miR-224靶向RASSF8在宫颈癌进展中的作用及其机制研究
发布时间：2019-03-20 13:37
【摘要】：目的： 验证miR-224在宫颈癌组织中表达上调,并明确miR-224对宫颈癌细胞生物学行为的作用及调控机制。 方法： 采用Stem-loop RT-PCR法检测148例宫颈鳞癌组织和64例宫颈正常上皮组织的miR-224的表达水平,收集整理临床病理资料,分析miR-224的表达水平与宫颈癌临床不良预后因素的相关性。 在宫颈癌细胞株Siha和CaSki中,利用细胞转染技术过表达miR-224以及siRNA干扰RASSF8的表达,利用MTT法检测细胞增殖能力,流式细胞技术检测细胞周期分布,划痕试验和Transwell侵袭试验检测细胞的迁移和侵袭能力,并利用Real time RT-PCR和Western Blot的方法检测宫颈癌组织中IASSF8的mRNA和蛋白水平,最后利用双荧光素酶报告基因的方法确认miR-224与RASSF8靶向关系 结果： 1.miR-224在宫颈鳞癌组织的表达水平与宫颈正常上皮组织相比显著下调。miR-224高表达组有更高的FIGO分期、更大的肿瘤体积,更易于发生盆腔淋巴结转移、深间质浸润、以及脉管浸润。 2.在宫颈癌细胞株中,过表达miR-224可增加Siha和CaSki细胞的增殖能力,促进细胞划痕的愈合和细胞的侵袭并且下调RASSF8的蛋白水平,但是对周期以及凋亡无明显影响。 3.宫颈癌组织中IASSF8的表达量明显低于正常宫颈组织。 4.miR-224可下调含RASSF83'UTR区报告质粒荧光素酶的活性,而不影响3'UTR区突变报告质粒的荧光素酶活性。 5.在宫颈癌细胞中,干扰RASSF8的表达可促进Siha和CaSki细胞的迁移和侵袭能力但是对增值无影响。结论： 1.miR-224的表达水平在宫颈癌组织与正常组织相比显著上调并且miR-224高表达与宫颈癌预后不良因素相关. 2.miR-224表达上调参与了宫颈癌的进展过程,miR-224通过靶向RASSF8调控宫颈癌细胞的生物学功能。
[Abstract]:Aim: to verify the up-regulation of miR-224 expression in cervical cancer tissues and to clarify the role of miR-224 in the biological behavior of cervical cancer cells and its regulatory mechanism. Methods: the expression of miR-224 was detected by Stem-loop RT-PCR in 148 cases of cervical squamous cell carcinoma and 64 cases of normal cervical epithelium, and the clinicopathological data were collected. To analyze the correlation between the expression of miR-224 and the prognostic factors of cervical cancer. In cervical cancer cell lines Siha and CaSki, the over-expression of miR-224 and siRNA interfered with the expression of RASSF8 in cervical cancer cell line Siha and CaSki, the proliferation ability of cells was detected by MTT method, and the cell cycle distribution was detected by flow cytometry. Scratch test and Transwell invasion test were used to detect the migration and invasion ability of cells, and the mRNA and protein levels of IASSF8 in cervical cancer tissues were detected by Real time RT-PCR and Western Blot methods. Finally, the double luciferase reporter gene method was used to confirm the target relationship between miR-224 and RASSF8. The expression level of 1.miR-224 in cervical squamous cell carcinoma was significantly down-regulated compared with that in normal cervical epithelium. 224 high expression group had higher FIGO stage. Larger tumor size, more prone to pelvic lymph node metastasis, deep interstitial infiltration, and vascular infiltration. 2. In cervical cancer cell lines, overexpression of miR-224 can increase the proliferation ability of Siha and CaSki cells, promote the healing of cell scratches and invasion of cells, and down-regulate the protein level of RASSF8, but have no obvious effect on cell cycle and apoptosis. 3. The expression of IASSF8 in cervical cancer tissues was significantly lower than that in normal cervical tissues. 4.miR-224 could down-regulate the luciferase activity of the reporter plasmid containing the RASSF83' UTR region, but did not affect the luciferase activity of the mutated reporter plasmid in the 3 'UTR region. 5. In cervical cancer cells, interference with the expression of RASSF8 can promote the migration and invasion of Siha and CaSki cells, but has no effect on proliferation. Conclusion: the expression of 1.miR-224 is significantly up-regulated in cervical cancer tissues and normal tissues, and the high expression of miR-224 is associated with poor prognosis of cervical cancer. The up-regulated expression of 2.miR-224 is involved in the progression of cervical cancer. MiR-224 regulates the biological function of cervical cancer cells by targeting RASSF8.
【学位授予单位】：浙江大学
【学位级别】：硕士
【学位授予年份】：2014
【分类号】：R737.33
本文编号：2444266
[Abstract]:Aim: to verify the up-regulation of miR-224 expression in cervical cancer tissues and to clarify the role of miR-224 in the biological behavior of cervical cancer cells and its regulatory mechanism. Methods: the expression of miR-224 was detected by Stem-loop RT-PCR in 148 cases of cervical squamous cell carcinoma and 64 cases of normal cervical epithelium, and the clinicopathological data were collected. To analyze the correlation between the expression of miR-224 and the prognostic factors of cervical cancer. In cervical cancer cell lines Siha and CaSki, the over-expression of miR-224 and siRNA interfered with the expression of RASSF8 in cervical cancer cell line Siha and CaSki, the proliferation ability of cells was detected by MTT method, and the cell cycle distribution was detected by flow cytometry. Scratch test and Transwell invasion test were used to detect the migration and invasion ability of cells, and the mRNA and protein levels of IASSF8 in cervical cancer tissues were detected by Real time RT-PCR and Western Blot methods. Finally, the double luciferase reporter gene method was used to confirm the target relationship between miR-224 and RASSF8. The expression level of 1.miR-224 in cervical squamous cell carcinoma was significantly down-regulated compared with that in normal cervical epithelium. 224 high expression group had higher FIGO stage. Larger tumor size, more prone to pelvic lymph node metastasis, deep interstitial infiltration, and vascular infiltration. 2. In cervical cancer cell lines, overexpression of miR-224 can increase the proliferation ability of Siha and CaSki cells, promote the healing of cell scratches and invasion of cells, and down-regulate the protein level of RASSF8, but have no obvious effect on cell cycle and apoptosis. 3. The expression of IASSF8 in cervical cancer tissues was significantly lower than that in normal cervical tissues. 4.miR-224 could down-regulate the luciferase activity of the reporter plasmid containing the RASSF83' UTR region, but did not affect the luciferase activity of the mutated reporter plasmid in the 3 'UTR region. 5. In cervical cancer cells, interference with the expression of RASSF8 can promote the migration and invasion of Siha and CaSki cells, but has no effect on proliferation. Conclusion: the expression of 1.miR-224 is significantly up-regulated in cervical cancer tissues and normal tissues, and the high expression of miR-224 is associated with poor prognosis of cervical cancer. The up-regulated expression of 2.miR-224 is involved in the progression of cervical cancer. MiR-224 regulates the biological function of cervical cancer cells by targeting RASSF8.
【学位授予单位】：浙江大学
【学位级别】：硕士
【学位授予年份】：2014
【分类号】：R737.33
【共引文献】
相关硕士学位论文 前2条
1 初菁菁;Markov决策模型在乳腺癌筛查卫生经济学评价中的应用[D];浙江大学;2014年
2 何石林;腹腔镜与开腹肝切除术治疗肝硬化肝癌的安全性及预后分析[D];浙江大学;2014年
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