锌指小脑蛋白5促进非小细胞肺癌恶性进展的机制探究

发布时间：2018-06-17 18:56

本文选题：锌指小脑蛋白 + 5　；参考：《南方医科大学》2017年硕士论文

【摘要】：第一部分:锌指小脑蛋白5在非小细胞肺癌中的表达及其与临床病理分期相关性的分析目的肺癌是是世界上发病率最高的恶性肿瘤,也是肿瘤中导致病人死亡的首要原因。锌指小脑蛋白家族成员蛋白(Zic family member)与多种肿瘤的发生发展密切相关。本研究旨在探索锌指小脑蛋白5(ZIC5)在NSCLC中的表达,并分析其与临床病理分期间的关系。方法应用肿瘤基因组图谱(the cancer genome atlas,TCGA)公共数据库筛选出在NSCLC中特异性高表达的ZIC5,并选取南京军区总医院心胸外科50例肺癌和癌旁正常组织,使用实时荧光定量逆转录聚合酶链式反应(qRT-PCR)方法检测ZIC5的信使RNA(mRNA)的表达情况并分析其表达与临床病理表现的关系。结果TCGA数据库分析显示:ZIC5在肺腺癌和肺鳞癌组织中的表达均高于其癌旁正常组织。在50例NSCLC组织中,45例(90%)肿瘤组织中的ZIC5 mRNA表达高于瘤旁正常组织(p0.01),且ZIC5的高表达与更大的肿瘤体积(p=0.032),性别(p=0.048)以及肿瘤类型(p=0.0017)相关。结论ZIC5在非小细胞型肺癌组织中高表达,且ZIC5可能参与了非小细胞型肺癌的恶性进展。第二部分:锌指小脑蛋白5促进非小细胞型肺癌恶性表型的机制研究目的锌指小脑蛋白家5(Zicfamilymember5,ZIC5)可能是一个潜在的促进非小细胞肺癌恶性表型的促癌基因。本研究旨在探讨ZIC5蛋白在非小细胞肺癌中的表达,并分析其与NSCLC恶性表型的关系及其潜在的调控机制,为肺癌的临床治疗提供新的靶点。方法1.使用qRT-PCR方法和western blot法检测NSCLC细胞系中ZIC5的mRNA和蛋白的表达情况。2.设计合适的siRNA干扰ZIC5的表达,体外功能实验观察ZIC5对肿瘤增殖、迁移能力及细胞周期、凋亡的影响。3.体内裸鼠移植瘤模型实验观察ZIC5的干扰对NSCLC肿瘤在裸鼠体内的生长的影响。4.使用qRT-PCR方法和western blot法检测沉默ZIC5后相关细胞周期蛋白的的mRNA和蛋白的表达情况变化。结果1.ZIC5在肺癌细胞系的表达明显高于肺上皮正常细胞。沉默ZIC5后H1703和H1299细胞株的增殖、迁移和侵袭能力明显降低,并使细胞周期阻滞在G2/M期。2.体内成瘤实验显示,沉默ZIC5后肿瘤的生长速度及体积均小于对照组。对移植瘤进行免疫组化染色显示,沉默ZIC5的实验组的KI67和CCNB1表达均低于对照组(p0.05)。3.qRT-PCR 和 westemblot 结果显示,沉默 ZIC5 后,CCNB1 和 phospho-CDKl的表达也受到了明显的抑制,而其它的细胞周期蛋白没有受到影响。结论我们的实验结果显示ZIC5在NSCLC中高表达,可促进NSCLC细胞的增殖和迁移侵袭能力。ZIC5可能通过影响CCNB1和CDK1复合物的表达来发挥其癌基因的功能,因此ZIC5可能作为一个潜在的治疗靶标,为未来NSCLC的诊治提供新的思路。
[Abstract]:The first part: the expression of zinc finger cerebellar protein 5 in non-small cell lung cancer and its correlation with clinicopathological staging; Lung cancer is the most common malignant tumor in the world and the leading cause of death in the tumor. Zic family memberships of zinc finger cerebellar protein family are closely related to the occurrence and development of many kinds of tumors. The purpose of this study was to investigate the expression of zinc finger cerebellar protein 5 (ZIC5) in NSCLC and its relationship with clinical pathological stages. Methods the tumor genome map and the cancer genome atlas TCGA public database were used to screen the specific high expression of ZIC5 in NSCLC. 50 cases of lung cancer and adjacent normal tissues were selected in cardiothoracic surgery of Nanjing military region General Hospital. The expression of messenger RNA-mRNAs of ZIC5 was detected by real-time fluorescence quantitative reverse transcriptase polymerase chain reaction (qRT-PCR) and the relationship between the expression and clinicopathological manifestations was analyzed. Results TCGA database analysis showed that the expression of WZIC5 in lung adenocarcinoma and lung squamous cell carcinoma was higher than that in adjacent normal tissues. The expression of ZIC5 mRNA in the tumor tissues was higher than that in the adjacent normal tissues, and the high expression of ZIC5 was related to the larger tumor volume (p0.032) and the tumor type (p0. 0017). Conclusion ZIC5 is highly expressed in non-small cell lung cancer, and ZIC5 may be involved in the malignant progression of non-small cell lung cancer. Part 2: mechanism of zinc finger cerebellar protein 5 promoting malignant phenotype of non-small cell lung cancer objective Zicfamily member 5 (ZIC5) may be a potential oncogene to promote the malignant phenotype of non-small cell lung cancer (NSCLC). The purpose of this study was to investigate the expression of ZIC5 protein in non-small cell lung cancer (NSCLC), and to analyze the relationship between ZIC5 protein and malignant phenotype of NSCLC and its potential regulatory mechanism, so as to provide a new target for the clinical treatment of lung cancer. Method 1. The expression of ZIC5 mRNA and protein in NSCLC cell line was detected by qRT-PCR and western blot. The expression of siRNA interference ZIC5 was designed, and the effects of ZIC5 on tumor proliferation, migration ability, cell cycle and apoptosis were observed in vitro. To observe the effect of interference of ZIC5 on the growth of NSCLC tumor in nude mice in vivo. QRT-PCR and western blot were used to detect the expression of cyclin mRNA and protein after silencing ZIC5. Results 1. The expression of ZIC5 in lung cancer cell line was significantly higher than that in normal lung epithelial cell line. After silencing ZIC5, the proliferation, migration and invasion of H1703 and H1299 cell lines were significantly decreased, and the cell cycle was blocked at G2 / M phase. In vivo tumorigenesis test showed that the growth rate and volume of tumor after ZIC5 silencing were smaller than those of control group. Immunohistochemical staining showed that the expression of KI67 and CCNB1 in the experimental group was lower than that in the control group (P 0.05). The results of RT-PCR and westemblot showed that the expression of CCNB1 and phospho-CDKl were also significantly inhibited after ZIC5 silencing. Other cyclins were not affected. Conclusion our results suggest that the high expression of ZIC5 in NSCLC can promote the proliferation and migration and invasion of NSCLC cells. ZIC5 may play the role of oncogene by affecting the expression of CCNB1 and CDK1 complexes. Therefore, ZIC5 may be a potential therapeutic target for the future diagnosis and treatment of NSCLC.
【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】：R734.2

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