本文選題：E蛋白 + 乳腺癌��； 參考：《天津醫(yī)科大學》2016年碩士論文

【摘要】：目的:E蛋白是一個新型膜蛋白,本實驗室前期通過蛋白組學分析,發(fā)現(xiàn)E蛋白與耐藥蛋白P-gp存在相互作用。本課題將在乳腺癌組織中評價E蛋白及耐藥蛋白P-gp的表達,分析E蛋白與耐藥蛋白P-gp以及臨床病理指標的相關性。同時,構建原代乳腺癌細胞培養(yǎng)方法,分析原代乳腺癌細胞中E蛋白的表達及定位與藥物敏感性的關系。此外,為更好的反應原代乳腺癌細胞對藥物的敏感性,我們將進一步比較二維及三維培養(yǎng)條件下,原代乳腺癌細胞的藥物敏感性情況,并分析E蛋白的表達與三維培養(yǎng)環(huán)境中藥物敏感性的關系。方法:1、利用免疫組化方法對87例乳腺癌患者進行指標染色,檢測E蛋白及耐藥蛋白P-gp的表達量,并分析E蛋白蛋白表達與耐藥蛋白P-gp及臨床病理之間的相關性。2、構建乳腺癌原代細胞培養(yǎng)方法,得到純度較高的原代乳腺癌細胞。3、采用MTT方法檢測不同患者來源的原代乳腺癌細胞對化療藥物(阿霉素和順鉑)的敏感性。4、采用Western Blot方法檢測不同患者來源的原代乳腺癌細胞中E蛋白的表達量,分析原代乳腺癌細胞中E蛋白的總體表達量與藥物敏感性的關系。5、采用免疫熒光方法,觀察對藥物敏感性不同的原代乳腺癌細胞中E蛋白與耐藥蛋白P-gp的表達及定位情況。6、構建Matrigel三維細胞培養(yǎng)方法,研究E蛋白表達水平與三維環(huán)境下藥物敏感性的關系。結果:1、乳腺癌組織中,E蛋白可以表達于胞膜、胞質、胞核中,E蛋白在膜漿的表達量與耐藥蛋白P-gp呈更明顯的相關性。且在E蛋白膜漿高表達組中,luminal型乳腺癌所占比例較高。2、通過膠原酶消化乳腺癌組織,梯度離心法及差異消化法進一步純化,建立原代乳腺癌細胞培養(yǎng)方法,成功培養(yǎng)原代乳腺癌細胞標本28例,其中20例獲得足夠數(shù)量的狀態(tài)良好的細胞進行后續(xù)實驗。3、對20例患者來源的原代乳腺癌細胞的總蛋白通過Western Blot方法,檢測了原代乳腺癌細胞內總體E蛋白表達水平,并計算了E蛋白/GAPDH值將患者分為E蛋白高表達組和E蛋白低表達組。4、20例患者中,對16例患者來源的原代乳腺癌細胞進行了對阿霉素的藥物敏感性檢測,12例進行了對順鉑的藥物敏感性檢測,并計算出IC50值。E蛋白表達水平與藥物敏感性數(shù)據(jù)結合分析顯示,在對DDP的藥物敏感性中,E蛋白總體水平高表達組,對順鉑的敏感性減低,耐受性增強;在對ADR的藥物敏感性中,兩組中無明顯差異。5、免疫熒光實驗發(fā)現(xiàn),在相對耐藥的患者中,E蛋白主要定位于胞膜上,同耐藥蛋白P-gp存在明顯胞膜共定位關系;在對藥物相對敏感的患者中,E蛋白在胞膜表達明顯減少,同耐藥蛋白P-gp無明顯共定位。6、三維環(huán)境中原代乳腺癌細胞對藥物的敏感性降低,在對藥物相對耐受的病例中,E蛋白主要表達于胞膜上,且E蛋白及耐藥蛋白P-gp均高表達,反之亦然。結論:1、乳腺癌組織中,E蛋白可以表達于胞膜、胞質、胞核中,E蛋白胞膜胞漿表達量與耐藥蛋白P-gp呈更明顯相關性。且在膜漿高表達組中,luminal型乳腺癌所占比例較高。2、原代乳腺癌細胞中,E蛋白總體表達水平與藥物敏感性無明顯相關性,表達于胞膜上的E蛋白調控耐藥蛋白P-gp的表達,影響細胞對藥物的敏感性。3、三維培養(yǎng)方法更能反應原代腫瘤細胞對藥物的敏感性,在三維環(huán)境中也表現(xiàn)出胞膜E蛋白高表達,細胞耐藥性增強,進一步驗證了二維培養(yǎng)條件下的結果。
[Abstract]:Objective: E protein is a new membrane protein. In the early stage of the laboratory, the interaction between E protein and drug resistant protein P-gp was found by proteomics analysis. This topic will evaluate the expression of E protein and drug resistant protein P-gp in breast cancer tissue, and analyze the correlation between E protein and drug resistant protein P-gp and the clinicopathological index. Meanwhile, the primary milk is constructed. In addition, in order to better respond to the sensitivity of the primary breast cancer cells to the drug sensitivity, we will further compare the drug sensitivity of the primary breast cancer cells in the two-dimensional and three-dimensional culture conditions and analyze the table of the E protein, in addition to the relationship between the expression of E protein in the primary breast cancer cells and the relationship between the localization of the protein and the drug sensitivity. The relationship between the drug sensitivity in the three-dimensional culture environment. Methods: 1, 87 cases of breast cancer were stained by immunohistochemical method, the expression of E protein and drug resistance protein P-gp were detected, and the correlation between the expression of E protein protein and drug resistant protein P-gp and the clinicopathology was analyzed, and the primary cell culture method of breast cancer was constructed. The MTT method was used to detect the sensitivity.4 of the primary breast cancer cells of different patients to chemotherapy drugs (adriamycin and cisplatin), and the Western Blot method was used to detect the expression of E protein in the primary breast cancer cells from different patients, and the overall table of E protein in the primary breast cancer cells was analyzed by using the Western Blot method to detect the.3 of the primary breast cancer cell with high purity. The relationship between dose and drug sensitivity.5, the expression and localization of E protein and drug resistant protein P-gp in primary breast cancer cells with different drug sensitivity were observed by immunofluorescence. The relationship between the expression of Matrigel and the relationship between the level of E protein expression and drug sensitivity in the three dimensional environment was studied. Results: 1, breast cancer group. In the fabric, E protein can be expressed in the membrane, cytoplasm and nucleus. The expression of E protein in the membrane pulp is more closely related to the drug resistant protein P-gp. And in the high expression group of E protein membrane pulp, the proportion of luminal type breast cancer is higher than.2, and further purified by collagenase digestion of breast cancer tissue, ladder centrifugation and differential digestion method, the original generation is established. In breast cancer cell culture, 28 cases of primary breast cancer cells were successfully cultured. 20 of them obtained sufficient numbers of good cells for follow-up experiment.3. The total protein of primary mammary cancer cells from 20 patients was detected by Western Blot method, and the expression of total E protein in the primary mammary gland cancer cells was measured. The /GAPDH value of E protein was divided into the high expression group of E protein and the low expression group of E protein in the.4,20 patients. The sensitivity of the drug sensitivity to adriamycin was detected in 16 cases of the primary breast cancer cells derived from the patients. The drug sensitivity of cisplatin was detected in 12 cases, and the expression of IC50 value.E protein was combined with the drug sensitivity data. The analysis showed that in the drug sensitivity to DDP, the high expression group of E protein was lower in sensitivity to cisplatin, and the tolerance was enhanced. In the sensitivity to ADR, there was no significant difference between the two groups. The immunofluorescence test found that in the relatively resistant patients, the E egg white was mainly located on the membrane, and there was a distinct cell with the drug resistant protein P-gp. In the patients with relatively sensitive drug, the expression of E protein decreased significantly in the cell membrane, and there was no obvious co localization of.6 with the drug resistant protein P-gp. The sensitivity of the breast cancer cells to the drug was reduced in the three-dimensional environment. In the cases of relative tolerance to the drug, the E protein was expressed on the cell membrane, and the E protein and the drug resistance protein P-gp were both high. Conclusion: 1, in breast cancer tissue, E protein can be expressed in the cell membrane, cytoplasm and nucleus. The cytoplasmic expression of E protein is more closely related to the drug resistant protein P-gp. In the high expression group, the proportion of luminal type breast cancer is higher.2. In the primary breast cancer cells, the overall expression level of E protein and drug sensitivity are in the primary breast cancer cells. No obvious correlation, the expression of E protein expressed on the cell membrane regulates the expression of drug resistant protein P-gp and affects the sensitivity of cell to drug sensitivity.3. The three-dimensional culture method is more responsive to the sensitivity of the original tumor cells to the drug. In the three-dimensional environment, the expression of E protein in the cell membrane is highly expressed and the cell resistance is enhanced. Further verification of the two-dimensional culture conditions The result.
【學位授予單位】：天津醫(yī)科大學
【學位級別】：碩士
【學位授予年份】：2016
【分類號】：R737.9

【相似文獻】

相關期刊論文 前10條

1 黃嘯原;用“印度閱兵”形容乳腺癌合適嗎?[J];診斷病理學雜志;2001年02期

2 張嘉慶,王殊,喬新民;乳腺癌的現(xiàn)狀和遠景[J];中華外科雜志;2002年03期

3 張維彬,汪波,石靈春;中醫(yī)藥在現(xiàn)代乳腺癌治療中的運用[J];中國中西醫(yī)結合急救雜志;2002年01期

4 薛志勇;食物與乳腺癌[J];山東食品科技;2002年04期

5 王旬果,王建軍,鄭國華;乳腺癌相關標志物的研究進展[J];山東醫(yī)藥;2002年33期

6 陸尚聞;;男人也患乳腺癌[J];環(huán)境;2003年12期

7 ;新技術清晰拍攝早期乳腺癌細胞[J];上海生物醫(yī)學工程;2005年04期

8 田富國;郭向陽;張華一;;乳腺癌診治研究新進展[J];腫瘤研究與臨床;2005年S1期

9 馬濤,谷俊朝;血管內皮生長因子與乳腺癌的臨床研究進展[J];國外醫(yī)學(外科學分冊);2005年01期

10 郭慶良,谷俊朝;乳腺癌和瘦素相關性研究進展[J];國外醫(yī)學.外科學分冊;2005年03期

相關會議論文 前10條

1 于永利;;抗乳腺癌免疫治療融合蛋白[A];中國免疫學會第四屆學術大會會議議程及論文摘要集[C];2002年

2 郭紅飛;;中醫(yī)治療乳腺癌的策略[A];江西省中醫(yī)、中西醫(yī)結合腫瘤學術交流會論文集[C];2012年

3 龐朋沙;伍會健;;乳腺癌治療靶標的研究進展[A];北方遺傳資源的保護與利用研討會論文匯編[C];2010年

4 陸勁松;邵志敏;吳炅;韓企夏;沈鎮(zhèn)宙;;新型維甲酸抑制乳腺癌細胞的生長及誘導凋亡的機制研究[A];2000全國腫瘤學術大會論文集[C];2000年

5 劉愛國;胡冰;;乳腺癌臨床治療進展[A];安徽省抗癌協(xié)會第四次代表大會暨乳腺癌、肺癌專業(yè)委員會成立會議、安徽省腫瘤防治進展學術研討會論文匯編[C];2001年

6 張嘉慶;王殊;喬新民;;乳腺癌的現(xiàn)狀和遠景[A];第一屆全國中西醫(yī)結合乳腺疾病學術會議論文匯編[C];2002年

7 劉清俊;;乳腺癌綜合治療的新進展[A];山西省抗癌協(xié)會第六屆腫瘤學術交流會論文匯編[C];2003年

8 邵志敏;;21世紀乳腺癌治療的展望[A];第三屆中國腫瘤學術大會教育論文集[C];2004年

9 陳松旺;張明;;乳腺癌治療的回顧與展望[A];西部地區(qū)腫瘤學學術會議論文匯編[C];2004年

10 白霞;傅建新;丁凱陽;王兆鉞;阮長耿;;組織因子途徑抑制物-2在乳腺癌細胞中的表達研究[A];第10屆全國實驗血液學會議論文摘要匯編[C];2005年

相關重要報紙文章 前10條

1 ;血檢有望揭示乳腺癌治療效果[N];醫(yī)藥經(jīng)濟報;2004年

2 記者 鄭曉春;乳腺癌細胞擴散基因被找到[N];科技日報;2007年

3 中國軍事醫(yī)學科學院腫瘤中心主任 宋三泰;乳腺癌有了新療法[N];中國婦女報;2002年

4 王艷紅;抑制ＤＮＡ修補可消滅乳腺癌細胞[N];醫(yī)藥經(jīng)濟報;2005年

5 詹建;乳腺癌飲食 兩個時期不一樣[N];中國中醫(yī)藥報;2006年

6 辛君;乳腺癌擴散基因“浮出水面”[N];大眾衛(wèi)生報;2009年

7 記者 毛黎;美發(fā)現(xiàn)有效抑制乳腺癌細胞生長的分子[N];科技日報;2010年

8 記者 吳春燕　通訊員 王麗霞;乳腺癌治療將有新途徑[N];光明日報;2011年

9 王樂　沈基飛;我科學家發(fā)現(xiàn)導致乳腺癌耐藥的新標志物[N];科技日報;2011年

10 劉霞;一種天然分子能阻止乳腺癌惡化[N];科技日報;2011年

相關博士學位論文 前10條

1 柴紅燕;疾病狀態(tài)下CYP4Z1和4A的生物學行為及其藥物干預研究[D];武漢大學;2012年

2 李凱;ID(inhibitor of DNA binding)家族蛋白調控乳腺細胞的分化并影響乳腺癌的預后[D];復旦大學;2014年

3 江一舟;乳腺癌新輔助化療前后基因變異檢測及其功能論證[D];復旦大學;2014年

4 馬邵;酪氨酸去磷酸化增強表皮生長因子受體在乳腺癌治療中靶向性的研究[D];山東大學;2015年

5 姚若斯;精氨酸甲基轉移酶PRMT7誘導乳腺癌細胞發(fā)生表皮—間質轉換及轉移的作用機制研究[D];東北師范大學;2015年

6 侯培鋒;α-酮戊二酸二甲酯(DM-2KG)上調缺氧誘導因子-1α(HIF-1α)誘發(fā)高致瘤性干細胞樣乳腺癌細胞機制研究[D];福建醫(yī)科大學;2014年

7 李麗麗;分泌蛋白SHON調控乳腺癌細胞EMT的分子機制研究[D];東北師范大學;2015年

8 陳麗艷;PI3K抑制劑聯(lián)合組蛋白去乙�；敢种苿⿲θ橄侔﹨f(xié)同殺傷作用的分子機制研究[D];延邊大學;2015年

9 樸俊杰;乳腺癌差異基因篩選及PAIP1對其生物學行為的影響[D];延邊大學;2015年

10 汪[?如;染色體6q25.1區(qū)域基因多態(tài)性與乳腺癌遺傳易感性的關聯(lián)研究[D];南方醫(yī)科大學;2015年

相關碩士學位論文 前10條

1 杜文英;乳腺癌分子亞型的臨床與病理特點[D];鄭州大學;2011年

2 賈曉菲;彩色多普勒超聲與乳腺癌病理及免疫組化指標的相關性研究[D];內蒙古大學;2015年

3 靳文;乳腺癌全基因組DNA甲基化修飾的研究[D];內蒙古大學;2015年

4 吳坤琳;TLR4/MyD88信號通路對乳腺癌侵襲性影響的實驗研究[D];福建醫(yī)科大學;2015年

5 葛廣哲;樹，

本文編號：2067919