本文選題：上皮性卵巢癌 + 上皮間質(zhì)轉(zhuǎn)化　； 參考：《山東大學(xué)》2016年碩士論文

【摘要】：研究背景及目的：上皮性卵巢癌是婦科惡性腫瘤中死亡率最高的腫瘤,主要原因是大多數(shù)患者確診時已發(fā)展為疾病的晚期,伴隨腹膜種植和遠處轉(zhuǎn)移。盡管近年來卵巢癌的治療已經(jīng)取得了很大進展,但是卵巢癌患者的高復(fù)發(fā)率和高轉(zhuǎn)移率使其預(yù)后并未得到明顯改善。在深入理解卵巢癌轉(zhuǎn)移的分子機制的基礎(chǔ)上,尋找可以抑制其轉(zhuǎn)移的藥物,對提高卵巢癌治療的有效率、改善其預(yù)后是十分必要的。上皮間質(zhì)轉(zhuǎn)化(epithelial-mesenchymal transition, EMT)是一種發(fā)育調(diào)控性程序,在胚胎發(fā)育、器官形成、傷口愈合等生理過程中發(fā)揮重要作用。近年來,在乳腺癌、肝癌、卵巢癌等研究中發(fā)現(xiàn),上皮性癌細胞模擬此過程后,即可獲得多種侵襲和遷移的能力,提示EMT與癌癥的侵襲和轉(zhuǎn)移密切相關(guān)。因此,尋找阻斷EMT的藥物具有潛在的臨床意義。鹽霉素是一種單羧基聚醚類抗生素,廣泛地用于雞的抗球蟲病治療。在乳腺癌的研究中發(fā)現(xiàn)鹽霉素能夠靶向殺滅乳腺癌干細胞。EMT與細胞獲得干性的密切關(guān)系提示我們EMT可能是鹽霉素靶向抑制干細胞的主要作用機制。近年來,鹽霉素抑制EMT的能力在肝癌、結(jié)腸癌、肺癌中被證實,但在卵巢癌中尚未見報道。本文旨在研究鹽霉素對人上皮性卵巢癌細胞系A(chǔ)2780和SK-OV-3增殖、侵襲、遷移等能力的影響并對其作用機制進行初步的探討。方法：CCK-8法檢測不同濃度鹽霉素(0,2,4,8,16μM)作用不同時間后A2780和SK-OV-3細胞的生存率。Transwell侵襲小室檢測鹽霉素對A2780細胞和SK-OV-3細胞侵襲和遷移能力的影響。Western-blot和RT-PCR檢測鹽霉素作用前后A2780和SK-OV-3細胞中上皮表型分子E-cadherin和Keratin,間質(zhì)表型分子N-cadherin和Vimentin以及Wnt/β-catenin信號通路相關(guān)分子表達的變化。免疫熒光方法用來定位P-catenin在卵巢癌細胞中的分布；GSK-3β抑制劑(SB216763)用來激活卵巢癌細胞中的Wnt/β-catenin信號通路。結(jié)果：(1)鹽霉素對卵巢癌細胞的增殖能力有抑制作用：應(yīng)用CCK-8法檢測不同濃度鹽霉素(0,2,4,8,16μM)作用于A2780和SK-OV-3細胞24,48,72小時后細胞的生長率,結(jié)果顯示：鹽霉素對這兩種細胞均有抑制增殖的作用,且呈時間依賴性和濃度依賴性。(2)鹽霉素對卵巢癌細胞的侵襲和遷移能力有抑制作用：應(yīng)用Transwell小室檢測鹽霉素作用前后A2780和SK-OV-3細胞侵襲和遷移能力的改變,結(jié)果顯示：與對照組相比,鹽霉素處理后的卵巢癌細胞的侵襲能力和遷移能力均明顯下降(P0.001)。(3)鹽霉素對卵巢癌細胞發(fā)生EMT有抑制作用：應(yīng)用Western-blot和RT-PCR方法檢測鹽霉素對A2780和SK-OV-3細胞上皮性分子E-cadherin、Keratin和間質(zhì)性分子N-cadherin、Vimentin表達的影響,結(jié)果顯示：與對照組相比,經(jīng)鹽霉素處理的卵巢癌細胞的上皮性分子表達明顯升高,而間質(zhì)性分子表達明顯下降,且隨著藥物濃度的增加變化越明顯。(4)鹽霉素對卵巢癌細胞中Wnt/β-catenin信號通路有阻斷作用：應(yīng)用Western-blot和RT-PCR方法檢測鹽霉素作用前后Wnt/β-catenin信號通路相關(guān)分子β-GSK-3β-ser9、β-catenin、slug、Axin2、CCND1的表達,結(jié)果顯示：與對照組相比,實驗組中各分子的表達隨著藥物濃度的增加逐漸下降；用免疫熒光方法定位鹽霉素作用前后β-catenin在卵巢癌細胞中的分布,結(jié)果顯示：與對照組相比,實驗組中細胞內(nèi)的β-catenin大部分集中于胞漿內(nèi),在胞核內(nèi)的分布明顯減少。(5)鹽霉素抑制卵巢癌細胞發(fā)生EMT的機制研究：GSK-3β抑制劑(SB216763)處理后,Wnt/β-catenin信號通路各相關(guān)分子表達均上調(diào),同時我們檢測EMT相關(guān)蛋白發(fā)現(xiàn),在兩個細胞系中,上皮表型分子均表達下調(diào)且間質(zhì)表型分子表達上調(diào)；我們將鹽霉素作用于SB216763預(yù)處理后的細胞,發(fā)現(xiàn)Wnt/β-catenin信號通路各相關(guān)分子的表達較SB216763單一處理后的細胞下調(diào)。結(jié)論：(1)鹽霉素通過抑制上皮性卵巢癌細胞發(fā)生EMT而抑制其侵襲和遷移能力。(2)鹽霉素抑制上皮性卵巢癌細胞發(fā)生EMT是通過抑制Wnt/β-catenin信號通路完成的。
[Abstract]:Background and objective: epithelial ovarian cancer is the highest death rate in gynecologic malignancies. The main reason is that most patients have been developed into late stage of the disease, accompanied by peritoneal implantation and distant metastasis. Although the treatment of ovarian cancer has been greatly expanded in recent years, the high recurrence rate and high rotation of ovarian cancer patients On the basis of understanding the molecular mechanism of ovarian cancer metastasis, it is necessary to improve the efficiency of ovarian cancer treatment and improve the prognosis of ovarian cancer. Epithelial-mesenchymal transition (EMT) is a developmental regulatory procedure. It plays an important role in embryonic development, organ formation, wound healing and other physiological processes. In recent years, in the studies of breast cancer, liver cancer, and ovarian cancer, epithelial cancer cells mimic the ability of multiple invasiveness and migration after this process, suggesting that EMT is closely related to the invasion and metastasis of cancer. Therefore, the search for drugs to block EMT is found. There is potential clinical significance. Salt mycin is a single carboxyl polyether antibiotic which is widely used in chicken anti coccidiosis treatment. In the study of breast cancer, it is found that salt mycin can target the killing of.EMT and cells in breast cancer stem cells, suggesting that EMT may be the main mechanism of salt mycin targeting to stem cells. In recent years, the ability of salt mycin to inhibit EMT has been confirmed in liver cancer, colon cancer and lung cancer, but it has not been reported in ovarian cancer. The purpose of this study is to investigate the effect of salt mycin on the proliferation, invasion and migration of human epithelial ovarian cancer cell lines, such as proliferation, invasion and migration, and to explore the mechanism of its action by CCK-8 method. The survival rate of A2780 and SK-OV-3 cells after the action of concentration of salt mycin (0,2,4,8,16 mu M).Transwell invasion of the chamber to detect the effect of salt mycin on the invasion and migration of A2780 cells and SK-OV-3 cells.Western-blot and RT-PCR detection of the epithelial phenotype molecules E-cadherin and interstitium in A2780 and SK-OV-3 cells in A2780 and SK-OV-3 cells before and after the action of salt mycin. Changes in the expression of phenotypic molecules N-cadherin and Vimentin and Wnt/ beta -catenin signaling pathway. Immunofluorescence is used to locate P-catenin in ovarian cancer cells; GSK-3 beta inhibitor (SB216763) is used to activate the Wnt/ beta -catenin signaling pathway in ovarian cancer cells. Results: (1) the proliferation of yammycin on ovarian cancer cells Ability has inhibitory effect: CCK-8 method was used to detect the growth rate of A2780 and SK-OV-3 cells after 24,48,72 hours of different concentrations of salt mycin (0,2,4,8,16 M). The results showed that the proliferation of these two cells was inhibited by salt mycin, and it was time dependent and concentration dependent. (2) the invasion and invasion of the ovarian cancer cells by the salt mycin. The migration ability was inhibited: the changes of invasion and migration ability of A2780 and SK-OV-3 cells before and after the effect of salt mycin were detected in the Transwell chamber. The results showed that the invasiveness and migration ability of the ovarian cancer cells after salt mycin treatment were significantly lower than those in the control group (P0.001). (3) the EMT of ovarian cancer cells was inhibited by salt mycin. The effect of Western-blot and RT-PCR method was used to detect the expression of E-cadherin, Keratin and interstitial molecules N-cadherin and Vimentin in A2780 and SK-OV-3 cells, and the results showed that the expression of skin molecules in the ovarian cancer cells treated by salinomycin was significantly higher than that of the control group, and the interstitial molecular table was compared with the control group. The change was obvious with the increase of drug concentration. (4) the Wnt/ beta -catenin signaling pathway in ovarian cancer cells was blocked by salt mycin: using Western-blot and RT-PCR methods to detect the expression of Wnt/ beta -catenin signaling pathway related molecules beta -GSK-3 beta -ser9, beta -catenin, slug, Axin2, CCND1, before and after the application of Western-blot and RT-PCR methods. Compared with the control group, the expression of each molecule in the experimental group decreased gradually with the increase of drug concentration; the distribution of beta -catenin in the ovarian cancer cells before and after the immunofluorescence method showed that the intracellular beta -catenin in the experimental group was mostly concentrated in the cytoplasm and in the nucleus compared with the control group. (5) the mechanism of the inhibition of EMT in ovarian cancer cells by salt mycin: after the treatment of GSK-3 beta inhibitor (SB216763), the expression of all related molecules in the Wnt/ beta -catenin signaling pathway was up-regulated, and we detected the expression of EMT related proteins. In the two cell lines, the expression of the upper cutaneous phenotypes was down and the expression of interstitial phenotypes was expressed. The effect of salt mycin on SB216763 pretreated cells showed that the expression of Wnt/ beta -catenin signaling pathway was down regulated by SB216763 single treated cells. Conclusion: (1) salt mycin inhibits the invasion and migration of epithelial ovarian cancer cells by inhibiting the occurrence of EMT in epithelial ovarian cancer cells. (2) yammycin inhibits epithelial ovarian cancer. EMT is produced by inhibiting the Wnt/ beta -catenin signaling pathway.
【學(xué)位授予單位】：山東大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2016
【分類號】：R737.31

【相似文獻】

相關(guān)期刊論文 前10條

1 張鵬偉;儲消和;郭美錦;儲炬;莊英萍;張嗣良;;響應(yīng)面設(shè)計優(yōu)化鹽霉素發(fā)酵條件[J];中國抗生素雜志;2010年02期

2 陳代杰,周光悹,許文思;鹽霉素生物合成的研究——Ⅰ.淺藍菌素對鹽霉素生物合成的影響[J];抗生素;1988年03期

3 陳代杰,周光悹,許文思;鹽霉素生物合成的研究 Ⅱ.短鏈脂肪酸在鹽霉素生物合成中作用機制的初步探討[J];抗生素;1988年03期

4 蔣順進;湯曉林;;鹽霉素發(fā)酵工藝的優(yōu)化研究[J];今日藥學(xué);2010年10期

5 高圣君;杭海峰;陳中兵;郭美錦;儲炬;莊英萍;張嗣良;;流加葡萄糖策略對鹽霉素生物合成調(diào)控影響[J];中國抗生素雜志;2013年08期

6 董秀云;徐文浩;謝志斌;殷娟;何鵬;張靜;楊菲;;成功救治急性鹽霉素中毒一例[J];臨床合理用藥雜志;2011年25期

7 劉朋;杭海峰;儲消和;郭美錦;儲炬;莊英萍;張嗣良;;鹽霉素自身抗性高產(chǎn)突變菌株篩選[J];中國抗生素雜志;2012年02期

8 董秀云;徐文浩;謝志斌;殷娟;何鵬;張靜;楊菲;;成功救治急性鹽霉素中毒一例[J];中國全科醫(yī)學(xué);2010年32期

9 楊亞勇;蔣順進;;提高白色鏈霉菌鹽霉素產(chǎn)量的工藝研究[J];中國抗生素雜志;2006年11期

10 邵明舉;邢介玲;王興蕾;邵華;劉紅;許榮廷;姜篤銀;;誤食鹽霉素中毒致死亡1例[J];山東大學(xué)學(xué)報(醫(yī)學(xué)版);2008年03期

相關(guān)會議論文 前3條

1 馬立農(nóng);陳杖榴;;高效液相色譜柱前衍生化法測定飼料中的鹽霉素[A];第四屆中國畜牧獸醫(yī)青年科技工作者學(xué)術(shù)研討會論文集[C];2001年

2 石永芝;王海;林建強;曲音波;;鹽霉素發(fā)酵補油工藝研究[A];中國資源生物技術(shù)與糖工程學(xué)術(shù)研討會論文集[C];2005年

3 陳智慧;;管碟法測定飼料中鹽霉素的不確定度評定[A];首屆中國獸藥大會動物藥品學(xué)暨中國畜牧獸醫(yī)學(xué)會動物藥品學(xué)分會2008學(xué)術(shù)年會論文集[C];2008年

相關(guān)重要報紙文章 前10條

1 山東魯抗動植物生物藥品事業(yè)部 張正海 孟偉民 朱志強;鹽霉素的應(yīng)用及前景[N];中國畜牧水產(chǎn)報;2000年

2 ;鹽霉素的應(yīng)用及前景[N];中國畜牧水產(chǎn)報;2001年

3 中國農(nóng)業(yè)科學(xué)院飼料研究所　武書庚 張海軍 李鵬 齊廣海 寧夏多維藥業(yè)有限公司　秦耀明;鹽霉素對豬生產(chǎn)性能的影響[N];中國畜牧獸醫(yī)報;2006年

4 ;鹽霉素的應(yīng)用及前景（未完待續(xù)）[N];中國畜牧水產(chǎn)報;2001年

5 $$四川農(nóng)業(yè)大學(xué)動物營養(yǎng)研究所 張克英 陳代文 $$山東魯抗動植物生物藥品事業(yè)部 張正海 周慶舉;鹽霉素對生長豬生產(chǎn)性能的影響及適宜添加量的研究[N];中國畜牧水產(chǎn)報;2001年

6 記者 陳青 通訊員 余飛;鹽霉素可抑制肝胰腺腫瘤生長[N];文匯報;2013年

7 ;鹽霉素的應(yīng)用及前景（未完待續(xù)）[N];中國畜牧水產(chǎn)報;2000年

8 ;鹽霉素的應(yīng)用及前景（未完待續(xù)）[N];中國畜牧水產(chǎn)報;2001年

9 萊州市畜牧獸醫(yī)站 原麗麗;隨意加大鹽霉素用量不可取[N];山東科技報;2006年

10 ;鹽霉素的應(yīng)用及前景（未完待續(xù)）[N];中國畜牧水產(chǎn)報;2000年

相關(guān)博士學(xué)位論文 前1條

1 馬馳;聯(lián)合應(yīng)用甲萘醌、維生素C和鹽霉素治療膠質(zhì)瘤的體外研究[D];吉林大學(xué);2014年

相關(guān)碩士學(xué)位論文 前10條

1 李睿;鹽霉素抑制上皮性卵巢癌細胞發(fā)生上皮間質(zhì)轉(zhuǎn)化的作用及其機制研究[D];山東大學(xué);2016年

2 儲消和;鹽霉素高產(chǎn)菌推理選育及發(fā)酵工藝研究[D];浙江工業(yè)大學(xué);2008年

3 張勁松;鹽霉素顆粒劑工藝的改進研究[D];天津大學(xué);2011年

4 張鵬偉;鹽霉素發(fā)酵過程優(yōu)化與放大研究[D];華東理工大學(xué);2010年

5 戚鳳娥;鹽霉素發(fā)酵工藝的調(diào)控分析與放大[D];山東大學(xué);2006年

6 石永芝;高脂酶活力鹽霉素高產(chǎn)菌株選育及發(fā)酵工藝優(yōu)化[D];山東大學(xué);2005年

7 劉朋;鹽霉素菌種選育和補油發(fā)酵工藝的優(yōu)化[D];華東理工大學(xué);2011年

8 趙福琴;鹽霉素對家兔急性、亞慢性毒性機理的研究[D];河北農(nóng)業(yè)大學(xué);2005年

9 吳丹心;鹽霉素抑制鼻咽癌細胞增殖并誘導(dǎo)凋亡的實驗研究[D];南方醫(yī)科大學(xué);2014年

10 張郭娟;鹽霉素通過調(diào)節(jié)β-catenin和mTOR信號通路抗前列腺癌干細胞的機理[D];南華大學(xué);2014年

，

本文編號：1914057