本文選題：胃癌 + 蛋白質(zhì)組學(xué)��； 參考：《重慶醫(yī)科大學(xué)》2016年碩士論文

【摘要】：目的:胃癌是危害人類健康的常見(jiàn)惡性腫瘤之一,名列全球第五大常見(jiàn)腫瘤。據(jù)最新統(tǒng)計(jì),全球每年新發(fā)病例約95萬(wàn)人,死亡約72萬(wàn)人。胃癌病人的預(yù)后與腫瘤分期密切相關(guān),而腫瘤的分期與腫瘤浸潤(rùn)深度、局部淋巴結(jié)和遠(yuǎn)端臟器是否轉(zhuǎn)移相關(guān)。目前胃癌的發(fā)病及轉(zhuǎn)移的機(jī)制尚不明確。因此深入研究胃癌的發(fā)生及轉(zhuǎn)移機(jī)制顯得尤為重要。近年來(lái),蛋白質(zhì)組學(xué)技術(shù)的快速發(fā)展為我們研究疾病的發(fā)病機(jī)制提供了強(qiáng)有力的工具。本研究旨在運(yùn)用同位素標(biāo)記的相對(duì)和絕對(duì)定量技術(shù)(iTRAQ)尋找與胃癌發(fā)病相關(guān)的蛋白,探討其在胃癌發(fā)生發(fā)展中的作用機(jī)制。方法:本研究運(yùn)用iTRAQ技術(shù)對(duì)胃癌組織和癌旁組織進(jìn)行定量蛋白質(zhì)組學(xué)研究,收集15例胃癌病人的胃癌組織和癌旁組織,通過(guò)i TRAQ篩選出胃癌組織和癌旁組織中差異表達(dá)的蛋白分子。從篩選出的蛋白分子中進(jìn)一步選出幾個(gè)蛋白,運(yùn)用免疫印跡、實(shí)時(shí)定量PCR驗(yàn)證在蛋白水平、RNA水平差異蛋白的表達(dá)情況,并且用組織芯片進(jìn)行免疫組化染色在組織水平驗(yàn)證差異蛋白的表達(dá)。然后運(yùn)用胃癌細(xì)胞株模型,利用小干擾RNA技術(shù)特異性的下調(diào)蛋白的表達(dá)后觀察其對(duì)胃癌細(xì)胞株生物學(xué)行為的影響。結(jié)果:通過(guò)iTRAQ技術(shù)以及質(zhì)譜分析,共篩選出了134個(gè)差異表達(dá)的蛋白,其中51個(gè)蛋白上調(diào),83個(gè)蛋白下調(diào)。通過(guò)免疫印跡和實(shí)時(shí)定量PCR驗(yàn)證了AZU1、CPVL、OLFM4、VIL1等差異蛋白的表達(dá),發(fā)現(xiàn)AZU1、CPVL、OLFM4、VIL1在胃癌組織的表達(dá)明顯高于癌旁組織,免疫組化檢測(cè)也有同樣的結(jié)果。結(jié)果與iTRAQ結(jié)果一致。從差異蛋白中篩選出OLFM4進(jìn)行下一步的研究,在兩株胃癌細(xì)胞AGS、MKN28中通過(guò)小干擾RNA技術(shù)下調(diào)胃癌細(xì)胞株中OLFM4的表達(dá)。結(jié)果發(fā)現(xiàn)胃癌細(xì)胞株中OLFM4的表達(dá)下調(diào)后,胃癌細(xì)胞的增殖生長(zhǎng)能力,遷移和侵襲能力均明顯降低。結(jié)論:本研究成功的運(yùn)用了定量蛋白質(zhì)組學(xué)方法篩選胃癌組織與癌旁組織中差異表達(dá)的蛋白,通過(guò)進(jìn)一步的驗(yàn)證表明iTRAQ技術(shù)的結(jié)果是可靠的。同時(shí)也發(fā)現(xiàn)過(guò)表達(dá)的OLFM4在胃癌的發(fā)病和轉(zhuǎn)移過(guò)程可能起了一定的促進(jìn)作用,而降低OLFM4的表達(dá)可以抑制胃癌細(xì)胞的增殖生長(zhǎng)能力、遷移侵襲能力。
[Abstract]:Objective: gastric cancer is one of the most common malignant tumors, which is the fifth most common tumor in the world. According to the latest statistics, the world's annual new cases of about 950000 people, about 720000 deaths. The prognosis of gastric cancer patients is closely related to the tumor stage, and the tumor stage is related to the depth of tumor invasion, the metastasis of local lymph nodes and distal organs. At present, the pathogenesis of gastric cancer and metastasis is not clear. Therefore, it is very important to study the pathogenesis and metastasis of gastric cancer. In recent years, the rapid development of proteomics technology provides a powerful tool for us to study the pathogenesis of diseases. The aim of this study was to explore the role of iTRAQ in the pathogenesis and progression of gastric cancer by using the relative and absolute quantitative techniques of isotopic labeling. Methods: the iTRAQ technique was used to study the quantitative proteomics of gastric cancer tissues and paracancerous tissues in 15 patients with gastric cancer. I TRAQ was used to screen differentially expressed protein molecules in gastric cancer tissues and paracancerous tissues. Several proteins were further selected from the selected protein molecules. The expression of differential proteins at the protein level was verified by Western blot and real-time quantitative PCR. Tissue microarray immunohistochemical staining was used to verify the differential protein expression at the tissue level. Then the biological behavior of gastric cancer cell line was observed by using a small interfering RNA technique to specifically down-regulate the expression of protein in gastric cancer cell line model. Results: 134 differentially expressed proteins were screened by iTRAQ and mass spectrometry, among which 51 proteins were up-regulated and 83 proteins were down-regulated. The differential protein expression of AZU1CPVLFL-OLFM4 / VIL1 was verified by Western blot and real-time PCR. It was found that the expression of AZU1CPVLFL-OLFM4 / VIL1 was significantly higher in gastric cancer tissues than that in adjacent tissues, and the same results were obtained by immunohistochemistry. The results were consistent with those of iTRAQ. OLFM4 was screened out from the differential protein for further study. The expression of OLFM4 was down-regulated by small interfering RNA in two gastric cancer cell lines AGSN MKN28. The results showed that the ability of proliferation, migration and invasion of gastric cancer cells decreased significantly after the expression of OLFM4 was down-regulated. Conclusion: quantitative proteomics was successfully used to screen differentially expressed proteins between gastric cancer tissues and paracancerous tissues, and the results of iTRAQ technique were proved to be reliable. It was also found that overexpression of OLFM4 might play a role in the pathogenesis and metastasis of gastric cancer, while the decrease of OLFM4 expression could inhibit the proliferation and invasion of gastric cancer cells.
【學(xué)位授予單位】：重慶醫(yī)科大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2016
【分類號(hào)】：R735.2

【參考文獻(xiàn)】

相關(guān)期刊論文 前1條

1 Dana Loomis;Wei Huang;Guosheng Che;;The International Agency for Research on Cancer(IARC) evaluation of the carcinogenicity of outdoor air pollution:focus on China[J];Chinese Journal of Cancer;2014年04期

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本文編號(hào)：1869622