LIMK2在乳腺癌侵袭转移中的作用及机制研究

发布时间：2018-04-02 08:12

本文选题：乳腺癌　切入点：增殖　出处：《第三军医大学》2016年博士论文

【摘要】：在全世界女性的所有恶性肿瘤中,乳腺癌的发病率位居第一。尽管中国女性乳腺癌的发病率相对较低,但近年来,其发病率的增长速度逐年加快,是全世界的两倍多,特别是在城市地区。目前,乳腺癌在中国女性全部恶性肿瘤的发病率中也位居第一,在全部女性恶性肿瘤导致的死因中,乳腺癌处于第六位。侵袭和转移是恶性肿瘤主要的不良生物学特征。许多癌症患者的死因也正是此。但目前对乳腺癌侵袭和转移的具体过程尚不完全清楚。因此,进一步明确乳腺癌侵袭和转移的机制,对乳腺癌的治疗和预防具有重要的意义。恶性肿瘤从形成到扩散、转移的过程是一个多水平、多因素参与的极其复杂的过程。其中癌基因在乳腺癌发生、发展中具有关键的作用。因此,探寻新的基因或重新认识原有基因的新功能,可以为深入了解恶性肿瘤的形成、侵袭和转移的具体过程;寻找最为有效的治疗靶点等提供有效帮助。转化生长因子-β可以诱导LIMK2作为连接体蛋白,使其具有促进形态发生,细胞增殖,黏附,迁移,分化和炎症作用。在人类许多恶性肿瘤组织中都检测到LIMK的过表达。目前,乳腺癌组织中LIMK2的表达情况、LIMK2与肿瘤临床病理参数及患者预后的关系仍无统一意见。而且,对LIMK2的调控机制知之甚少。针对以上问题,本研究试图运用免疫组织化学(immunohistochemistry,IHC)的方法,检测了我科212例乳腺肿瘤组织标本中LIMK2蛋白的表达情况,探讨LIMK2与肿瘤临床病理参数及患者的预后是否有关联。采用q PCR和western blot、过表达和干扰等技术初步探讨LIMK2在乳腺癌中的生物学作用及分子机制。通过上述实验试图初步阐述在乳腺癌侵袭转移过程中LIMK2所起的作用及其作用机理,为初步揭示LIMK2在乳腺癌中的侵袭性行为提供实验数据据,探讨LIMK2是否能作为治疗乳腺癌的新的分子靶点。实验方法和主要结果1.乳腺恶性肿瘤标本中LIMK2与患者临床病理参数和预后的关联方法:运用免疫组织化学(immunohistochemistry,IHC)的方法,检测了我科212例乳腺肿瘤组织标本中LIMK2蛋白的表达情况,探讨LIMK2与肿瘤临床病理参数及患者的预后是否有关联。结果:LIMK2主要表达于乳腺癌细胞的细胞质,很少见于细胞核中。LIMK2的表达与患者年龄、性别、肿瘤部位、肿瘤的病理类型和TNM分期无关(P0.05)。但是,LIMK2的高表达与乳腺癌的淋巴结转移、远处转移和肿瘤的组织学分级相关(P0.05)。乳腺癌患者中高表达LIMK2较低表达LIMK2者其生存率显著降低。另外,肿瘤大小(HR:5.27;95%CI:1.61-17.20;P=0.01),TNM分期(HR:1.84;95%CI:1.01-3.33;P=0.04)和LIMK2高表达(HR:2.83;95%CI:1.63-4.93;P=0.00)是乳腺癌患者的独立预后因素。在乳腺癌的发生和进展中,LIMK2可能起重要作用。LIMK2是乳腺癌预后不良的独立因素之一。2.LIMK2在乳腺癌侵袭、转移中的作用及作用机制的初步探讨方法:利用基因敲低和过表达、q PCR、western blot等技术,检测LIMK2对乳腺癌侵袭转移的作用,及探寻相关作用机制。结果:1.MCF-7、SUM159、MDA-MB-231细胞中均表达LIMK2,其中以SUM159细胞表达最高,MDA-MB-231细胞次之,MCF-7细胞表达最低;2.LIMK2在乳腺癌中是促癌基因,在敲低LIMK2后,乳腺癌的增殖、迁移、侵袭能力均减弱,而过表达后,乳腺癌的增殖、迁移、侵袭能力均增强;3.LIMK2可增加乳腺癌细胞的增殖、侵袭和迁移,作用途径主要是通过失活下游分子cofilin。结论1、LIMK2可增加乳腺癌细胞的增殖、侵袭和迁移,与乳腺癌的恶性程度相关。LIMK2可能是乳腺癌形成、发展中的癌基因。依据有:(1)LIMK2的高表达与乳腺癌的淋巴结转移、远处转移和肿瘤的组织学分级相关。乳腺癌患者中高表达LIMK2较低表达LIMK2者其生存率显著降低。LIMK2是乳腺癌预后不良的独立因素之一。(2)在侵袭力较强的三阴性乳腺癌MDA-MB-231细胞和SUM159细胞中,LIMK2的表达水平较高。而在恶性程度相对较低的乳腺癌MCF-7细胞中,LIMK2的表达水平较低。(3)敲低LIMK2后可减少MDA-MB-231细胞的增殖、迁移和侵袭潜能。而在MCF-7细胞中,过表达LIMK2基因可提高癌细胞的增殖、迁移和侵袭能力。2、LIMK2可增加乳腺癌细胞的增殖、侵袭和迁移,作用途径主要是通过失活下游分子cofilin。
[Abstract]:In all malignant tumors of women all over the world, the incidence of breast cancer in the first place. Although the incidence of female breast cancer Chinese rate is relatively low, but in recent years, the incidence rate of growth accelerated year by year, which is more than two times around the world, especially in the area of the city. At present, the incidence of breast cancer in Chinese all the female malignant tumor rate also ranked first, in all female malignant tumor related deaths, breast cancer in sixth. The invasion and metastasis of malignant tumor is the main adverse biological characteristics. Many cancer patients the cause of death is this. But the process of invasion and metastasis of breast cancer is not completely clear. Therefore, to further clarify the mechanism of invasion and metastasis of breast cancer, has important significance to prevention and treatment of breast cancer. The malignant tumor from the formation to the diffusion transfer process is a multi level, multi factor and pole The complex process which oncogenes in the pathogenesis of breast cancer, which plays a key role in the development. Therefore, to explore new genes or new features to recognize the original gene, for in-depth understanding of the formation of malignant tumors, the specific process of invasion and metastasis; for providing effective help for the most effective therapeutic targets. Transforming growth factor beta can induce LIMK2 as the connecting body protein, which promotes morphogenesis, cell proliferation, adhesion, migration, differentiation and inflammation. In many human malignant tumor tissues were detected the expression of LIMK. At present, the expression of LIMK2 in breast cancer, the relationship between LIMK2 and tumor clinical pathology the parameters and prognosis of patients is still no unified opinion. Moreover, little is known on the regulation mechanism of LIMK2. To solve the above problems, this study attempts to use immunohistochemistry (immunohistochemistry, IHC) detection method The expression of LIMK2 protein of 212 cases of breast cancer tissues, to explore the LIMK2 and prognosis of tumor and the clinical pathological parameters of the patients are associated with Q. PCR and Western blot, over expression and interference technology to investigate the biological effect of LIMK2 in breast cancer and its molecular mechanism. Through the experiment tentatively this invasion and its mechanism of action of LIMK2 metastasis in breast cancer, and provide experimental data according to reveal the invasive behavior of LIMK2 in breast cancer, to investigate whether LIMK2 can be used as a new molecular target for the treatment of breast cancer. Related experimental methods and main results of the 1. breast cancer specimens and LIMK2 patients with clinical pathological parameters and prognosis: using immunohistochemistry (immunohistochemistry, IHC) method, the detection of LIMK2 protein in our department in 212 cases of breast tumor tissues expression To investigate, LIMK2 and prognosis of tumor and the clinical pathological parameters of the patients are related. Results: LIMK2 was mainly expressed in the cytoplasm of breast cancer cells, the expression is very rare in the nucleus.LIMK2 and patient age, gender, tumor location, tumor pathological type and TNM staging (P0.05). However, the high expression and breast LIMK2 cancer lymph node metastasis, distant metastasis and tumor histological grading (P0.05). The high expression of LIMK2 in breast cancer patients with low expression of LIMK2 the survival rate decreased significantly. In addition, the tumor size (HR:5.27; 95%CI: 1.61-17.20; P=0.01), TNM stage (HR:1.84; 95%CI:1.01-3.33; P=0.04) and high expression LIMK2 (HR:2.83; 95%CI:1.63-4.93; P=0.00) is an independent prognostic factor in patients with breast cancer. The occurrence and progression of breast cancer, LIMK2 may play an important role in the prognosis of breast cancer is.LIMK2 adverse independent factors of.2.LIMK2 in milk Cancer invasion, metastasis and preliminary study on the role of the mechanism methods: using gene knockdown and overexpression of Q, PCR, western, blot, LIMK2 detection of invasion and metastasis of breast cancer, and explore the related mechanism. Results: 1.MCF-7, SUM159, LIMK2 were expressed in MDA-MB-231 cells, which the highest expression of SUM159 cells, MDA-MB-231 cells, MCF-7 cells was the lowest; 2.LIMK2 in breast cancer is a cancer promoting gene knockdown of LIMK2 in breast cancer, proliferation, migration and invasion ability were weakened, and after the expression of breast cancer cell proliferation, migration and invasion ability were enhanced; 3.LIMK2 can increase breast cancer cell proliferation, invasion and migration pathways, mainly through inactivation of downstream molecules cofilin. conclusion 1, LIMK2 can increase breast cancer cell proliferation, invasion and migration, the malignant degree of breast cancer and breast cancer may be related to.LIMK2 formation, development Csllt. Based on: (1) the high expression of LIMK2 in breast cancer with lymph node metastasis, distant metastasis and tumor histological grade. High expression of LIMK2 in breast cancer patients with low expression of LIMK2 significantly decreased the survival rate of.LIMK2 is the independent factors of poor prognosis in breast cancer. A (2) in an aggressive three negative breast cancer MDA-MB-231 cells and SUM159 cells, the expression level of LIMK2 is higher. The breast cancer MCF-7 cells in the malignant degree is relatively low, the expression level of LIMK2 was low. (3) knockdown of LIMK2 can reduce MDA-MB-231 cell proliferation, migration and invasion potential. In MCF-7 cells, overexpression of LIMK2 gene can enhance cancer cell proliferation, migration and invasion of.2, LIMK2 can increase the proliferation of breast cancer cells, invasion and migration pathways, mainly through inactivation of downstream molecules cofilin.

【学位授予单位】：第三军医大学
【学位级别】：博士
【学位授予年份】：2016
【分类号】：R737.9

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