高效表达RKIP基因对人喉鳞癌化疗增敏作用的实验研究

发布时间：2018-04-15 09:20

本文选题：喉癌 + RKIP　；参考：《吉林大学》2010年博士论文

【摘要】： 探讨高效表达RKIP基因联合化疗药物对人喉鳞癌细胞的作用,为喉癌基因联合化疗减低喉癌化疗抵抗提供了理论依据。应用DNA克隆技术构建干涉和高效表达RKIP的重组质粒,并进行测序鉴定和效果鉴定;应用真核细胞转染技术将重组质粒转染人喉癌Hep-2细胞,观察RKIP表达水平变化对喉鳞癌细胞增殖、周期、凋亡及侵袭的影响,并分析其作用机制;高效表达RKIP的重组质粒联合顺铂后,探讨其减低喉癌化疗抵抗和增强喉癌对顺铂敏感性的可能性。(1)经酶切和测序证实,成功构建重组质粒。荧光定量RT-PCR和Western印迹检测RKIP mRNA和蛋白的表达,高效表达组升高,干涉组表达降低,其中干涉组的RKIP-shRNA501抑制作用最为明显。(2)MTT检测结果显示,高效表达组出现增殖抑制,干涉组出现增殖促进。FCM检测结果显示,高效表达组的G1期细胞增多,出现明显的凋亡峰。干涉组的G1期细胞减少,未出现明显的凋亡峰。Matrigel侵袭实验结果显示,高效表达组细胞侵袭力减弱,干涉组细胞侵袭力增强。MAPK通路的改变或许是RKIP引起喉鳞癌Hep-2细胞增殖、周期、凋亡、侵袭等改变的原因或部分原因。(3)高效表达RKIP的重组质粒联合顺铂后,MTT检测结果显示,细胞增殖受到明显抑制。FCM检测结果显示,细胞凋亡明显增强,MAPK通路的改变或许是RKIP-pIRES2-EGFP增强DDP对喉鳞癌Hep-2细胞影响的原因或部分原因。高效表达RKIP基因联合化疗对人喉鳞癌具有显著的治疗效果。
[Abstract]:To investigate the effect of high expression of RKIP gene combined with chemotherapeutic drugs on human laryngeal squamous carcinoma cells, and to provide a theoretical basis for the combination of laryngeal oncogene and chemotherapy to reduce the chemotherapeutic resistance of laryngeal carcinoma.The recombinant plasmids expressing RKIP were constructed by DNA cloning, sequenced and identified, and the recombinant plasmids were transfected into human laryngeal carcinoma Hep-2 cells by eukaryotic cell transfection.To observe the effect of RKIP expression on proliferation, cell cycle, apoptosis and invasion of laryngeal squamous cell carcinoma cells, and to analyze its mechanism.To investigate the possibility of reducing chemotherapeutic resistance and enhancing the sensitivity of laryngeal carcinoma to cisplatin, the recombinant plasmid was successfully constructed by restriction endonuclease digestion and sequencing.Fluorescence quantitative RT-PCR and Western blotting were used to detect the expression of RKIP mRNA and protein. The expression of RKIP mRNA and protein increased in high expression group and decreased in intervention group. The inhibitory effect of RKIP-shRNA501 in intervention group was the most obvious. The results showed that proliferation inhibition appeared in high expression group.The results of proliferation-promoting. FCM assay showed that the G1 phase cells in the high expression group increased and the apoptotic peak appeared.In the intervention group, the G1 phase of cells decreased, and no obvious apoptotic peak. Matrigel invasion assay showed that the cell invasion of the high expression group was weakened, and the increase of the cell invasion and MAPK pathway in the intervention group might be due to the proliferation and cycle of Hep-2 cells induced by RKIP.The reason or part of apoptosis, invasion, etc.) the recombinant plasmid expressing RKIP efficiently combined with cisplatin showed that the cell proliferation was obviously inhibited.The effect of RKIP-pIRES2-EGFP on DDP in laryngeal squamous cell carcinoma (LSCC) might be due to the change of apoptosis.High expression of RKIP gene combined with chemotherapy has a significant therapeutic effect on human laryngeal squamous cell carcinoma.
【学位授予单位】：吉林大学
【学位级别】：博士
【学位授予年份】：2010
【分类号】：R739.65

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