长链非编码RNANONHSAT070806在膀胱癌中的表达水平及功能研究

发布时间：2018-06-14 00:19

本文选题：膀胱癌 + 长链非编码RNA　；参考：《中国人民解放军医学院》2017年硕士论文

【摘要】：背景膀胱癌是泌尿系统最常见的肿瘤,在其发生发展过程中,肿瘤侵袭起着重要的作用,根据肿瘤侵袭程度的不同,膀胱癌又被分为非肌层浸润型膀胱癌(non-muscle invasive bladder cancer, NMIBC)及肌层浸润型(muscle invasive bladder cancer, MIBC),其中,MIBC恶性程度高,复发率高,预后较差,死亡率明显高于NMIBC。近年来,虽然相关诊疗手段有了较大提高,但临床确诊膀胱癌仍然依靠有创性的膀胱镜检,这对早起筛查膀胱癌造成了不利影响,因此,深入探讨研究膀胱癌病因学相关机制,具有重要的临床意义。长链非编码RNA(Long-noncoding RNA, LncRNA)是一类长度超过200个nt,但没有编码蛋白质的一类RNA,占人体RNA总数的80%以上,LncRNA与肿瘤的相关研究是近年来的热点,包括多种LncRNA被认为在胃癌、前列腺癌、乳腺癌、肺癌、膀胱癌等恶性肿瘤中表达水平发生显著改变,但缺乏相关机制研究。方法通过基因芯片对膀胱癌及癌旁组织差异表达的LncRNA进行分析,选取其中表达差异显著的LncRNA,利用实时定量PCR技术(qPCR)进一步筛选,接着在34例患者中进行验证,在此基础上对LncRNA表达水平与膀胱癌患者临床指标进行关联性分析。构建过表达NONHSAT070806载体,将其瞬时转染入膀胱癌细胞T24和5637中,检测细胞过表达NONHSAT070806后凋亡、增殖、侵袭、迁移和克隆形成能力的变化,探讨NONHSAT070806在体外对膀胱癌细胞生物学特性的影响。更进一步,将转染NONSHAT070806过表达载体和对照空白载体的5637细胞注射入裸鼠皮下,构建裸鼠成瘤模型,通过测量瘤体体积和重量,分析NONHSAT070806对膀胱癌细胞体内成瘤能力的影响。结果发现正义链长链非编码RNANONHSAT070806在大部分患者膀胱癌组织中表达水平下降(24/34, 70.6%),且其表达水平与患者肿瘤侵袭程度呈负相关(p0.05),而与其他临床指标,如年龄、性别、吸烟史无统计学相关性。细胞功能学实验结果表明,NONHSAT070806过表达后,可诱导细胞凋亡,抑制细胞增殖、迁移、侵袭和细胞克隆形成能力(p0.05 ),动物实验结果同样表明,NONHSAT070806过表达可以在体内抑制膀胱癌细胞成瘤能力(p0.05)。结论NONHSAT070806在膀胱癌中低表达,且其表达水平与肿瘤侵袭有关,其可能通过抑制膀胱癌侵袭发挥其抑癌作用,下一步关于其作用靶点及相关机制的研究将有助于揭示膀胱癌的发病机制。
[Abstract]:Background bladder cancer is the most common tumor in the urinary system. Tumor invasion plays an important role in the process of its occurrence and development, according to the degree of tumor invasion. Bladder cancer was also divided into non-muscle invasive bladder cancer, (non-muscle invasive bladder cancer,) and muscle invasive bladder cancer, (invasive bladder cancer,), in which the malignant degree, recurrence rate and prognosis were higher, and the mortality was significantly higher than that of NMIBC. In recent years, although the methods of diagnosis and treatment have been greatly improved, the clinical diagnosis of bladder cancer still depends on invasive cystoscopy, which has a negative impact on early screening of bladder cancer. It has important clinical significance. Long-noncoding RNAs (LncRNAs) are a class of RNAs with length of more than 200 nts but no coding proteins, which account for more than 80% of the total RNA of human body. The research on the relationship between LncRNA and tumors has been a hot topic in recent years, including many kinds of LncRNAs are considered to be involved in gastric cancer and prostate cancer. The expression level of breast cancer, lung cancer, bladder cancer and other malignant tumors have changed significantly, but lack of related mechanism. Methods the differentially expressed LncRNAs in bladder cancer and its adjacent tissues were analyzed by gene microarray. LncRNAs with significant difference in expression were selected and further screened by real-time quantitative PCR, and then verified in 34 patients. On this basis, the correlation between the expression of LncRNA and the clinical indexes of bladder cancer was analyzed. Overexpression of NONHSAT070806 was constructed, and its transient transfection into bladder cancer cells T24 and 5637 was used to detect the changes of apoptosis, proliferation, invasion, migration and clone formation after overexpression of NONHSAT070806. To investigate the effect of NONHSAT070806 on the biological characteristics of bladder cancer cells in vitro. Furthermore, 5637 cells transfected with over expression vector of NONSHAT070806 and control vector were injected subcutaneously into nude mice to construct tumor-forming model of nude mice. The effect of NONHSAT070806 on the tumorigenesis of bladder cancer cells was analyzed by measuring the volume and weight of tumor. The results showed that the expression level of NONHSAT070806 in most patients with bladder cancer decreased by 24 / 34,70.660. The expression level of NONHSAT070806 was negatively correlated with the degree of tumor invasion, but with other clinical parameters, such as age, sex, etc. There was no statistical correlation in smoking history. The results of cell function test showed that the overexpression of NONHSAT070806 could induce cell apoptosis, inhibit cell proliferation and migration. The results of animal experiments also showed that the overexpression of NONHSAT070806 could inhibit the tumorigenesis ability of bladder cancer cells in vivo. Conclusion the expression of NONHSAT070806 is low in bladder cancer, and its expression level is related to tumor invasion, which may play a role in inhibiting the invasion of bladder cancer. Further studies on its target and related mechanisms will be helpful to reveal the pathogenesis of bladder cancer.
【学位授予单位】：中国人民解放军医学院
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R737.14

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