瘤的大小、浸润的深度及淋巴结转移相关;差异有统计学意义(P0.05)。而与患者的性别、年龄、肿瘤的分化程度、肿瘤位置和远处转移无关(P0.05)。生存分析结果显示:linc-UBC1高表达组的生存时间明显小于linc-UBC1低表达组,差异有统计学意义(P0.01)。多因素COX回归分析表明:linc-UBC1的表达水平可作为CRC患者的独立预后因子(Hazard ratio:2.434,95%CI:1.093-5.420,P=0.029)。3.与 NCM460 相比,SW620、HCT116 和 HT29 中 linc-UBC1 表达水平均明显升高(P0.01),而在SW480细胞中无显著差异(P0.05),其中SW620的linc-UBC1表达量最高。4.与阴性对照组相比,linc-UBC1干扰组的SW620的增殖、迁移及侵袭能力降低,凋亡率升高及周期受到抑制(P0.05)。5.与阴性对照组相比,linc-UBC1干扰组的SW620的凋亡相关蛋白cleaved caspase-3和cleavedcaspase-9的表达量显著上升,Bcl-2的表达量显著下降,差异有统计学意义(P0.05)。结论linc-UBC1在CRC组织中的表达量显著升高,且linc-UBC1的表达量与CRC的TNM分期、肿瘤的大小、浸润的深度、淋巴结转移及不良预后正相关,并且linc-UBC1可以促进CRC细胞的增殖、迁移及侵袭。因此,linc-UBC1有可能成为判断CRC患者预后和发展的新的肿瘤标记物。
[Abstract]:Objective A large number of studies have demonstrated the regulatory effect of long chain noncoding RNAs (LNRNAs) on the carcinogenesis and development of malignant tumors.It has been reported that long chain non-coding RNA linc-UBC1 can affect the prognosis of bladder cancer by binding PRC2.This study was designed to explore the effect of linc-UBC1 on the biological function of CRC cells by detecting the expression of linc-UBC1 in colorectal carcinoma, and to reveal the effect of linc-UBC1 on CRC, and to provide a new idea for the diagnosis and treatment of CRC in the future.Method 1.The clinical and pathological data of 96 patients with colorectal cancer (including cancer tissues and adjacent tissues) and the clinicopathological data were collected. The colorectal cancer cell lines SW480, SW620, HCT116 and HT29, as well as normal colonic epithelial cells NCM460, were donated by Southern Medical University.Real-time fluorescence quantitative polymerase chain reaction (RT-qPCR) was used to detect the expression of linc-UBC1 in colorectal cancer tissues and adjacent tissues, and the relationship between linc-UBC1 and survival data was analyzed by Kaplan-Meier method.The expression of linc-UBC1 in NCM460 of four colorectal cancer cell lines and normal colonic epithelial cells was detected by RT-qPCR. The cell line SW620 with the highest linc-UBC1 expression was selected, and the linc-UBC1 gene in SW620 cell line was interfered with by liposome transfection technique.And use RT-qPCR to detect its interference efficiency. 4. 4.CCK8, flow cytometry and Transwell assay were used to detect the changes of proliferation, apoptosis rate and cycle, migration and invasion ability of SW620 cell lines after linc-UBC1 gene interference.Western blot assay was used to detect the expression of apoptosis-related proteins (caspase-9 and Bcl-2) in SW620 cell lines after linc-UBC1 gene interference.Results the relative expression of 1.linc-UBC1 in colorectal cancer was 6.55 卤6.60, which was significantly higher than that in paracancerous tissues (1.67 卤1.54).By analyzing the relationship between the expression of linc-UBC1 and clinicopathology, it was concluded that the expression of linc-UBC1 was correlated with the TNM stage, tumor size, depth of invasion and lymph node metastasis of colorectal cancer, and the difference was statistically significant (P 0.05).There was no correlation between sex, age, tumor differentiation, tumor location and distant metastasis (P 0.05).Survival analysis showed that the survival time of the high expression group of linc-UBC1 was significantly shorter than that of the low expression group of linc-UBC1, and the difference was statistically significant (P 0.01).Multivariate COX regression analysis showed that the expression level of BC1 in CRC patients could be regarded as the independent prognostic factor of CRC patients.Compared with NCM460, the expression of linc-UBC1 in HCT116 and HT29 was significantly higher than that in NCM460, but there was no significant difference in SW480 cells. The linc-UBC1 expression of SW620 was the highest. 4.Compared with the negative control group, the SW620 proliferation, migration and invasion ability of the control group were decreased, the apoptosis rate was increased and the cycle was inhibited by P0.05. 5.Compared with the negative control group, the expression of apoptosis-related protein cleaved caspase-3 and cleavedcaspase-9 in the SW620 interference group was significantly increased, and the expression of Bcl-2 was significantly decreased in the control group, and the difference was statistically significant (P 0.05).Conclusion the expression of linc-UBC1 in CRC tissues was significantly increased, and the expression of linc-UBC1 was positively correlated with the TNM stage of CRC, tumor size, depth of invasion, lymph node metastasis and poor prognosis. Linc-UBC1 could promote the proliferation, migration and invasion of CRC cells.Therefore, linc-UBC1 may be a new tumor marker for the prognosis and development of CRC patients.
【学位授予单位】：南方医科大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R735.3
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本文编号：1769346