ING5抑制乳腺癌上皮间质转化机制的研究

发布时间：2018-02-01 22:44

本文关键词： 乳腺癌上皮-间质转化 ING5 侵袭转移　出处：《青岛大学》2016年博士论文　论文类型：学位论文

【摘要】：目的:探讨ING5在乳腺癌组织中的表达水平,与临床病理参数的关系,研究ING5对乳腺癌上皮间质化的影响及其作用机制。方法:(1)采集乳腺癌患者接受手术切除的癌组织标本共68例,荧光定量PCR检测ING5 m RNA的表达,分析与乳腺癌患者临床病理参数的相关性;该方法和western blot法分别检测HMEC-1、MDA-MB-231和MCF-7细胞中ING5 m RNA及其蛋白的表达。(2)构建ING5过表达乳腺癌细胞系,用Transwell小室法检测ING5过表达乳腺癌细胞侵袭和迁移,流式细胞术检测细胞凋亡,MTT法检测细胞的增殖活性。(3)荧光定量PCR和western blot法分别检测ING5过表达的人MDA-MB-231和MCF-7细胞中E-cadherin和N-cadherin m RNA和蛋白的表达;用western blot检测ING5过表达的MDA-MB-231细胞中p-PI3K、PI3K、p-Akt、Akt蛋白的表达,采用Image-Pro Plus6.0软件对其灰度值进行分析。结果:(1)乳腺癌组织中ING5 m RNA的表达显著低于癌旁组织,与肿瘤大小、临床分期有关。ING5 m RNA及其蛋白在乳腺癌细胞系MDA-MB-231和MCF-7中的表达显著低于其在HMEC-1细胞中的表达。(2)ING5过表达的细胞构建成功,Transwell小室法检测结果是ING5过表达细胞的迁移及侵袭能力显著低于对照组,NG5过表达细胞的凋亡细胞数量显著高于对照组,ING5过表达细胞的各时间点的生存率均显著降低。(3)ING5过表达细胞中的E-cadherin m RNA及其相应蛋白的表达量明显增高;而N-cadherin m RNA及其相应蛋白的表达量则明显降低。ING5过表达细胞中p-PI3K和p-Akt蛋白的表达量明显降低,灰度值分析表明p-PI3K和p-Akt蛋白的表达水平明显低于对照组,而PI3K和Akt蛋白的表达量与对照组相比无显著差异。结论:ING5在乳腺癌组织中呈异常低表达状态,上调其表达能够明显抑制细胞增殖、诱导凋亡、侵袭及迁移能力下降;对EMT具有抑制作用,而该抑制作用可能是通过抑制PI3K和Akt的磷酸化,抑制PI3K/Akt信号转导通路。
[Abstract]:Objective: to investigate the expression of ING5 in breast cancer and its relationship with clinicopathological parameters. To study the effect and mechanism of ING5 on epithelial stroma of breast cancer. The expression of ING5 m RNA was detected by fluorescence quantitative PCR, and the correlation with clinicopathological parameters of breast cancer was analyzed. This method and western blot method were used to detect HMEC-1. The expression of ING5 m RNA and its protein in MDA-MB-231 and MCF-7 cells. Transwell chamber assay was used to detect the invasion and migration of breast cancer cells overexpression of ING5 and apoptosis was detected by flow cytometry. Detection of cell proliferation activity by MTT. Fluorescence quantitative PCR and western. Detection of E-cadherin and N-cadherin in MDA-MB-231 and MCF-7 cells overexpressed by ING5 by blot method. Expression of RNA and protein; Western blot was used to detect the expression of p-PI3KPI3KPI3KPI3KPI3KPI3KPI3KP- Akt protein in MDA-MB-231 cells overexpressed by ING5. The expression of ING5 m RNA in breast cancer tissues was significantly lower than that in adjacent tissues. With the size of the tumor. The expression of .ING5m RNA and its protein in breast cancer cell line MDA-MB-231 and MCF-7 was significantly lower than that in HMEC-1 cell line. ING5 overexpression cells were successfully constructed. The results of Transwell chamber assay showed that the number of apoptotic cells of ING5 overexpression cells was significantly lower than that of control group. The survival rate of ING5 overexpression cells decreased significantly at each time point. The expression of E-cadherin m RNA and its corresponding proteins in the overexpression cells of ING5 was significantly increased. On the other hand, the expression of p-PI3K and p-Akt proteins in the overexpression cells of N-cadherin m RNA and its corresponding proteins were significantly decreased. The expression of p-PI3K and p-Akt protein was significantly lower than that of the control group. There was no significant difference in the expression of PI3K and Akt between the two groups. Conclusion the expression of PI3K and Akt is abnormally low in breast cancer, and up-regulation of the expression can significantly inhibit cell proliferation. Induced apoptosis, invasion and migration decreased; The inhibitory effect on EMT may be by inhibiting the phosphorylation of PI3K and Akt and inhibiting the PI3K/Akt signal transduction pathway.
【学位授予单位】：青岛大学
【学位级别】：博士
【学位授予年份】：2016
【分类号】：R737.9

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