本文選題：Cystatin + B　； 參考：《復(fù)旦大學(xué)》2014年碩士論文

【摘要】：研究背景及目的：卵巢癌是導(dǎo)致女性死亡的首位生殖道惡性腫瘤,目前尚無(wú)有效的早期檢測(cè)方法及治療手段,因此尋找早期卵巢癌的生物標(biāo)志物對(duì)卵巢癌的診斷及治療具有重要意義。半胱氨酸蛋白酶抑制素B(cystatin B, stefin B, CSTB)是細(xì)胞內(nèi)半胱氨酸蛋白超家族中的一員,在多種腫瘤中均有表達(dá),而在卵巢癌中的報(bào)道尚且不多。轉(zhuǎn)化生長(zhǎng)因子-β (transforming growth factor-β, TGF-β)在卵巢癌的發(fā)生發(fā)展過程中起著重要作用。本課題主要研究CSTB的表達(dá)與卵巢腫瘤進(jìn)展的相關(guān)性、TGF-p信號(hào)轉(zhuǎn)導(dǎo)通路是否對(duì)它產(chǎn)生調(diào)控作用以及CSTB對(duì)卵巢癌細(xì)胞增殖的調(diào)控功能。方法：1.卵巢組織標(biāo)本：收集自2005年1月至2012年12月以來(lái)復(fù)旦大學(xué)附屬金山醫(yī)院的27例卵巢腫瘤患者(包括漿液性、黏液性和透明細(xì)胞性卵巢腫瘤,分為良性腫瘤組、交界性腫瘤組和惡性腫瘤組)和6例卵巢無(wú)病變患者(對(duì)照組)的卵巢石蠟標(biāo)本；2.免疫組織化學(xué)染色及分析：免疫組織化學(xué)染色法檢測(cè)上述33例卵巢組織石蠟標(biāo)本中CSTB是否表達(dá),并對(duì)染色結(jié)果進(jìn)行評(píng)分,分析其表達(dá)情況。收集患者的病史資料,對(duì)評(píng)分結(jié)果及臨床病理參數(shù)(年齡、卵巢腫瘤組織學(xué)類型、腫瘤大小、淋巴結(jié)轉(zhuǎn)移及臨床分期等方面)之間的相關(guān)性做統(tǒng)計(jì)分析；3.卵巢癌細(xì)胞株培養(yǎng),TGF-β與拮抗劑SB-431542干預(yù)：TGF-β不同濃度及時(shí)間干預(yù)上皮性卵巢癌OVCAR-3和SK-OV-3細(xì)胞株,觀察TGF-β是否調(diào)控這兩種細(xì)胞株內(nèi)CSTB的表達(dá)。進(jìn)而使用TGF-βI型受體拮抗劑SB-431542,觀察細(xì)胞內(nèi)CSTB的表達(dá)情況；4. Western blot和Real-time PCR:檢測(cè)TGF-β與其拮抗劑SB-431542干預(yù)下卵巢癌細(xì)胞株內(nèi)CSTB的表達(dá)情況；5.細(xì)胞轉(zhuǎn)染及增殖檢測(cè)：CSTB特異性的siRNA轉(zhuǎn)染卵巢癌OVCAR-3細(xì)胸.實(shí)現(xiàn)CSTB基因敲減后進(jìn)行細(xì)胞增殖檢測(cè)。檢測(cè)方法包括WST-1比色法及流式細(xì)胞儀分析細(xì)胞周期法；6.統(tǒng)計(jì)分析：所有數(shù)據(jù)應(yīng)用SPSS19.0統(tǒng)計(jì)軟件進(jìn)行分析。結(jié)果：1.CSTB在人卵巢腫瘤中過表達(dá)：免疫組織化學(xué)染色發(fā)現(xiàn),CSTB在良性、交界性和惡性卵巢腫瘤中有不同程度的表達(dá),呈現(xiàn)依次增高的趨勢(shì),在三種組織學(xué)類型的卵巢癌(漿液性、粘液性和透明細(xì)胞性卵巢癌)中呈強(qiáng)陽(yáng)性表達(dá)。卵巢腫瘤中CSTB的表達(dá)陽(yáng)性率顯著高于卵巢無(wú)病變組織(P0.01),而良性、交界性和惡性腫瘤之間的差異不顯著(P0.05)。卵巢腫瘤中CSTB的免疫組化評(píng)分亦顯著高于卵巢無(wú)病變組織(P0.01)。交界性和惡性腫瘤的CSTB免疫組化評(píng)分較良性卵巢腫瘤顯著增高(P0.05),后兩者間的差異不顯著；2.CSTB與卵巢腫瘤臨床病理參數(shù)的相關(guān)性無(wú)統(tǒng)計(jì)學(xué)意義：CSTB的表達(dá)與卵巢癌患者的臨床病理特征(如年齡、組織學(xué)類型、腫瘤大小、淋巴結(jié)轉(zhuǎn)移和臨床分期等)無(wú)顯著相關(guān)性；3. TGF-β1調(diào)控卵巢癌細(xì)胞CSTB的表達(dá),且這種調(diào)控作用可被TGF-βI型受體阻斷劑阻斷：TGF-β1干預(yù)上皮性卵巢癌OVCAR-3和SK-OV-3細(xì)胞株后,CSTB表達(dá)下調(diào),呈劑量依賴性。兩種細(xì)胞中TGF-β1最佳抑制濃度分別為10ng/ml、1 ng/ml;最佳抑制時(shí)間是24 h(梯度設(shè)定為1 h、6 h和24 h)。使用TGF-βⅠ型受體抑制劑(SB-431542)后,出現(xiàn)CSTB mRNA和蛋白的表達(dá)上調(diào)；4.CSTB促進(jìn)卵巢癌細(xì)胞的增殖：由WST-1比色法繪制細(xì)胞生長(zhǎng)曲線知,CSTB特異性的siRNA干預(yù)組出現(xiàn)細(xì)胞增殖抑制；流式細(xì)胞儀分析,細(xì)胞周期阻滯于G2/M期,細(xì)胞增殖抑制。結(jié)論：1.CSTB是卵巢腫瘤的進(jìn)展性標(biāo)志物；2.CSTB的表達(dá)受TGF-p信號(hào)轉(zhuǎn)導(dǎo)通路調(diào)控；3.CSTB可能促進(jìn)卵巢癌細(xì)胞的增殖。
[Abstract]:Research background and objective: ovarian cancer is the first reproductive tract malignant tumor that causes female death. There is no effective early detection method and treatment. Therefore, it is of great significance to find the biomarkers of early ovarian cancer for the diagnosis and treatment of ovarian cancer. Cysteine egg white enzyme inhibin B (cystatin B, stefin B, CSTB) is fine. A member of the cysteine protein superfamily is expressed in a variety of tumors, but there are not many reports in ovarian cancer. Transforming growth factor- beta (TGF- beta) plays an important role in the development of ovarian cancer. This topic is to study the relationship between the expression of CSTB and the progress of ovarian tumors. Whether or not the TGF-p signal transduction pathway regulates it and regulates the proliferation of ovarian cancer cells by CSTB. Methods: 1. ovarian tissue specimens: 27 ovarian tumor patients (including serous, mucous and transparent cell ovarian tumors) were collected from January 2005 to December 2012 at Jinshan Hospital Affiliated to Fudan University. Ovarian paraffin specimens of benign tumor group, borderline tumor group and malignant tumor group) and 6 cases of ovarian non pathological patients (control group); 2. immunohistochemical staining and analysis: immunohistochemical staining method was used to detect the expression of CSTB in the paraffin specimens of the 33 cases of ovarian tissue, and the staining results were scored and the expression was analyzed. A statistical analysis of the correlation between scores and clinicopathological parameters (age, ovarian tumor histology type, tumor size, lymph node metastasis and clinical staging) was collected, and 3. ovarian cancer cell lines were cultured, TGF- beta and antagonist SB-431542 intervention: TGF- beta concentration and time intervention in epithelial ovarian ovary OVCAR-3 and SK-OV-3 cell lines, observe whether TGF- beta regulates the expression of CSTB in these two cell lines, and then uses TGF- beta I receptor antagonist SB-431542 to observe the expression of CSTB in the cell, and 4. Western blot and Real-time PCR: to detect the expression of the ovarian cancer cell line under the interference of beta and its antagonist; 5. thin. Cell transfection and proliferation detection: CSTB specific siRNA transfected OVCAR-3 fine chest. Cell proliferation detection was performed after CSTB knockout. The detection methods included WST-1 colorimetric assay and flow cytometry analysis of cell cycle method; 6. statistical analysis: all data were analyzed by SPSS19.0 statistical soft parts. Results: 1.CSTB in human ovarian swelling. Over expression in the tumor: immunohistochemical staining showed that CSTB was expressed in a different degree in benign, borderline and malignant ovarian tumors, showing a trend in turn, strongly positive in three histological types of ovarian cancer (serous, mucinous and transparent cell ovarian cancer). The positive rate of CSTB in ovarian tumors was significant. The difference between borderline and malignant tumor was not significant (P0.05). The immunohistochemical score of CSTB in ovarian tumors was also significantly higher than that of ovarian nonpathological tissue (P0.01). The CSTB immunohistochemical score of borderline and malignant tumors was significantly higher than that of benign ovarian tumors (P0.05). The difference between the two groups was not significant (P0.05). There was no significant correlation between 2.CSTB and the clinicopathological parameters of ovarian tumors. There was no significant correlation between the expression of CSTB and the clinicopathological features of ovarian cancer patients (such as age, histological type, tumor size, lymph node metastasis and clinical staging, etc.); 3. TGF- beta 1 regulates the expression of CSTB in ovarian cancer cells, and this regulation can be used. Blocked by TGF- beta I receptor blocker: TGF- beta 1 interfered with OVCAR-3 and SK-OV-3 cell lines in epithelial ovarian cancer, CSTB expression was downregulated and dose-dependent. The optimal inhibitory concentration of TGF- beta 1 in two cells was 10ng/ml and 1 ng/ml, and the best inhibition time was 24 h (1 h, 6 h and 24). After that, the expression of CSTB mRNA and protein was up-regulated; 4.CSTB promoted the proliferation of ovarian cancer cells: the cell growth curve was plotted by WST-1 colorimetry, and the proliferation inhibition of CSTB specific siRNA intervention group appeared; flow cytometer analysis, cell cycle arrest in G2/M phase and cell proliferation inhibition. Conclusion: 1.CSTB is the progress of ovarian tumor. The expression of 2.CSTB is regulated by TGF-p signal transduction pathway; 3.CSTB may promote the proliferation of ovarian cancer cells.
【學(xué)位授予單位】：復(fù)旦大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2014
【分類號(hào)】：R737.31

【共引文獻(xiàn)】

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1 徐嘉文;葉海燕;陳建國(guó);;miRNA-200在卵巢癌中的研究進(jìn)展[J];廣東醫(yī)學(xué);2012年02期

2 高娜;吳濤;;TGF-β在卵巢癌中作用的研究進(jìn)展[J];大連醫(yī)科大學(xué)學(xué)報(bào);2012年04期

3 劉貴鵬;張春梅;趙曉東;;轉(zhuǎn)化生長(zhǎng)因子β_1及其Ⅰ型和Ⅱ型受體在子宮內(nèi)膜腺癌中的表達(dá)及意義[J];廣東醫(yī)學(xué);2013年02期

4 曾孔鑫;石巍;胡光勝;;胃癌組織中TGFBR1的基因突變研究[J];中南醫(yī)學(xué)科學(xué)雜志;2014年02期

5 楊建華,石一復(fù);卵巢癌的病因流行病學(xué)研究進(jìn)展[J];國(guó)外醫(yī)學(xué).婦幼保健分冊(cè);2004年06期

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4 劉霞;山西省婦女卵巢癌危險(xiǎn)因素的調(diào)查研究[D];山西醫(yī)科大學(xué);2008年

5 王瑞雪;卵巢惡性腫瘤術(shù)后化療患者60例證候分布特點(diǎn)及影響因素初步研究[D];成都中醫(yī)藥大學(xué);2009年

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