| siRNA沉默人胃癌细胞COX-2基因对Notch信号通路影响的研究 |
| Alternative Title | The effect of silencing COX-2 genes on Notch signaling pathway in human gastric carcinoma cells
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| 王璟 |
| Thesis Advisor | 刘宏斌
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| 2014-05-23
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| Degree Grantor | 兰州大学
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| Place of Conferral | 兰州
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| Degree Name | 硕士
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| Keyword | 胃肿瘤
环氧合酶-2
RNA干扰
Notch信号通路
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| Abstract | 目的：1.研究siRNA沉默COX-2基因表达对人胃癌细胞增殖和凋亡能力的影响。2.研究siRNA沉默COX-2基因表达对人胃癌细胞Notch信号通路的影响及相关机制。
方法：siRNA干扰技术沉默胃癌MKN45细胞 COX-2基因，将实验分为三组，转染siRNA-COX-2的胃癌细胞为实验组，转染siRNA Control为阴性对照组，以未转染的MKN45细胞为空白组。应用荧光定量-PCR检测靶向沉默COX-2基因的表达和沉默效率，以及Notch信号通路相关基因的表达水平；蛋白免疫印记法检测COX-2，N1ICD，N2ICD和Hes1蛋白表达水平；细胞计数法检测各组细胞增殖，流式细胞术检测分析各组细胞凋亡。
结果：转染siRNA沉默COX-2基因可明显下调COX-2的表达量，实验组与阴性对照组比较差异有统计学意义（P＜0.05），表达抑制率为（81.81±3.82）%；Notch信号通路下游基因Hes1 mRNA表达水平也下调，差异有统计学意义（P＜0.05），表达抑制率为（74.01±4.35）%；Notch1和Notch2 mRNA表达水平无明显变化，与阴性对照组比较，差异无统计学意义（P＞0.05）；沉默COX-2基因后COX-2，N1ICD，N2ICD和Hes-1蛋白表达明显受到抑制，实验组与阴性对照组比较，差异有统计学意义（P＜0.05）。细胞计数法检测在转染后第4天细胞增值能力明显下降（P<0.05）；沉默COX-2基因可提高细胞凋亡率，转染48小时后实验组与阴性对照组比较，差异有统计学意义（P＜0.05）。
结论：靶向沉默COX-2基因抑制人胃癌细胞增殖和促进细胞凋亡可能是通过抑制Notch信号通路实现。 |
| Other Abstract | Objective: 1. To investigate the effect of silencing COX-2 gene on proliferation and apoptosis. 2. To investigate the underlying mechanisms with Notch signaling pathway in human gastric carcinoma cell line by target RNA interference.
Methods: The siRNA of COX-2 was transfected into gastric cancer cell line MKN45 by Lipofectamine 2000. MKN45 cells transfected with blank vector were served as negative control group. MKN45 cells non-transfected served as blank group. Application of qRT-PCR to detect the COX-2 gene expression and silencing efficiency. The mRNA expression of Notch1, Notch2 and Hes-1 were determined by qRT-PCR. The protein expressions of COX-2, N1ICD, N2ICD and Hes-1 were determined by Western blotting. Proliferation of MKN45 was detected after transfection by cell counting method. Cell apoptosis was detected by flow cytometry before and after transfection.
Results: The mRNA expression of COX-2 and Hes-1(P<0.05) but not Notch1 and Notch2(P>0.05) were significantly decreased after transfection. Inhibition of COX-2 gene expression rate was (81.81 ± 3.82)% and inhibition of Hes1 gene expression rate was (74.01 ± 4.35)%. The protein expressions of COX-2, N1ICD, N2ICD and Hes-1 were obviously inhibited(P<0.05) and the difference was statistically significant. Compared with experiment group and negative group, it can Inhibit cell proliferation and induce apoptosis after COX-2 silencing.The difference was statistically significant (P< 0.05).
Conclusion: RNA silence targeting COX-2 can inhibit cell proliferation and induce apoptosis of gastric cell line probably by inhibiting Notch signaling pathway. |
| URL | 查看原文
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| Language | 中文
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| Document Type | 学位论文
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| Identifier | https://ir.lzu.edu.cn/handle/262010/223929
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| Collection | 学院待认领
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| Affiliation | 临床医学院
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Recommended Citation
GB/T 7714 |
王璟. siRNA沉默人胃癌细胞COX-2基因对Notch信号通路影响的研究[D]. 兰州. 兰州大学,2014.
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