CBX8在肝细胞肝癌中的表达及对SMMC-7721细胞增殖、侵袭能力和细胞周期的影响

发布时间：2018-04-15 14:05

本文选题：CBX8 + 肝细胞肝癌　；参考：《青岛大学》2017年硕士论文

【摘要】：目的通过检测CBX8 m RNA和蛋白质在肝细胞肝癌组织中的表达及对肝癌细胞系SMMC-7721增殖、侵袭能力和细胞周期的影响,初步探讨CBX8在肝细胞肝癌发生发展中的作用及其意义。方法懫用实时荧光定量PCR和western blot分别检测15对新鲜肝细胞肝癌手术标本及癌旁组织中CBX8 m RNA和蛋白质的表达;懫用组织芯片技术及免疫组织化学方法检测CBX8蛋白质在120例肝细胞肝癌手术标本及癌旁组织中的表达情况;分析CBX8的表达与肝癌发生及临床病理特征之间的关系。分别在肝癌细胞系SMMC-7721中瞬时转染小干扰RNA和过表达质粒,通过MTT、平板克隆形成实验及transwell小室检测CBX8的敲降和过表达对肝癌细胞系SMMC-7721细胞增殖及侵袭能力的影响,通过细胞同步化及流式细胞仪检测CBX8对肝癌细胞系SMMC-7721细胞周期的影响。结果实时荧光定量PCR结果显示CBX8 m RNA在13例(13/15)新鲜肝细胞肝癌手术标本中表达量增高,15例肝细胞肝癌与癌旁组织中的差异具有统计学意义(t=2.742,P0.05)。Western blot结果表明CBX8蛋白质在11例(11/15)新鲜肝细胞肝癌组织中表达量增高,15例肝细胞肝癌与癌旁组织中的差异具有统计学意义(t=2.686,P0.05)。组织芯片及免疫组化检测结果显示CBX8在100例(83.33%)肝细胞肝癌手术标本中表达,在46例(38.33%)癌旁组织中表达,CBX8在120例肝细胞肝癌及癌旁组织的表达差异具有统计学意义(Z=-8.072,P0.05)。CBX8在肝细胞肝癌中的表达水平与患者乙肝表面抗原(HBs Ag)、肿物直径、被膜侵犯以及TNM分期有关(?2=10.900、4.043、46.570、8.374,P0.05)。敲降CBX8后,肝癌细胞系SMMC-7721细胞的增殖、克隆形成率和迁移侵袭能力均下降,且与对照组相比差异均具有统计学意义(F=9.395、24.300、71.180、8.745、63.700、9.044,P0.05);过表达CBX8则结果相反,并且与对照组相比差异均具有统计学意义(F=7.177、36.100、24.680、9.045、7.037、6.525,P0.05)。通过检测细胞周期,我们发现敲降CBX8后,实验组G1期细胞所占比例明显增加,S期细胞所占比例减少,差异有统计学意义(t=3.170、3.269,P0.05);过表达CBX8后,实验组G1期细胞所占比例明显减少,S期细胞所占比例增加,差异有统计学意义(t=2.884、4.443,P0.05)。结论CBX8在肝细胞肝癌中呈显著高表达;CBX8在肝细胞肝癌的发生发展中发挥重要作用,能够促进肝癌细胞系SMMC-7721的增殖、迁移、侵袭等生物学行为,并可能通过调控细胞周期G1/S转换进而影响SMMC-7721细胞周期进程;CBX8在肝细胞肝癌发生发展中发挥癌基因作用。
[Abstract]:Objective to investigate the effect of CBX8 m RNA and protein on the proliferation, invasion and cell cycle of hepatocellular carcinoma (HCC) cell line SMMC-7721 by detecting the expression of CBX8 m RNA and protein in HCC tissues.Methods Real-time quantitative PCR and western blot were used to detect the expression of CBX8 m RNA and protein in 15 fresh hepatocellular carcinoma specimens and adjacent tissues.Tissue microarray technique and immunohistochemical method were used to detect the expression of CBX8 protein in 120 HCC specimens and adjacent tissues, and the relationship between the expression of CBX8 and the occurrence and clinicopathological features of HCC was analyzed.Transient transfection of small interfering RNA and overexpression plasmids were carried out in hepatocellular carcinoma (SMMC-7721) cell lines. The effects of CBX8 knockdown and overexpression on the proliferation and invasiveness of SMMC-7721 cells were detected by MTT, plate clone formation assay and transwell chamber.Cell synchronization and flow cytometry were used to detect the effect of CBX8 on the cell cycle of hepatocellular carcinoma cell line SMMC-7721.Results the results of real-time fluorescence quantitative PCR showed that the expression of CBX8 m RNA was increased in 13 cases of fresh hepatocellular carcinoma and 15 cases of hepatocellular carcinoma. The difference between 15 cases of hepatocellular carcinoma and adjacent tissues was statistically significant. Western blot showed that CBX8 protein was expressed in 13 cases of fresh hepatocellular carcinoma.In 11 cases of fresh hepatocellular carcinoma (11 / 15), there was a significant difference between 15 cases of hepatocellular carcinoma (HCC) and adjacent tissues (P 0.05).The results of tissue microarray and immunohistochemistry showed that CBX8 was expressed in 100 specimens of hepatocellular carcinoma (HCC).There were significant differences in the expression of CBX8 in 120 cases of hepatocellular carcinoma (HCC) and its adjacent tissues in 46 cases (38.33). The expression level of CBX8 in hepatocellular carcinoma (HCC) was significantly higher than that in patients with Hepatocellular carcinoma (HCC).The membrane invasion and TNM staging were related to 46.570 ~ 8.374g / L P0.05A, P = 10.900 ~ 4.043n ~ 46.570g / L, P = 0.05a, P = 0.05a, respectively.After knockdown of CBX8, the proliferation, clone formation rate and migration and invasion ability of SMMC-7721 cell line decreased, and the difference was statistically significant compared with the control group. The difference was statistically significant compared with the control group.Compared with the control group, the difference was statistically significant (P 0.05).By measuring cell cycle, we found that the percentage of G 1 phase cells in the experimental group decreased significantly after knocking down CBX8, and the difference was statistically significant (P 0.05) after overexpression of CBX8.The percentage of G 1 phase cells in the experimental group was significantly decreased and the percentage of S phase cells was increased. The difference was statistically significant (P 0.05).Conclusion CBX8 plays an important role in the carcinogenesis and development of hepatocellular carcinoma (HCC), and can promote the proliferation, migration, invasion and other biological behaviors of HCC cell line SMMC-7721.CBX8 may play an oncogene role in the carcinogenesis and development of hepatocellular carcinoma (HCC) by regulating the G1 / S transition of cell cycle and then affecting the progression of SMMC-7721 cell cycle.
【学位授予单位】：青岛大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R735.7

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