PDCD5在子宫内膜样子宫内膜癌的表达及与临床病理参数的相关性研究

发布时间：2018-06-19 19:32

本文选题：PDCD5 + 表达　；参考：《山东大学》2017年硕士论文

【摘要】：实验目的:子宫内膜癌被称为是女性的生殖系统中最为常见的恶性肿瘤其中之一。流行病学研究表明,包括高龄、初潮提前、更年期推迟、未经产、长期使用雌激素、肥胖和糖尿病在内的某些危险因素与子宫内膜癌的风险增加有关。绝大多数子宫内膜癌是源于单层上皮细胞的癌症。根据其临床特征和发病机制,子宫内膜癌大体可以分为两种类型:Ⅰ型和Ⅱ型。Ⅰ型子宫内膜癌最常发生于更年期之前和更年期左右,占子宫内膜癌的75-90%。它们通常是子宫内膜样癌,依赖雌激素,药物治疗效果较好。Ⅱ型子宫内膜癌通常发生于较年长的绝经后人群。它们通常是非子宫内膜样癌(例如浆液性癌、透明细胞癌和粘液癌等),这种类型不依赖雌激素并且预后较差。据报告,目前有很多肿瘤抑制基因和致癌基因(例如PTEN和K-ras)与子宫内膜癌的发生有着一定的联系。细胞程序性死亡5(PDCD5),一种新近发现的促凋亡基因,它是由北京大学的人类疾病基因研究中心从他们正常培养的白血病细胞株细胞中通过克隆而获得到的,它也被称为 TFAR19(TF-1 cell apoptosis related gene 19)。PDCD5 通过与Tip60(组蛋白乙酰转移酶)的相互作用促进DNA损伤诱发的细胞凋亡,由多功能激酶CK2诱导的PDCD5磷酸化是PDCD5发挥促凋亡作用的重要步骤。PDCD5的功能与抑癌基因P53有关。PDCD5与YY1相关因子2(YAF2)相互作用,促进P53介导的基因毒性应激反应。依托泊苷治疗时去泛素化酶OTU(OTUD5)与PDCD5结合,有效的介导了 PDCD5及P53的顺序激活。PDCD5与P53结合后对P53途径起正向调控作用。PDCD5选择性介导组蛋白去乙酰化酶(HDAC3)与p53解离,诱导HDAC3的裂解及泛素依赖的蛋白酶体降解。PDCD5表达受DNAJB1负相调控,DNAJB1可以靶向PDCD5抑制P53依赖的癌细胞凋亡。此外,PDCD5可通过抑制人类骨肉瘤细胞系MG-63中的Ras/Raf/MEK/ERK信号通路来抑制肿瘤的发生,携带PDCD5基因的腺病毒可对人类白血病细胞系发挥有效的抗肿瘤作用。不仅如此,而且,目前已经在多种癌症中观察到PDCD5的下调,比如说乳腺癌、卵巢癌、胃癌、肝细胞癌、急性和慢性粒细胞白血病、胶质瘤和喉鳞状细胞癌,PDCD5的下调与肿瘤的发展和预后密切相关。这些研究表明,PDCD5可能发挥了肿瘤抑制基因的作用,在癌症的发病机制中扮演重要角色。然而,PDCD5在子宫内膜癌的表达及其在临床的应用价值尚未得到充分阐述。本课题旨在利用qRT-PCR、western blot、免疫组化及免疫细胞化学检测对照子宫内膜组织、子宫内膜样子宫内膜癌组织,以及对照子宫内膜腺上皮细胞、子宫内膜癌细胞系KLE中PDCD5的表达水平,分析其与子宫内膜癌患者临床与病理特征的相关性,初步探寻PDCD5在子宫内膜癌发生、发展过程中所起到的作用。实验方法:一、采用qRT-PCR技术来检测对照子宫内膜组织与子宫内膜样子宫内膜癌中的PDCD5mRNA表达的情况二、采用western blot技术来检测对照子宫内膜组织与子宫内膜样子宫内膜癌中的PDCD5蛋白表达的情况三、采用免疫组织化学(IHC)技术来检测子对照宫内膜组织与子宫内膜样子宫内膜癌中的PDCD5蛋白表达的情况四、采用免疫细胞化学(ICC)技术来检测对照子宫内膜腺上皮细胞与子宫内膜癌细胞系KLE中的PDCD5表达的情况五、利用卡方检验来分析PDCD5表达与子宫内膜癌患者的临床病理参数的相关性实验结果:一、qRT-PCR检测对照子宫内膜组织与子宫内膜样子宫内膜癌中的PDCD5 mRNA表达水平17份对照子宫内膜组织和16份新鲜冷冻的子宫内膜样子宫内膜癌组织中的PDCD5 mRNA检测显示,正常对照子宫内膜组织和子宫内膜样子宫内膜癌组织之间的PDCD5 mRNA表达没有显著差异(P0.05)。二、Western blot检测对照子宫内膜组织与子宫内膜样子宫内膜癌中的PDCD5蛋白表达水平17份对照子宫内膜组织和16份新鲜冷冻的子宫内膜样子宫内膜癌组织中的PDCD5蛋白检测显示,子宫内膜样子宫内膜癌组织中的PDCD5蛋白的表达明显低于对照子宫内膜组织(P0.001),与qRT-PCR的结果不一致。三、IHC检测正常对照子宫内膜组织与子宫内膜样子宫内膜癌中的PDCD5蛋白表达水平1、IHC结果显示,PDCD5蛋白主要位于对照子宫内膜腺细胞或子宫内膜样子宫内膜癌细胞的细胞质中,这些细胞的胞核中也有低水平的PDCD5表达。2、51份子宫内膜样子宫内膜癌组织中的PDCD5蛋白IHC检测结果明显低于53份对照子宫内膜组织中的水平(P0.01)。3、53份对照子宫内膜组织的增殖期和分泌期之间的PDCD5表达水平没有明显差异(P0.05)。4、中、低分化的子宫内膜样子宫内膜癌组织中的PDCD5蛋白表达明显少于对照子宫内膜组织(P0.001);高分化的子宫内膜样子宫内膜癌组织和对照子宫内膜组织之间没有明显差别。四、ICC技术检测对照子宫内膜腺上皮细胞与子宫内膜癌细胞系KLE中的PDCD5表达水平1、子宫内膜腺上皮细胞表达细胞角蛋白,子宫内膜间质细胞表达波形蛋白。2、PDCD5阳性染色主要位于对照子宫内膜腺上皮细胞和KLE细胞的细胞质中,在上述细胞的胞核中也发现了 PDCD5的弱表达。KLE细胞中的PDCD5蛋白表达比对照子宫内膜腺上皮细胞中的表达弱。五、子宫内膜样子宫内膜癌组织中的PDCD5蛋白表达与患者临床病理参数的相关性PDCD5蛋白水平和年龄、子宫肌层浸润、FIGO期、雌激素受体或孕激素受体之间没有明显的关联,但与肿瘤分化程度有显著的相关性(P0.05)。与高分化子宫内膜样子宫内膜癌样本相比,中、低分化子宫内膜样子宫内膜癌中的PDCD5的表达明显降低(P0.05),这表明,PDCD5的表达可能与子宫内膜癌的发生发展相关。实验结论:一、子宫内膜样子宫内膜癌组织与对照子宫内膜组织之间的PDCD5 mRNA表达没有显著差异,但子宫内膜样子宫内膜癌组织中PDCD5蛋白比对照子宫内膜组织中减少,提示子宫内膜癌PDCD5蛋白表达可能存在转录后调控。二、子宫内膜样子宫内膜癌组织中PDCD5表达在子宫内膜的增生期和分泌期无明显差别,提示PDCD5表达可能不受卵巢激素水平的调控。三、子宫内膜样子宫内膜癌组织中PDCD5蛋白表达水平与肿瘤分化程度有关,提示PDCD5表达可能在子宫内膜样子宫内膜癌的发生发展过程中起了重要作用,并可能有助于改善预后。创新性及意义:一、本项研究首次发现PDCD5蛋白在子宫内膜样子宫内膜癌组织中表达降低,并且表达水平与肿瘤分化程度有关。二、本项研究结果提示PDCD5可能在子宫内膜癌的发生发展过程中发挥重要的作用,并有可能作为子宫内膜癌诊断和治疗的新靶点。本研究的局限性:本项研究所有结果均是临床标本及细胞的检测,缺乏体外实验和体内实验的进一步确证,有关PDCD5在子宫内膜癌中的作用及其机制还有待于进一步的探讨。
[Abstract]:Objective: endometrial cancer is known as one of the most common malignant tumors in the female reproductive system. Epidemiological studies have shown that some dangerous risk factors, including estrogen, obesity, and diabetes, are associated with increased risk of endometrial cancer, including advanced age, early menarche, postponement of menopause, and non oestrus, and the risk factors for long-term use of estrogen, obesity and diabetes. Endometrial carcinoma is a cancer of single layer epithelial cells. According to its clinical characteristics and pathogenesis, endometrial carcinoma can be divided into two types: type I and type II. Type I endometrial carcinoma is most often occurring before and around menopause, and the 75-90%. of endometrium cancer is usually endometrioid carcinoma, depending on female excitation. Type II endometrial carcinoma usually occurs in older postmenopausal women. They are usually non endometrioid carcinomas (such as serous, transparent and mucous), which are not dependent on estrogen and have a poor prognosis. It is reported that there are many tumor suppressor genes and oncogenic genes (such as PTEN, for example. And K-ras) associated with the occurrence of endometrial cancer. Cell programmed death 5 (PDCD5), a newly discovered proapoptotic gene, was obtained by cloning of the human disease gene center of Peking University from their normal cultured leukemic cell lines, and also known as TFAR19 (TF-1 cell apoptosi). S related gene 19).PDCD5 can promote apoptosis induced by DNA damage through interaction with Tip60 (histone acetyltransferase). PDCD5 phosphorylation induced by multifunctional kinase CK2 is an important step for PDCD5 to play a role in promoting apoptosis; the function of.PDCD5 is associated with the interaction of.PDCD5 and associated factor 2, which is associated with the P53 of the tumor suppressor gene. Mediated gene toxicity stress response. Etoposide treated deubiquitinase OTU (OTUD5) combined with PDCD5, effectively mediated the sequence activation of PDCD5 and P53,.PDCD5 and P53 binding to P53 pathway..PDCD5 selective mediated histone deacetylase (HDAC3) and p53 dissociation, inducing HDAC3 fragmentation and ubiquitin dependence. The expression of proteasome degradation.PDCD5 is regulated by DNAJB1 negative phase, and DNAJB1 can target PDCD5 to inhibit the apoptosis of P53 dependent cancer cells. In addition, PDCD5 can inhibit the occurrence of tumor by inhibiting the Ras/Raf/MEK/ERK signaling pathway in human osteosarcoma cell line MG-63, and adenosoncosis with PDCD5 gene can play an effective role in human leukemia cell lines. Not only that, but also the downregulation of PDCD5, such as breast, ovarian, gastric, hepatocellular carcinoma, acute and chronic granulocytic leukemia, glioma and laryngeal squamous cell carcinoma, and the downregulation of PDCD5 are closely related to the development and prognosis of the tumor. These studies suggest that PDCD5 may play an important role. The role of tumor suppressor genes plays an important role in the pathogenesis of cancer. However, the expression of PDCD5 in endometrial cancer and its clinical application have not been fully explained. This topic aims to use qRT-PCR, Western blot, immunohistochemistry and immunocytochemical test to compare endometrium and endometrium in uterus. The expression level of PDCD5 in the endometrial adenocarcinoma cell line KLE and the endometrial carcinoma cell line, and the correlation between the clinical and pathological features of endometrial carcinoma, and the preliminary exploration of the use of PDCD5 in the development and development of endometrial carcinoma. The expression of PDCD5mRNA in endometrial tissue and endometrioid carcinoma two, Western blot technique was used to detect the expression of PDCD5 protein in endometrioid carcinoma and endometrioid endometrioid carcinoma. Immunohistochemistry (IHC) was used to detect endometrium and endometrium in the endometrium. The expression of PDCD5 protein in endometrial carcinoma (four), the expression of PDCD5 in endometrial adenocarcinoma and endometrial carcinoma cell line KLE was detected by immunocytochemistry (ICC). The correlation between PDCD5 expression and the clinicopathological parameters of endometrium cancer patients was analyzed by chi square test. The expression of PDCD5 mRNA in endometrial endometrial carcinoma and endometrioid endometrioid carcinoma by qRT-PCR and PDCD5 mRNA in 17 control endometrium tissues and 16 freshly frozen endometrioid endometrioid endometrioid carcinoma tissues; PDCD5 between normal endometrium and endometrioid endometrioid carcinoma tissue There was no significant difference in mRNA expression (P0.05). Two, Western blot detected the expression level of PDCD5 protein in endometrioid endometrioid carcinoma and endometrioid endometrioid carcinoma, PDCD5 protein detection in endometrioid endometrioid carcinoma tissues of 16 fresh frozen endometrioid endometrioid carcinoma tissues and endometrioid endometrial carcinoma tissues were detected. The expression of PDCD5 protein in the endometrium was significantly lower than that of the control endometrium (P0.001), which was not consistent with the results of qRT-PCR. Three, IHC was used to detect the expression of PDCD5 protein in endometrioid endometrioid carcinoma and endometrioid endometrioid carcinoma by IHC. The results of IHC showed that the PDCD5 protein was mainly in the endometrium or endometrioid uterus of the endometrium. In the cytoplasm of endometrial cancer cells, there are also low levels of PDCD5 expression in the nucleus of these cells, and the results of PDCD5 protein IHC in endometrioid endometrioid carcinoma tissues of.2,51 are significantly lower than that in 53 controls (P0.01).3,53 portion of PDCD5 expression water between the proliferative and secretory phases of the endometrial tissue. There was no significant difference (P0.05).4. The expression of PDCD5 protein in endometrioid endometrioid carcinoma tissues of low differentiated endometrioid endometrioid carcinoma was significantly less than that of the control endometrium (P0.001), and there was no significant difference between the highly differentiated endometrioid endometrioid carcinoma tissue and the control endometrium. Four, ICC technique was used to detect the endometrium epithelium. The expression level of PDCD5 in cell and endometrial carcinoma cell line KLE was 1, the epithelial cells of endometrium expressed cytokeratin, and the endometrial stromal cells expressed vimentin.2. The positive staining of PDCD5 was mainly in the cytoplasm of the endometrial gland epithelial cells and KLE cells of the control uterus. The weak table of PDCD5 was also found in the nuclei of the above cells. The expression of PDCD5 protein in the.KLE cells was weaker than that in the control of endometrial adenoepithelial cells. Five, the expression of PDCD5 protein in endometrioid endometrial carcinoma tissues was related to the level and age of the clinicopathological parameters of the patients, PDCD5 protein level and age, the uterine myometrium infiltration, the FIGO stage, the estrogen receptor or progesterone receptor. Correlation, but has a significant correlation with the degree of tumor differentiation (P0.05). Compared with highly differentiated endometrioid endometrioid carcinoma samples, the expression of PDCD5 in endometrioid endometrioid carcinoma in low differentiated endometrioid endometrioid carcinoma is significantly decreased (P0.05), which suggests that the expression of PDCD5 may be associated with the development of endometrial carcinoma. There was no significant difference in the expression of PDCD5 mRNA between endometrial carcinoma and control endometrium, but PDCD5 protein in endometrioid endometrioid carcinoma decreased in endometrium endometrium than in control endometrium, suggesting that the expression of PDCD5 protein in endometrial carcinoma may have post transcriptional regulation. Two, PDCD5 in endometrioid endometrium carcinoma tissue. There is no obvious difference in the expression of PDCD5 expression in endometrium, suggesting that the expression of PDCD5 may not be regulated by the level of ovarian hormone. Three, the expression of PDCD5 protein in endometrioid endometrium carcinoma is related to the degree of tumor differentiation, suggesting that the expression of PDCD5 may play a role in the development of endometrioid endometrial carcinoma. The important role, and may help to improve the prognosis, innovation and significance. First, this study first found that the expression of PDCD5 protein in endometrioid endometrial carcinoma is reduced, and the level of expression is related to the degree of tumor differentiation. Two, the results of this study suggest that PDCD5 may play an important role in the development of endometrial cancer. The role, and may be a new target for the diagnosis and treatment of endometrial cancer. The limitations of this study: all the results are clinical specimens and cells, the lack of further confirmation of in vitro and in vivo experiments. The role of PDCD5 in endometrial cancer and its mechanism remain to be further explored.
【学位授予单位】：山东大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R737.33

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