不同病理类型胶质瘤标本MiRNA表达谱分析及MiR-125a-5p在GBM中生物学功能的研究

发布时间：2018-06-29 23:05

本文选题：胶质瘤 + 病理类型　；参考：《中南大学》2014年博士论文

【摘要】：背景：高级别胶质瘤是成人最常见的原发性颅内恶性肿瘤。尽管近年来针对其手术、放疗及化疗等治疗方案均取得了长足的进展,患者的整体生存率依然较低,预后极差。世界卫生组织(WHO)依据组织病理学将其分为4级,其中胶质母细胞瘤(GBM)是其中最常见、恶性程度最高的一类。GBM患者预后极差,其中位生存期为14.6个月,中位无进展生存期为6.9个月,5年生存率为9.8%。 MicroRNA(MiRNA)是一种不参与蛋白质编码的小分子RNA,通过与靶基因mRNA3'-UTR序列互补的结合,抑制其表达,实现对靶基因的负调控作用。最近的研究表明MiRNA的表达谱异常具有肿瘤特异性,并通过影响关键致癌／抑癌基因的表达参与肿瘤的发生、侵袭、抗凋亡等多种进程。有证据表明单个MiRNA可影响多个靶基因的表达。目前,已有多个针对胶质瘤与不同正常脑组织间MiRNA表达谱的差异的研究,均表现出了表达谱的差异,其在肿瘤进展中的作用机制尚不完全明了。因此,发现并对在胶质瘤中差异表达的MiRNA及其涉及的靶基因进行深入研究有助于揭示其在细胞迁移、侵袭细胞骨架重排及血管形成中的作用,最终实现在诊断、治疗中的应用。目的：针对不同病理类型胶质瘤样本MiRNA结果的筛查,筛选差异表达的MiRNA,用以发现新的胶质瘤相关MiRNA并结合其表达模式推测其在胶质瘤发生发展中的作用。深入研究MiR-125a-5p在GBM中的功能,并通过生物信息学的方式对其靶基因做出预测。方法：分析胶质瘤MiRNA数据库,筛选出与病理类型相关的MiRNA。收集各种病理类型胶质瘤标本,并用qRT-PCR方法验证其表达量。向GBM细胞株转染外源性MiR-125a-5p mimics,通过MTT法、划痕实验与transwell侵袭实验分别检测转染前后细胞增殖、迁移、侵袭能力有无改变；应用生物信息学方法,预测侵袭相关GMBMiRNAs可能的靶基因。结果：(1)在胶质瘤组织中发现42个在不同病理类型胶质瘤中特异性表达的MiRNA,其中特异性下调的有7个,特异性上调的有35个。(2)qRT-PCR实验结果与基因芯片结果相符,MiR-125a-5p在A/O等恶性程度较低的胶质瘤中无明显下调,在AA/AOA/AO/GBM等恶性程度较高的胶质瘤中下调明显。其表达与MGMT的表达有相关性。(3)与对照组相比,转染MiR-125a-5p mimics后96小时内细胞增殖力的变化无统计学意义(P0.05)；(4)与阴性和空白对照组相比,转染MiR-125a-5p mimics的GBM8401细胞侵袭及迁移能力明显增强(P0.01);(5) MiR-125a-5p有多个靶基因,可能参与调节细胞生长、分化、凋亡、转录和信号转导等多种过程相关。结论：1.在胶质瘤组织中发现42个在不同病理类型胶质瘤中特异性表达的MiRNA,其中特异性下调的有7个,特异性上调的有35个。 2. MiR-125a-5p在A/O等恶性程度较低的胶质瘤中无明显下调,在AA/AOA/AO/GBM等恶性程度较高的胶质瘤中下调明显。其表达与MGMT的表达有相关性。 3.MiR-125a-5p的过表达可抑制GBM细胞的迁移及侵袭,对其生长增殖无明显影响。
[Abstract]:Background : High - grade glioma is the most common primary intracranial malignant tumor in adults . Although in recent years , the overall survival rate of patients is low and the prognosis is extremely poor . Although in recent years , the overall survival rate of the patients is low and the prognosis is extremely poor . The World Health Organization ( WHO ) is divided into four grades according to the histopathology . The prognosis of the patients is extremely poor . Among them , the survival time is 14.6 months . Median progression - free survival is 6.9 months , and the 5 - year survival rate is 9.8 % .

MicroRNA ( MiRNA ) is a kind of small molecule RNA which is not involved in protein coding . It can inhibit the expression of target gene by binding to mRNA of mRNA3 ' - 3 ' of target gene . The recent research shows that the expression profile of MiRNA can influence the expression of multiple target genes .

Objective : To screen differentially expressed MiRNA for the screening of MiRNA results of different pathological types of glioma samples and to investigate the role of MiR - 125a - 5p in the development of glioma .

Methods : MiRNA of glioma was analyzed . MiRNA related to pathological type was selected . The expression of MiRNA was verified by qRT - PCR . The exogenous MiR - 125a - 5p - labeled cells were transfected with qRT - PCR . The proliferation , migration and invasion ability of transfected cells were detected by MTT assay , scratch test and transwell invasion assay .
Bioinformatic methods were used to predict the possible target genes of invasive related MRNA .

Results : ( 1 ) The specific expression of MiR - 125a - 5p and MiR - 125a - 5p in glioma tissues was significantly lower than that of control group , and the expression of MiR - 125a - 5p was significantly lower than that of control group ( P0.05 ) .
( 5 ) MiR - 125a - 5p has multiple target genes , which may be involved in regulating cell growth , differentiation , apoptosis , transcription and signal transduction .

Conclusion : 1 . MiRNA specifically expressed in 42 different pathological types of glioma was found in glioma tissues , among which there were 7 specific downregulation , 35 of which were up - regulated .

2 . MiR - 125a - 5p down - regulated in glioma with lower malignant degree , such as A / O and so on , down - regulated in high grade gliomas with higher malignant degree of AA / AOA / AO / . The expression of MiR - 125a - 5p was related to the expression of MGMT .

3 . The overexpression of MiR - 125a - 5p can inhibit the migration and invasion , and has no obvious effect on growth and proliferation .
【学位授予单位】：中南大学
【学位级别】：博士
【学位授予年份】：2014
【分类号】：R739.41

【参考文献】