【摘要】：背景：上皮性卵巢癌（epithelial ovarian carcinoma，EOC）是來源于卵巢上皮組織的惡性腫瘤，發(fā)病率占女性生殖系統(tǒng)惡性腫瘤的第三位。惡性程度高，不易早期發(fā)現(xiàn)，75%的患者就診時為FIGO III期或IV期，是死亡率最高的女性生殖系統(tǒng)腫瘤�；钚匝酰╮eactive oxygen species，ROS）是真核細胞有氧呼吸時的產(chǎn)物是正常細胞代謝自然產(chǎn)生的，由氧分子自然形成的。氧分子（molecularoxygen）通過線粒體的有氧代謝對ATP的產(chǎn)生是必需的。腫瘤細胞的快速生長和增殖會導致氧需的激增。同時又因消耗氧供而處于低氧環(huán)境。腫瘤細胞的改編碼代謝主要依賴著糖酵解和需氧呼吸的降低。因此，氧耗促進了腫瘤細胞的永生，而且直接作用于新生血管的生長以促進腫瘤生長。大量研究表明，，ROS與腫瘤的發(fā)生發(fā)展以及轉(zhuǎn)移有關(guān)1，2，包括參與上皮間充質(zhì)轉(zhuǎn)化（epithelial mesenchymaltransition，EMT）等復雜過程2，成為基礎醫(yī)學和生命科學的研究熱點。但是，在卵巢癌患者中，ROS是否參與及其作用機制目前仍不清楚。E-鈣粘蛋白（E-cadherin,簡稱E-cad）是上皮間充質(zhì)改變（epithelial-mesenchymaltransition，EMT） I型標志性分子，可維持細胞間緊密連接，阻止細胞活動侵襲及轉(zhuǎn)移擴散3。目前在腫瘤轉(zhuǎn)移通路上，有學者提出HIF-1α及LOX可能是E-cadherin的上游分子4，5。因而本研究大膽假設并驗證，HIF-1α、LOX參與ROS介導的卵巢癌間充質(zhì)轉(zhuǎn)化通路。 目的：本研究在調(diào)節(jié)ROS水平情況下，檢測相對應EOC E-cadherin水平，來研究ROS水平高低是否影響EOC EMT發(fā)生，并進一步探討其影響機制。即ROS是否會介導HIF-1α的高表達，從而上調(diào)LOX，并通過抑制E-cadherin等介導EMT使腫瘤細胞發(fā)生上皮間充質(zhì)轉(zhuǎn)化，增強腫瘤細胞的侵襲能力，促進腫瘤細胞的轉(zhuǎn)移,為上皮性卵巢癌的轉(zhuǎn)移機制提供新的理論基礎。 方法：研究分為四部分進行，前兩部分以人上皮性卵巢癌細胞株SKOV-3為切入點，探索ROS水平與EOC轉(zhuǎn)移的相關(guān)性。并通過SiRNA基因沉默技術(shù)轉(zhuǎn)染細胞、藥物干預分別抑制了HIF-1α和LOX，并測定HIF-1α、LOX及E-cadherin蛋白和或mRNA表達，推測其可能的分子機制。第三部分以動物實驗為著手，著重分析大黃素單藥以及未予藥物干預對照組小鼠間的成瘤差異，并對EMT標志分子E-cadherin進行蛋白免疫組化實驗，驗證活性氧與上皮間充質(zhì)改變和腫瘤轉(zhuǎn)移之間的關(guān)系。第四部分進行卵巢癌患者臨床資料分析，并對收集的標本進行E-cadherin蛋白的免疫組化實驗。 結(jié)論：我們發(fā)現(xiàn)，臨床標本的E-cadherin與卵巢癌分期關(guān)系密切�；谏鲜鏊牟糠謱嶒�，我們獲得以下通路推測：上皮性卵巢癌ROS↑—— HIF-1α↑—— LOX↑—— E-cadherin↑——腫瘤細胞發(fā)生EMT——侵襲性↑——腫瘤轉(zhuǎn)移
[Abstract]:Background: epithelial ovarian cancer (epithelial ovarian carcinoma,EOC) is a malignant tumor from ovarian epithelial tissue, which accounts for the third highest incidence of malignant tumor of female reproductive system. The malignant degree was high and it was not easy to detect early. 75% of the patients were in FIGO III stage or IV stage, which was the female reproductive system tumor with the highest mortality. Reactive oxygen species (reactive oxygen species,ROS) is the product of aerobic respiration in eukaryotic cells, which is naturally produced by normal cell metabolism and formed by oxygen molecules. Oxygen molecule (molecularoxygen) is necessary for ATP production through aerobic metabolism of mitochondria. The rapid growth and proliferation of tumor cells can lead to a surge in oxygen demand. At the same time, it is in a hypoxia environment because of the consumption of oxygen supply. The modification metabolism of tumor cells mainly depends on glycolysis and the decrease of aerobic respiration. Therefore, oxygen consumption promotes the immortality of tumor cells and directly acts on the growth of neovascularization to promote tumor growth. A large number of studies have shown that ROS is related to the occurrence, development and metastasis of tumors. 2, including participation in complex processes such as epithelial mesenchymal transformation (epithelial mesenchymaltransition,EMT), has become a hot research topic in basic medicine and life science. However, it is still unclear whether ROS is involved and its mechanism in patients with ovarian cancer. E-cadherin (E-cad) is the iconic molecule of epithelial mesenchyma change (epithelial-mesenchymaltransition,EMT) type I, which is known as E-cadherin (E cadherin, E cadherin). It can maintain the tight junction between cells and prevent the invasion and metastasis and diffusion of cell activity. 3. At present, in the tumor metastasis pathway, some scholars have suggested that HIF-1 偽 and LOX may be the upstream molecules of E-cadherin 4, 5. Therefore, this study boldly hypothesized and verified that HIF-1 偽 and LOX are involved in ROS-mediated mesenchymal transformation pathway in ovarian cancer. Objective: to detect the corresponding EOC E-cadherin level under the condition of adjusting ROS level, to study whether ROS level affects the occurrence of EOC EMT, and to further explore its mechanism. That is, whether ROS mediates the high expression of HIF-1 偽, thus upregulating LOX, and promoting the metastasis of tumor cells by inhibiting E-cadherin and other mediating EMT to transform tumor cells into epithelial mesenchyma, enhance the invasiveness of tumor cells and promote the metastasis of tumor cells. It provides a new theoretical basis for the metastasis mechanism of epithelial ovarian cancer. Methods: the study was divided into four parts. The first two parts were based on human epithelial ovarian cancer cell line SKOV-3 to explore the relationship between ROS level and EOC metastasis. The expression of HIF-1 偽 and LOX, were inhibited by SiRNA gene silencing technique, and the expression of HIF-1 偽, LOX, E-cadherin protein and mRNA were detected, and the possible molecular mechanism was speculated. In the third part, based on the animal experiment, the tumor formation difference between emodin single drug and control group without drug intervention was analyzed, and the protein Immunohistochemical test of EMT marker E-cadherin was carried out. To verify the relationship between reactive oxygen species (Ros) and epithelial mesenchymal changes and tumor metastasis. In the fourth part, the clinical data of patients with ovarian cancer were analyzed, and the collected samples were studied by immunohistochemistry of E-cadherin protein. Conclusion: E-cadherin in clinical specimens is closely related to the stage of ovarian cancer. Based on the above four parts of the experiment, we obtain the following pathway speculation: epithelial ovarian cancer ROS-HIF-1 偽-LOX-E-cadherin-tumor cells develop EMT-- invasive tumor metastasis-tumor metastasis
【學位授予單位】：上海交通大學
【學位級別】：碩士
【學位授予年份】：2014
【分類號】：R737.31

【共引文獻】

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