本文選題：上皮細胞粘附分子　切入點：前列腺癌　出處：《蘇州大學》2016年博士論文　論文類型：學位論文

【摘要】：目的:前列腺癌是目前西方社會導致死亡的第二大因素。盡管在過去數(shù)十年中,前列腺癌患者的總生存率有顯著提高,但是最近的數(shù)據(jù)表明轉移性前列腺癌患者生存的改善并沒有對死亡率的下降產生顯著影響。上皮細胞粘附分子(EpCAM,Epithelial cell adhesion molecule)是一種上皮細胞跨膜糖蛋白,最初是以一種腫瘤相關抗原而被發(fā)現(xiàn)的,由于EpCAM在多種腫瘤組織中的表達升高而備受關注。有較多研究成果顯示EpCAM在腫瘤的發(fā)生、發(fā)展中發(fā)揮重要作用,有望作為腫瘤診斷的分子標志物和可能的治療靶點。因此本文選擇EpCAM作為研究對象,分析其在前列腺癌特別是雄激素依賴性前列腺癌發(fā)生與發(fā)展中的作用。方法:選取我中心近五年內接受前列腺癌根治術的50例患者的手術標本,標本包括癌和癌旁組織,均經(jīng)冰凍連續(xù)切片首尾病理證實為癌或正常組織后留取標本,其中術前血清PSA≤10ng/ml 23例、PSA10ng/ml 27例,T2期18例、T3期24例、T4期8例,淋巴結轉移18例,無遠處轉移病例。通過Western-blot方法和免疫組化檢測50對癌和癌旁組織中EpCAM蛋白表達的差異,并且定期隨訪患者術后PSA等預后情況。在體外細胞水平,運用Western-blot方法檢測3種人前列腺癌細胞株(LNCap、DU-145、PC-3)中EpCAM蛋白表達的水平,并分析其差異;設計shRNA,以慢病毒為載體構建EpCAM沉默的LNCap穩(wěn)轉細胞株,并運用流式細胞技術檢測細胞周期,運用細胞功能學實驗檢測細胞增殖、侵襲和轉移能力。通過Western-blot方法檢測EpCAM及雄激素受體(Androgen receptor,AR)、前列腺特異性抗原(prostate specific antigen,PSA)水平,細胞周期相關蛋白,以及上皮-間質轉化(EMT)蛋白標志物的表達水平,深入探討EpCAM對前列腺癌細胞生物學特性的影響。結果:EpCAM在前列腺癌組織及癌旁組織中均有表達,但EpCAM在前列腺癌組織中的表達明顯高于癌旁前列腺組織,差異具有統(tǒng)計學意義(P0.05)。EpCAM蛋白的高表達與前列腺癌的手術前血清中總的PSA水平、臨床病理分級、淋巴結轉移以及生化復發(fā)情況等具有明顯相關性,且EpCAM高表達的前列腺癌患者,其生化復發(fā)時間顯著短于EpCAM低表達的前列腺癌患者。3種前列腺癌細胞系DU-145、PC-3、LNCaP中雄激素依賴性前列腺癌細胞LNCaP的EpCAM表達明顯高于其它兩種雄激素抵抗性前列腺癌細胞(P0.05)。運用shRNA穩(wěn)定轉染LNCaP細胞后,相較空白對照組和陰性對照組,實驗組EpCAM明顯下調,而且細胞周期明顯阻滯在G1期,且LNCaP細胞的增殖及侵襲能力明顯受到抑制。LNCaP中沉默EpCAM的表達后,AR和PSA的表達水平顯著降低,細胞周期相關蛋白Cycling D和E表達水平明顯降低,上皮細胞特異性蛋白E-cadherin的水平上升而Vimentin、N-cadherin水平下降。結論:高表達水平的EpCAM與前列腺癌疾病的演變,特別是雄激素依賴性前列腺癌的發(fā)生發(fā)展有關。其過表達可促進前列腺癌細胞周期進展,增強前列腺癌細胞的增殖及侵襲能力,干擾EpCAM可能是激素依賴性前列腺癌治療的一個靶點。此外,EpCAM的過表達可促進AR和PSA表達,可能與雄激素非依賴性前列腺癌的發(fā)生相關,因此EpCAM具有重要的研究價值,為將來前列腺癌細胞的發(fā)生和發(fā)展提供更多的理論依據(jù),及為研制該階段有效的藥物提供了可靠的靶點。
[Abstract]:Objective: prostate cancer is the second leading cause of death in western society. Although in the past few decades, was significantly higher in patients with prostate cancer survival rates, but recent data suggest that prostate cancer metastasis survival patients do not have a significant impact on mortality decreased. The epithelial cell adhesion molecule (EpCAM, Epithelial cell adhesion molecule) is a kind of epithelial cell membrane glycoprotein, originally as a tumor associated antigen was found, due to the increased expression of EpCAM in tumor tissues and concern. There are more research results show that EpCAM in the incidence of cancer, plays an important role in the development, is expected to serve as molecular tumor diagnosis signs and possible therapeutic target. So this paper chooses EpCAM as the research object, analysis of the prostate cancer is androgen dependent prostate cancer occurrence and development Show in vitro. Methods: the center for radical prostatectomy in 50 cases of surgical specimens in the past five years, including specimens of carcinoma and adjacent tissues, were confirmed by pathology and frozen serial sections for cancer or normal tissue specimens, including preoperative serum PSA 10ng/ ml in 23 cases. PSA10ng/ml 27 cases, T2 18 cases, T3 24 cases, T4 8 cases, 18 cases with lymph node metastasis, distant metastasis cases. By using the methods of Western-blot and immunohistochemistry to detect EpCAM protein expression in 50 carcinoma and paracancerous tissues, and regular follow-up of patients with postoperative prognosis in PSA. In vitro, using Western-blot method to detect 3 kinds of human prostate cancer cell lines (LNCap, DU-145, PC-3) level of expression of EpCAM protein, and to analyze the difference; design of shRNA, a lentivirus vector stably transfected cell line EpCAM silencing of LNCap, and using flow cytometry. Cell cycle, cell function by cell proliferation assay, invasion and metastasis. Detected by Western-blot EpCAM and androgen receptor (Androgen receptor, AR), prostate specific antigen (prostate specific, antigen, PSA) level, cell cycle related protein, and epithelial mesenchymal transition (EMT) protein expression level mark the in-depth study of effect of EpCAM on cell biological characteristics of prostate cancer. The results showed the expression of EpCAM in prostate cancer tissues and paracancerous tissues, but the expression of EpCAM in prostate cancer tissues was significantly higher than adjacent noncancerous prostate tissue, the difference was statistically significant (P0.05) and the high expression of.EpCAM protein in prostate cancer surgery the serum total PSA level, clinical pathological grading, lymph node metastasis and biochemical recurrence and had a significant correlation, and the high expression of EpCAM in prostate cancer patients, the biochemical The recurrence time was significantly shorter than that of.3 in patients with prostate cancer DU-145 and prostate cancer cell lines with low expression of EpCAM PC-3 and LNCaP in androgen independent prostate cancer cell LNCaP EpCAM expression was significantly higher than the other two androgen resistant prostate cancer cells (P0.05). Using shRNA stable transfected LNCaP cells compared with the blank control group and negative control group the experimental group, EpCAM was significantly decreased, and the cell cycle was arrested in G1 phase, and the proliferation and invasion of LNCaP cells was significantly inhibited to silence the expression of EpCAM in.LNCaP, the expression level of AR and PSA were significantly decreased and the expression of cell cycle related protein Cycling, D and E were significantly decreased, increased levels of epithelial cell specific protein E-cadherin and Vimentin, N-cadherin levels decreased. Conclusion: the evolution of a high level of EpCAM expression in prostate cancer and disease, especially in androgen dependent prostate cancer Related to the development. Its overexpression may promote the progression of prostate cancer cell cycle, enhance the ability of proliferation and invasion of prostate cancer cells, EpCAM interference may be a target for hormone dependent prostate cancer treatment. In addition, the overexpression of EpCAM may promote the expression of AR and PSA may be related to androgen independent prostate cancer therefore, EpCAM has important research value, provides theoretical basis for the occurrence and development of prostate cancer cells in the future, and for the development of the effective drug target provides reliable.

【學位授予單位】：蘇州大學
【學位級別】：博士
【學位授予年份】：2016
【分類號】：R737.25

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