抑癌基因PAX5在非小细胞肺癌中的甲基化及功能研究

发布时间：2018-04-26 14:22

本文选题：PAX5 + 非小细胞肺癌　；参考：《重庆医科大学》2015年硕士论文

【摘要】：目的：检测PAX5在NSCLC中的表达及甲基化状态,研究PAX5甲基化与临床病理特征的相关性,并探讨PAX5在NSCLC中的生物学功能。方法：半定量PCR、real-time PCR检测PAX5在正常成人、癌和癌旁组织及细胞株中的表达。通过MSP检测癌和正常肺组织、良恶性胸水及细胞株中PAX5的甲基化,并用χ2分析PAX5甲基化与临床病理参数的关系。去甲基化药物处理细胞后检测PAX5表达的变化。将PAX5过表达质粒转染NSCLC细胞株A549. H1975 (PAX5表达缺失),通过克隆形成、CCK8、Transwell、划痕、流式细胞等实验,分别检测PAX5对增殖、转移、周期和凋亡的作用。通过裸鼠成瘤实验,验证PAX5的体内抑癌作用。结果：PAX5在正常成人组织中广泛高表达,在86%(12/14)的NSCLC组织中表达下调(P0.0001),在8株细胞中有7株下调或缺失87.5%。在NSCLC组织和正常肺组织中,PAX5甲基化率分别为70%(33/47)和14%(2/16),P0.001。在恶性胸水和良性胸水中,PAX5甲基化为65%(20/31)和11%(1/9),P=0.004。χ2检验显示,PAX5的甲基化状态与组织学类型相关(P=0.006),肺腺癌中,PAX5甲基化率更高,和年龄、分级、分化等不相关(P0.05)。过表达PAX5后,A549、H1975细胞的增殖、迁移和侵袭能力明显降低,细胞阻滞于G2期,凋亡增加。体内裸鼠成瘤实验显示PAX5可明显抑制非小细胞肺癌的生长(P0.001)。结论：非小细胞肺癌中PAX5因启动子区域高甲基化而表达下调或缺失,且对非小细胞肺癌生物学行为有明显的抑制作用,可作为非小细胞肺癌新的候选抑癌基因,为非小细胞肺癌的诊断和指导合理的“个体化治疗”提供有价值的科学依据。
[Abstract]:Aim: to detect the expression and methylation of PAX5 in NSCLC, to study the correlation between PAX5 methylation and clinicopathological features, and to explore the biological function of PAX5 in NSCLC. Methods: semi-quantitative real-time PCR was used to detect the expression of PAX5 in normal adults, cancer and adjacent tissues and cell lines. The methylation of PAX5 in benign and malignant pleural effusion and cell lines was detected by MSP. The relationship between PAX5 methylation and clinicopathological parameters was analyzed by 蠂 2. The changes of PAX5 expression in cells treated with demethylated drugs were detected. PAX5 overexpression plasmid was transfected into NSCLC cell line A549. The effects of PAX5 on proliferation, metastasis, cell cycle and apoptosis were detected by clone formation, scratch, flow cytometry and so on. The tumor inhibition effect of PAX5 in vivo was verified by tumorigenesis in nude mice. Results the expression of 10% PAX5 was highly expressed in normal adult tissues, down-regulated P0.0001 in 86 / 12 / 14 NSCLC, and down-regulated or absent 87.5% in 7 of 8 cell lines. The methylation rates of PAX5 in NSCLC and normal lung tissues were 70 / 33 / 47 and 14 / 2 / 16 / P0.001, respectively. In malignant pleural effusion and benign pleural effusion, PAX5 methylation was 65 / 20 / 31) and 11 / 1 / 9 / 9 P0. 004. 蠂 2 test showed that the methylation status of PAX5 was correlated with histological type (P 0. 006), and the methylation rate of PAX5 in lung adenocarcinoma was higher, not related to age, grade, differentiation and so on (P 0. 05). After overexpression of PAX5, the proliferation, migration and invasion of H1975 cells were significantly decreased, the cells were blocked in G2 phase and apoptosis was increased. In vivo tumorigenesis in nude mice showed that PAX5 could significantly inhibit the growth of NSCLC (P 0.001). Conclusion: the expression of PAX5 in NSCLC is down-regulated or absent due to hypermethylation of promoter region, and it has obvious inhibitory effect on biological behavior of NSCLC. It can be used as a new candidate tumor suppressor gene for NSCLC. To provide valuable scientific basis for the diagnosis and guidance of rational individualized treatment of non-small cell lung cancer.
【学位授予单位】：重庆医科大学
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：R734.2

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