MiR-124对胶质瘤生长和新的血管生成的影响及分子机制

发布时间：2019-01-10 09:16

【摘要】：胶质瘤代表了人类脑部肿瘤中最具侵袭性,最致命的一类恶性肿瘤,起源于脑组织和脊髓,发生于神经胶质细胞,约占中枢神经系统肿瘤的40%。近年来,胶质瘤的临床治疗主要依靠手术的切除,并结合放疗和／或化疗的联合疗法,但由于其呈浸润性生长,具有高度侵袭性和富集血管性的特点,决定了胶质瘤手术不能完全切除,其术后复发率高,五年存活率仍旧不理想。因此,进一步探究神经胶质瘤的发病机制,寻找新的诊断和治疗方法显得尤为重要。微小RNAs (MicroRNAs, miRNAs)是一类长度约18-24个核苷的小的、非编码RNA分子,可以在植物和动物中被发现,在进化上高度保守,通过与靶基因nRNA3'-UTR区域配对调控基因的表达,调节细胞分化、凋亡、细胞周期、增殖和分裂等生理病理过程。MiRNAs在多种肿瘤中出现特征性的表达种类和量的变化,在肿瘤发生发展中扮演着癌基因或者抑癌基因的角色,并且以miRNAs为基础治疗的方法已经得到越来越多的关注。我们收集了6例正常脑组织和2/3/4级胶质瘤组织各8例,共30例临床组织标本,检测发现在脑胶质瘤组织中,miR-124的表达显著低于正常脑组织,与其他报道结果一致,并且miR-124表达水平的高低与胶质瘤临床分级密切相关,提示miR-124在胶质瘤中起着重要的作用。同时,我们还构建了过表达miR-124的胶质瘤稳定细胞株,分别在分子水平和动物水平上研究miR-124的生物学功能。我们发现,在胶质瘤细胞系中,过表达miR-124可以显著抑制细胞增长速率和细胞侵袭能力,并且可以在裸鼠体内成瘤实验中抑制肿瘤的生长和肿瘤新的血管生成。为了进一步研究其潜在的分子机制,我们通过生物信息学软件预测和报告基因检测发现miR-124可直接靶向R-RAS和N-RAS。R-RAS和N-RAS是参与细胞信号转导的致癌基因,它们的激活可影响下游PI3K/AKT和RAF/ERK1/2两条关于肿瘤生长和新的血管生成的重要信号通路。我们发现miR-124可通过降低R-RAS和N-RAS的蛋白水平,并通过下游信号通路下调HIF-1a及VEGF的表达,抑制肿瘤生长及肿瘤新的血管生成。我们还发现,与正常脑组织相比,R-RAS和N-RAS在胶质瘤临床标本中表达上调,并且和miR-124的表达水平呈负相关。综上所述,miR-124可通过直接靶向调节R-RAS和N-RAS,影响其下游信号的表达,从而抑制胶质瘤的生长及新的血管生成。MiR-124在体内和体外都能抑制胶质瘤的发生发展,并且miR-124及其靶基因R-RAS/N-RAS与胶质瘤的发展进程密切相关。通过研究,我们发现了miR-124的两个新的作用靶点,阐明了miR-124通过其靶点抑制胶质瘤生长的作用及其分子机制,为胶质瘤诊断及预后的miRNAs应用提供了新的理论依据,并为今后开发以niRNAs为基础的治疗药物提供了新的理论支持。
[Abstract]:Gliomas represent the most aggressive and fatal type of malignant tumors in the human brain, originating in the brain and spinal cord and occurring in glial cells, accounting for about 40 percent of the tumors in the central nervous system. In recent years, the clinical treatment of glioma mainly depends on surgical resection, combined with radiotherapy and / or chemotherapy. However, because of its invasive growth, glioma has the characteristics of highly invasive and vascular enrichment. It is determined that glioma surgery can not be completely resected, its recurrence rate is high, five-year survival rate is still unsatisfactory. Therefore, it is very important to explore the pathogenesis of glioma and find new methods of diagnosis and treatment. Tiny RNAs (MicroRNAs, miRNAs) are a class of small, non-encoded RNA molecules with a length of about 18-24 nucleosides that can be found in plants and animals and are highly conserved in evolution, regulating the expression of genes by pairing with the nRNA3'-UTR region of the target gene. Regulation of cell differentiation, apoptosis, cell cycle, proliferation and mitosis, and other physiological and pathological processes. MiRNAs in a variety of tumors have a characteristic variety of expression and quantity changes, and play the role of oncogene or tumor suppressor gene in tumorigenesis and development. And more attention has been paid to the treatment based on miRNAs. We collected 6 cases of normal brain tissue and 8 cases of grade glioma in 2-3-4, 30 clinical tissue specimens were collected. We found that the expression of miR-124 in glioma tissue was significantly lower than that in normal brain tissue, which was consistent with other reported results. The level of miR-124 expression is closely related to the clinical grade of glioma, suggesting that miR-124 plays an important role in glioma. At the same time, we also constructed a stable glioma cell line with overexpression of miR-124 to study the biological function of miR-124 at the molecular level and animal level, respectively. We found that overexpression of miR-124 could significantly inhibit cell growth and cell invasion in glioma cell lines, and inhibit tumor growth and tumor angiogenesis in nude mice. In order to further study its potential molecular mechanism, we found that miR-124 can directly target R-RAS, N-RAS.R-RAS and N-RAS as the oncogenes involved in cell signal transduction through bioinformatics software prediction and report gene detection. Their activation may affect downstream PI3K/AKT and RAF/ERK1/2 signaling pathways related to tumor growth and new angiogenesis. We found that miR-124 can inhibit tumor growth and tumor angiogenesis by decreasing the protein levels of R-RAS and N-RAS and down-regulating the expression of HIF-1a and VEGF through downstream signaling pathway. We also found that the expression of R-RAS and N-RAS was up-regulated in clinical specimens of glioma compared with normal brain tissue and negatively correlated with the expression of miR-124. In conclusion, miR-124 can inhibit the growth and angiogenesis of gliomas by directly targeting the expression of R-RAS and N-RAS. MiR-124 can inhibit the occurrence and development of gliomas in vivo and in vitro. MiR-124 and its target gene R-RAS/N-RAS are closely related to the development of glioma. We have found two new targets of miR-124, elucidated the role of miR-124 in inhibiting glioma growth and its molecular mechanism, and provided a new theoretical basis for miRNAs application in glioma diagnosis and prognosis. It provides new theoretical support for the development of niRNAs-based drugs in the future.
【学位授予单位】：南京医科大学
【学位级别】：硕士
【学位授予年份】：2014
【分类号】：R739.41

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