【作者】 李力；

【导师】 邓艳春；

【作者基本信息】 第四军医大学 ， 神经病学， 2010， 博士

【摘要】 中枢神经系统肿瘤,尤其是胶质瘤,以其恶劣的预后严重地危险着人们的健康。寻求疗效更佳的治疗方案和更可靠的预后评估因子,是神经科学家和临床医生们一直的追求。在这个过程中,我校研究人员首先发现了人类的NDRG2基因。该基因在包括胶质瘤在内的多种肿瘤中低表达,其表达受到经典癌基因MYC的负调控;人为调高NDRG2在肿瘤细胞中的表达,会对肿瘤细胞的增殖产生明显的抑制作用;基于上述发现,NDRG2被认为是一种抑癌候选基因,受到我们的持续关注。一方面,通过对NDRG2的解构分析,我们希望发现其作用的具体机制;另一方面,我们也想知道该基因的表达状态对于肿瘤患者的实际意义如何。本项研究的第一部分是从人类NDRG2蛋白的分子分析入手,预测其三维结构为两个结构域构成的球性蛋白,根据结构域,设计出若干分别代表其N末端、C末端,以及两个结构域的截短肽片段,构建其真核表达载体,转染胶质瘤细胞系U87MG和U251以及经典肿瘤细胞HeLa,通过流式细胞术观察肿瘤细胞的凋亡、周期阻滞和增殖(MTT法,仅对HeLa)情况,发现C末端的129肽对NDRG2发挥抗肿瘤作用最为关键,且N、C末端折叠成的结构域和C末端自身都可在某些情况下发挥较全长蛋白更强的抗肿瘤作用,且以诱导细胞周期阻滞为主。在本项研究的第二部分,我们以一个时间段于我院进行常规手术治疗和放、化疗的胶质瘤患者为研究对象,通过免疫组化分析其肿瘤组织中的NDRG2的表达情况,比较其表达与肿瘤分级的关系;并通过对NDRG2表达和术后患者生存状态、生活质量的相关分析,对NDRG2作为预后因子的可能性进行评价,结果表明:NDRG2蛋白表达与病理分级呈负相关;与星形细胞瘤患者的术后累计生存率、生存时间、生活质量呈正相关,可以作为独立危险因素来评估胶质瘤患者的预后。总之,通过这项研究,我们对NDRG2发挥作用的关键部位进行了定位,也评价了NDRG2在胶质瘤预后判断中的价值,既为今后机制研究明确了方向,也为其作为预后指标及治疗手段而应用于临床奠定了基础。更多还原

【Abstract】 The awful prognosis of glioma is serious threaten for people’s heath. It is of a persistent work for all neuroscientists and neurologists to pursue the reliable prognositic biomarker and the new therapies with well-prognosis.Our team members found the human NDRG2 gene first and confirmed its low expression in many tumor tissues and cell lines. It is downstream regulated by MYC, and inhibits the cell proliferation when it is transfected into glioma cell lines and induced to overexpress. Thus the gene is regarded as a candidate for tumor suppressor gene and attracts our attention for a long time.Two questions focus our attention on the present study: Which part of the NDRG2 plays the leading role in its antitumor function? What correlation exists between the expression of NDRG2 and the patients’overall survival?The first part of the present study is about the correlation between the structure of NDRG2 and its antitumor function. Here, we started our research from the analysis for the molecular structure of NDRG2, predicted its 3-dimentional structure according to its homologous protein of mouse. Then, we truncated the NDRG2 protein into several parts theoretically, designed and constructed the eukaryotic expression plasimds for each truncated peptides of NDRG2. Two glioma cell lines (U87MG and U251) and a classical cervical cancer cell line (HeLa) were transfected by the plasmids and their cell cycle arrest and apoptosis were dectected by flowcytometry. More than that, the proliferation of HeLa cells after being transfected by our plasminds was also observed by MTT assay. We found that the C-terminal of NDRG2 plays the leading role in its antitumor fuction, and sometimes the truncated Peptide AC and Peptide C showed more powerful antitumor function than the whole length protein. Peptide AC is the mimic of the big domain of original NDRG2 protein and Peptide C is consisted of 129 amino acids adjacent to C-terminal.In the second part, we studied the expression profile of NDRG2 in human astrocytoma tissues by immunohistochemistry and found it was significantly downregulated in astrocytomas. The level of expression of NDRG2 showed an inverse correlation with the WHO grade of tumors and a positive one with the life span of patients. Our results suggested that NDRG2 may play a pivotal role as a potent tumor suppressor and a prognostic biomarker candidate for human astrocytoma.In brief, we located the part playing the leading role for NDRG2 antitumor function and assessed the value of NDRG2 as prognostic biomarker in the study.更多还原