本文選題：胃癌 + 腫瘤標(biāo)記物��； 參考：《寧夏醫(yī)科大學(xué)》2015年碩士論文

【摘要】：目的 胃癌是目前全球最常見的惡性消化道腫瘤之一,篩查敏感性、特異性高的胃癌腫瘤標(biāo)志物,能夠達(dá)到早期發(fā)現(xiàn)、早期診斷、早期治療胃癌的目的,進(jìn)而明顯改善患者的預(yù)后和提高其生存率。目前廣泛用于臨床的腫瘤標(biāo)志物CEA、CA199、CA724等診斷胃癌的靈敏度和特異性均不理想,而近些年發(fā)現(xiàn)在胃癌中相對(duì)特異的標(biāo)志物MG7-抗原、胃蛋白酶原在我國(guó)胃癌篩查中尚無大樣本的證據(jù)支持。因此,本課題擬通過利用同位素標(biāo)記相對(duì)和絕對(duì)定量技術(shù)(i TRAQ)聯(lián)合液相色譜串聯(lián)質(zhì)譜(LC-MS/MS)蛋白組學(xué)技術(shù)篩查早期、進(jìn)展期胃癌和健康人群血清中差異表達(dá)的蛋白,并經(jīng)生物信息學(xué)分析后篩選出與胃癌發(fā)生、發(fā)展相關(guān)的特異蛋白,最后用Western blot和免疫組化方法共同驗(yàn)證,以期發(fā)現(xiàn)與胃癌分期及預(yù)后有關(guān)的蛋白標(biāo)志物。方法 用iTRAQ定量蛋白質(zhì)組學(xué)方法分析具有明顯差異的早期與進(jìn)展期胃癌患者血清各3例以及2例正常體檢者血清,登陸GO及KEGG數(shù)據(jù)庫(kù)對(duì)鑒定蛋白質(zhì)進(jìn)行生物信息學(xué)分析。通過蛋白差異表達(dá)情況、生物信息學(xué)分析并結(jié)合文獻(xiàn)查閱,選定與胃癌發(fā)生發(fā)展、信號(hào)轉(zhuǎn)導(dǎo)通路、侵襲轉(zhuǎn)移等相關(guān)的差異表達(dá)蛋白。然后從早期、進(jìn)展期胃癌與正常對(duì)照各30例血清樣本中各組隨機(jī)抽取9例血清,利用Western blot技術(shù)對(duì)其中的2個(gè)差異蛋白進(jìn)行驗(yàn)證。另收集與早期、進(jìn)展期胃癌血清對(duì)應(yīng)的術(shù)后石蠟包埋的胃癌組織及癌旁正常組織各30例,應(yīng)用免疫組化方法對(duì)2個(gè)差異蛋白進(jìn)行驗(yàn)證。最后,應(yīng)用SPSS16.0統(tǒng)計(jì)軟件分別采用t檢驗(yàn)和Kaplan-Meier生存分析法,分析2個(gè)差異蛋白的表達(dá)與胃癌患者復(fù)發(fā)轉(zhuǎn)移的關(guān)系及其對(duì)預(yù)后的影響。結(jié)果 iTRAQ定量蛋白質(zhì)組學(xué)方法共鑒定得到119個(gè)差異蛋白質(zhì)(53個(gè)上調(diào),66個(gè)下調(diào))。登陸GO數(shù)據(jù)庫(kù)搜索發(fā)現(xiàn),差異蛋白主要定位于細(xì)胞內(nèi)、細(xì)胞漿、細(xì)胞器及細(xì)胞膜等處。其功能與結(jié)合、催化、酶調(diào)節(jié)等相關(guān)。參與的生理和病理生理過程有信號(hào)傳導(dǎo)通路、代謝、單細(xì)胞生物過程、抗氧化、調(diào)節(jié)通道等。登陸KEGG pathway數(shù)據(jù)庫(kù)搜索,它們主要參與MAPK信號(hào)通路、P53信號(hào)通路等。根據(jù)篩選條件共選出7個(gè)關(guān)鍵性差異表達(dá)蛋白:IGFBP-3、ARF-1、HRG、PON3、FN1、HGF和CETP,并對(duì)其中IGFBP-3與ARF-1蛋白進(jìn)行相關(guān)驗(yàn)證。Western blot結(jié)果顯示IGFBP-3蛋白在進(jìn)展期胃癌血清中的表達(dá)低于早期胃癌與正常對(duì)照血清中的表達(dá),ARF-1蛋白在進(jìn)展期胃癌血清中的表達(dá)則高于早期胃癌與正常對(duì)照血清中的表達(dá),差異均具有統(tǒng)計(jì)學(xué)意義(P值均0.05),兩個(gè)差異蛋白在早期血清與正常對(duì)照血清中的表達(dá),經(jīng)比較差異無統(tǒng)計(jì)學(xué)意義(P0.05)。免疫組化結(jié)果顯示IGFBP-3蛋白與ARF-1蛋白陽(yáng)性表達(dá)均定位于細(xì)胞漿及部分細(xì)胞核。IGFBP-3在組織中的表達(dá)水平與Western blot結(jié)果一致,ARF-1在進(jìn)展期胃癌組織中的表達(dá)與早期胃癌、正常對(duì)照組織中的表達(dá)比較結(jié)果與Western blot結(jié)果一致,且ARF-1在早期組表達(dá)水平也高于正常組表達(dá)水平,差異具有統(tǒng)計(jì)學(xué)意義(P值0.05)。IGFBP-3與胃癌的復(fù)發(fā)轉(zhuǎn)移相關(guān),ARF-1與胃癌的臨床分期、復(fù)發(fā)轉(zhuǎn)移均相關(guān),差異均具有統(tǒng)計(jì)學(xué)意義(P值均0.05)。生存分析顯示:IGFBP-3陰性表達(dá)患者的平均生存時(shí)間明顯小于陽(yáng)性表達(dá)患者的平均生存時(shí)間,而ARF-1陰性表達(dá)患者的平均生存時(shí)間明顯大于陽(yáng)性表達(dá)患者的平均生存時(shí)間,差異均具有統(tǒng)計(jì)學(xué)意義(P值均0.05)。結(jié)論 本實(shí)驗(yàn)應(yīng)用定量蛋白質(zhì)組學(xué)i TRAQ技術(shù)研究初步篩選了早期胃癌、進(jìn)展期胃癌與正常人血清中的差異蛋白,得到了胃癌血清差異表達(dá)蛋白質(zhì)譜,共發(fā)現(xiàn)了119個(gè)差異蛋白。重點(diǎn)篩選出了7個(gè)關(guān)鍵性差異蛋白:IGFBP-3、ARF-1、HRG、PON3、FN1、HGF和CETP。對(duì)其中IGFBP-3與ARF-1兩個(gè)差異蛋白進(jìn)行相關(guān)驗(yàn)證,發(fā)現(xiàn)IGFBP-3蛋白在進(jìn)展期胃癌患者的組織和血清中均表達(dá)下調(diào),且與胃癌的復(fù)發(fā)轉(zhuǎn)移和預(yù)后相關(guān);ARF-1蛋白在胃癌患者的組織和血清中均表達(dá)上調(diào),且與胃癌的臨床分期、復(fù)發(fā)轉(zhuǎn)移和預(yù)后相關(guān)。ARF-1可能是潛在的價(jià)值較大的胃癌腫瘤標(biāo)志物,IGFBP-3雖為胃癌腫瘤標(biāo)志物的研究提供了新的線索,但其價(jià)值尚需進(jìn)一步擴(kuò)大樣本量研究證實(shí)。
[Abstract]:Objective gastric cancer is one of the most common malignant digestive tract tumors in the world at present. Screening sensitive and specific tumor markers for gastric cancer can achieve early detection, early diagnosis, early treatment of gastric cancer, and then obviously improve the prognosis and improve the survival rate of the patients. It is widely used in clinical tumor markers CEA, CA199, CA724. The sensitivity and specificity of the diagnosis of gastric cancer are not ideal, but in recent years, it is found that the relative specific marker MG7- antigen in gastric cancer is not supported by large samples in the screening of gastric cancer in our country. Therefore, we should use the relative and absolute quantitative technique of isotope labeling (I TRAQ) combined with liquid chromatography tandem mass spectrometry. LC-MS/MS) proteomics was used to screen differentially expressed proteins in the sera of early, progressive gastric cancer and healthy people. After bioinformatics analysis, specific proteins related to the development of gastric cancer were screened. Finally, Western blot and immunohistochemical method were used to verify the protein markers in order to detect the protein markers related to the stage and prognosis of gastric cancer. Methods the iTRAQ quantitative proteomics method was used to analyze the serum of 3 patients with early and progressive gastric cancer in early and progressive gastric cancer and 2 cases of normal physical examination. Bioinformatics analysis was carried out on the identification of protein by GO and KEGG database. The differential expression proteins related to the occurrence and development of gastric cancer, signal transduction pathway, invasion and metastasis, and then 9 serum samples were randomly selected from 30 serum samples of early, progressive gastric cancer and normal control, and 2 of them were verified by Western blot technique. 30 cases of paraffin embedded gastric carcinoma and 30 normal tissues adjacent to cancer were used to verify 2 differential proteins by immunohistochemical method. Finally, t test and Kaplan-Meier survival analysis were used to analyze the relationship between the expression of 2 differential proteins and the recurrence and metastasis of gastric cancer patients and the effect on prognosis. Results iTRA A total of 119 different proteins (53 up-regulated and 66 down-regulated) were identified by the Q quantitative proteomics method. The GO database search found that the differential proteins were located mainly in cells, cytoplasm, organelles and cell membranes. Their functions were related to binding, catalysis, enzyme regulation and so on. The physiological and pathophysiological processes involved were signaling pathways. Metabolism, single cell biological processes, antioxidation, regulation channels, etc.. KEGG pathway database search, they are mainly involved in the MAPK signaling pathway, P53 signaling pathway and so on. According to the screening conditions, 7 key differentially expressed proteins are selected: IGFBP-3, ARF-1, HRG, PON3, FN1, HGF and CETP. The blot results showed that the expression of IGFBP-3 protein in the serum of advanced gastric cancer was lower than that of early gastric cancer and normal control sera, and the expression of ARF-1 protein in the serum of advanced gastric cancer was higher than that in the early gastric cancer and normal control sera, and the difference was statistically significant (the value of P was 0.05), and the two differential proteins were in the early sera. The expression of serum in the normal control serum was not statistically significant (P0.05). The immunohistochemical results showed that the expression of IGFBP-3 protein and ARF-1 protein expressed in the cytoplasm and part of the nucleus.IGFBP-3 in the tissue was consistent with the Western blot results. The expression of ARF-1 in the advanced gastric cancer tissues and the early gastric cancer, The results of expression comparison in normal control tissues were consistent with the results of Western blot, and the expression level of ARF-1 in early group was also higher than that of normal group. The difference was statistically significant (P 0.05) and.IGFBP-3 was related to the recurrence and metastasis of gastric cancer. ARF-1 was correlated with the clinical stage and relapse of gastric cancer, and the difference was statistically significant (P values were all). 0.05). The survival analysis showed that the average survival time of IGFBP-3 negative expression patients was significantly less than the mean survival time of positive expression patients, while the average survival time of ARF-1 negative expression patients was significantly greater than that of positive expression patients, and the difference was statistically significant (P value was 0.05). Conclusion this experiment applied quantitative eggs. The white matter group study I TRAQ technology preliminarily screened the differential proteins in the serum of the early gastric cancer, the advanced gastric cancer and the normal human, and obtained the differential expression protein mass spectrum of the gastric cancer serum, and found 119 different proteins. The emphasis was on screening 7 key differential proteins: IGFBP-3, ARF-1, HRG, PON3, FN1, HGF and CETP., of which IGFBP-3 and ARF-1 were two. The differential protein was proved to be down regulated in the tissues and serum of the patients with advanced gastric cancer, and it was associated with the recurrence and metastasis of gastric cancer and the prognosis of the gastric cancer. The expression of ARF-1 protein in the tissues and serum of gastric cancer patients was up-regulated, and.ARF-1 may be the potential price associated with the clinical stage of gastric cancer, the recurrence and metastasis and prognosis of.ARF-1. Although the value of the tumor marker, IGFBP-3 provides a new clue for the research of tumor markers for gastric cancer, its value still needs to be further expanded to confirm the sample size.

【學(xué)位授予單位】：寧夏醫(yī)科大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2015
【分類號(hào)】：R735.2

【參考文獻(xiàn)】

相關(guān)期刊論文 前9條

1 谷艷嬌;包翠芬;魏嘉;梁佳;高志安;;ERK1/2和p-ERK1/2在肺癌組織中的表達(dá)及意義[J];中國(guó)組織化學(xué)與細(xì)胞化學(xué)雜志;2009年03期

2 李鑫;;比較蛋白質(zhì)組學(xué)研究與應(yīng)用進(jìn)展[J];國(guó)際免疫學(xué)雜志;2006年03期

3 李永文;劉紅雨;陳軍;;肺癌細(xì)胞中HGF/c-Met信號(hào)通路的異常調(diào)控及其靶向藥物研究進(jìn)展[J];中國(guó)肺癌雜志;2014年08期

4 楊麗麗;阮秋蓉;;對(duì)氧磷酶3基因研究進(jìn)展[J];中國(guó)動(dòng)脈硬化雜志;2012年10期

5 甄朱;蛋白質(zhì)組學(xué)進(jìn)展[J];生物工程學(xué)報(bào);2001年05期

6 王洋;王歡;莫佳美;蘭雪玲;褚福樓;余憲;;血清腫瘤標(biāo)志物在胃癌診斷中的價(jià)值[J];現(xiàn)代腫瘤醫(yī)學(xué);2014年04期

7 鄒小農(nóng);孫喜斌;陳萬青;鄭榮壽;張思維;代珍;劉衛(wèi)東;趙德利;;2003—2007年中國(guó)胃癌發(fā)病與死亡情況分析[J];腫瘤;2012年02期

8 林河春;張方琳;耿沁;趙方瑜;孫磊;孔韓衛(wèi);李靜;姚明;;應(yīng)用iTRAQ技術(shù)篩選高轉(zhuǎn)移和低轉(zhuǎn)移肺腺癌細(xì)胞中差異表達(dá)的膜蛋白[J];腫瘤;2013年02期

9 李艷輝;高穎;汪宏波;金志珊;;子宮內(nèi)膜癌腫瘤微環(huán)境內(nèi)IGFBP-3表達(dá)及其對(duì)腫瘤侵襲能力的影響[J];現(xiàn)代婦產(chǎn)科進(jìn)展;2014年06期

，

本文編號(hào)：1843511