胃癌中E2F3相關(guān)調(diào)控網(wǎng)絡(luò)的分析及miR-320a，let-7d對(duì)其調(diào)控作用的研究
發(fā)布時(shí)間：2019-05-28 11:52
【摘要】：20世紀(jì)以來(lái),胃癌這一惡性腫瘤發(fā)病率極高,世界范圍內(nèi)胃癌患者病死率僅次于肺癌位居世界第二位。據(jù)統(tǒng)計(jì),截止2016年,70%以上的胃癌患者來(lái)自于發(fā)展中國(guó)家,側(cè)面說(shuō)明了胃癌發(fā)病與空氣、污染、水源、人口數(shù)量等生存環(huán)境息息相關(guān)。目前胃癌晚期患者的五年生存率仍然較低,因此進(jìn)一步探究胃癌細(xì)胞浸潤(rùn)生長(zhǎng)模式以及胃癌發(fā)生轉(zhuǎn)移精確的分子機(jī)制,篩選出胃癌診斷過程中易于檢測(cè)的關(guān)鍵靶點(diǎn),對(duì)于胃癌的預(yù)防和治療具有十分重要的意義。轉(zhuǎn)錄因子E2F3在促進(jìn)癌癥轉(zhuǎn)移進(jìn)程方面具有十分重要的作用,精確構(gòu)建E2F3基因相關(guān)的調(diào)控網(wǎng)絡(luò)可以幫助人們深入了解胃癌各個(gè)時(shí)期發(fā)生發(fā)展的機(jī)制。Micro RNA與相應(yīng)靶基因結(jié)合調(diào)控基因的轉(zhuǎn)錄,在腫瘤的各個(gè)時(shí)期發(fā)揮了重要作用。本研究旨在系統(tǒng)、全面的解析胃癌中E2F3相關(guān)的轉(zhuǎn)錄調(diào)控網(wǎng)絡(luò)的表達(dá)模式,并進(jìn)一步探討miR-320a和let-7d對(duì)E2F3表達(dá)水平的影響。研究通過基因芯片技術(shù)篩選胃癌組織中差異表達(dá)明顯的基因和micro RNA,結(jié)合生物信息學(xué)軟件預(yù)測(cè)E2F3相關(guān)的micro RNA、轉(zhuǎn)錄因子以及靶基因,構(gòu)建以轉(zhuǎn)錄因子E2F3為中心的轉(zhuǎn)錄調(diào)控網(wǎng)絡(luò),結(jié)合大量文獻(xiàn)富集分析調(diào)控網(wǎng)絡(luò)中E2F3與相關(guān)micro RNA之間的調(diào)控關(guān)系。生物信息學(xué)分析表明E2F3可能是miR-320a和let-7d的靶基因,通過實(shí)驗(yàn)進(jìn)一步驗(yàn)證miR-320a和let-7d與E2F3之間的調(diào)控關(guān)系。第一部分胃癌中TF-micro RNA轉(zhuǎn)錄調(diào)控網(wǎng)絡(luò)的構(gòu)建通過分析10對(duì)胃癌及癌旁正常組織的差異基因表達(dá)譜和10對(duì)micro RNA表達(dá)譜芯片數(shù)據(jù),我們篩選到809個(gè)上調(diào)和92個(gè)下調(diào)的差異表達(dá)基因,58個(gè)上調(diào)和8個(gè)下調(diào)的差異表達(dá)micro RNA。綜合TRED數(shù)據(jù)庫(kù)篩選,其中共有32個(gè)差異基因與E2F家族調(diào)控相關(guān),以差異基因?yàn)檠芯繉?duì)象,進(jìn)行功能顯著性分析和信號(hào)通路分析,此部分研究為尋找胃癌相關(guān)致病基因提供了數(shù)據(jù)信息及理論依據(jù)。結(jié)合Targetscan等micro RNA數(shù)據(jù)庫(kù)的預(yù)測(cè),篩選出miR-320a,miR-200,miR-150,miR-125a-5p,miR-92a,miR-31,miR-30,miR-17,let-7d,let-7i等10個(gè)micro RNA與E2F3相關(guān)。經(jīng)過大量文獻(xiàn)富集分析,我們推測(cè)miR-320a和let-7d極有可能通過調(diào)控E2F3從而影響胃癌的轉(zhuǎn)移分化進(jìn)程。第二部分miR-320a和let-7d對(duì)E2F3調(diào)控作用的研究通過qRT-PCR及Western blot檢測(cè)轉(zhuǎn)錄因子E2F3和miR-320a、let-7d在胃癌中的表達(dá)水平,發(fā)現(xiàn)E2F3在胃癌組織中表達(dá)量升高,其表達(dá)水平與胃癌的分化程度,臨床分期等因素顯著相關(guān);miR-320a,let-7d在胃癌組織和細(xì)胞中顯著低表達(dá),其表達(dá)水平與E2F3的表達(dá)有明顯的相關(guān)性。使用miR-320a和let-7d mimics轉(zhuǎn)染SGC-7901及MGC-803細(xì)胞系驗(yàn)證過表達(dá)miR-320a和let-7d對(duì)E2F3的影響,結(jié)果顯示在miR-320a和let-7d過表達(dá)細(xì)胞中E2F3的表達(dá)水平顯著降低,表明E2F3的表達(dá)與miR-320a,let-7d的表達(dá)水平呈顯著負(fù)相關(guān),由此可見miR-320a和let-7d可能通過調(diào)控轉(zhuǎn)錄因子E2F3的表達(dá)進(jìn)而影響胃癌的轉(zhuǎn)移分化進(jìn)程。
[Abstract]:Since the 20th century, the incidence of gastric cancer is very high, and the mortality of gastric cancer patients ranks second only to lung cancer in the world. According to statistics, up to 2016, more than 70% of patients with gastric cancer come from developing countries, which shows that the incidence of gastric cancer is closely related to air, pollution, water source, population and other living environment. At present, the 5-year survival rate of patients with advanced gastric cancer is still low, so we further explore the infiltration and growth pattern of gastric cancer cells and the accurate molecular mechanism of metastasis of gastric cancer, and screen out the key targets that are easy to detect in the diagnosis of gastric cancer. It is of great significance for the prevention and treatment of gastric cancer. Transcription factor E2F3 plays a very important role in promoting cancer metastasis. The accurate construction of E2F3 gene-related regulatory network can help people to understand the mechanism of gastric cancer occurrence and development. Micro RNA and the transcription of corresponding target gene binding regulatory genes, and play an important role in each stage of tumor. The purpose of this study was to systematically and comprehensively analyze the expression patterns of E2F3-related transcriptional regulatory networks in gastric cancer and to further explore the effects of miR-320a and let-7d on the expression of E2F3. In this study, gene chip technique was used to screen differentially expressed genes in gastric cancer tissues and micro RNA, combined with bioinformatics software to predict micro RNA, transcription factors and target genes related to E2F3, and a transcriptional regulatory network centered on transcription factor E2F3 was constructed. Combined with a large number of literature enrichment, the regulatory relationship between E2F3 and related micro RNA in the regulatory network is analyzed. Bioinformatics analysis showed that E2F3 may be the target gene of miR-320a and let-7d. The regulatory relationship between miR-320a and let-7d and E2F3 was further verified by experiments. In the first part, the construction of TF-micro RNA transcriptional regulatory network in gastric cancer was carried out by analyzing the differential gene expression profiles of 10 pairs of gastric cancer and adjacent normal tissues and the microarray data of 10 pairs of micro RNA expression profiles. We screened 809 up-regulated and 92 down-regulated differentially expressed genes, 58 up-regulated and 8 down-regulated differentially expressed micro RNA.. A total of 32 differential genes were related to the regulation of E2F family by comprehensive TRED database screening. The functional significance analysis and signal pathway analysis were carried out with the difference gene as the research object. This part of the study provides data information and theoretical basis for searching for gastric cancer related pathogenic genes. Combined with the prediction of micro RNA database such as Targetscan, 10 micro RNA, such as miR-320a,miR-200,miR-150,miR-125a-5p,miR-92a,miR-31,miR-30,miR-17,let-7d,let-7i, are selected to be related to E2F3. After a large number of literature enrichment analysis, we speculate that miR-320a and let-7d may affect the metastasis and differentiation of gastric cancer by regulating E2F3. The second part is the study on the regulatory effect of miR-320a and let-7d on E2F3. The expression of transcription factors E2F3 and miR-320a,let-7d in gastric cancer was detected by qRT-PCR and Western blot. It was found that the expression of E2F3 was increased in gastric cancer. The expression level of gastric cancer was significantly correlated with the degree of differentiation and clinical stage of gastric cancer. The expression of miR-320a,let-7d was significantly low in gastric cancer tissues and cells, and its expression level was significantly correlated with the expression of E2F3. SGC-7901 and MGC-803 cell lines were transformed with miR-320a and let-7d mimics to verify the effect of overexpression of miR-320a and let-7d on E2F3. The results showed that the expression level of E2F3 in miR-320a and let-7d overexpressed cells was significantly decreased. The results showed that the expression of E2F3 was negatively correlated with the expression of miR-320a,let-7d. It can be seen that miR-320a and let-7d may affect the metastasis and differentiation of gastric cancer by regulating the expression of transcription factor E2F3.
【學(xué)位授予單位】：吉林大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2017
【分類號(hào)】：R735.2
本文編號(hào)：2487028
[Abstract]:Since the 20th century, the incidence of gastric cancer is very high, and the mortality of gastric cancer patients ranks second only to lung cancer in the world. According to statistics, up to 2016, more than 70% of patients with gastric cancer come from developing countries, which shows that the incidence of gastric cancer is closely related to air, pollution, water source, population and other living environment. At present, the 5-year survival rate of patients with advanced gastric cancer is still low, so we further explore the infiltration and growth pattern of gastric cancer cells and the accurate molecular mechanism of metastasis of gastric cancer, and screen out the key targets that are easy to detect in the diagnosis of gastric cancer. It is of great significance for the prevention and treatment of gastric cancer. Transcription factor E2F3 plays a very important role in promoting cancer metastasis. The accurate construction of E2F3 gene-related regulatory network can help people to understand the mechanism of gastric cancer occurrence and development. Micro RNA and the transcription of corresponding target gene binding regulatory genes, and play an important role in each stage of tumor. The purpose of this study was to systematically and comprehensively analyze the expression patterns of E2F3-related transcriptional regulatory networks in gastric cancer and to further explore the effects of miR-320a and let-7d on the expression of E2F3. In this study, gene chip technique was used to screen differentially expressed genes in gastric cancer tissues and micro RNA, combined with bioinformatics software to predict micro RNA, transcription factors and target genes related to E2F3, and a transcriptional regulatory network centered on transcription factor E2F3 was constructed. Combined with a large number of literature enrichment, the regulatory relationship between E2F3 and related micro RNA in the regulatory network is analyzed. Bioinformatics analysis showed that E2F3 may be the target gene of miR-320a and let-7d. The regulatory relationship between miR-320a and let-7d and E2F3 was further verified by experiments. In the first part, the construction of TF-micro RNA transcriptional regulatory network in gastric cancer was carried out by analyzing the differential gene expression profiles of 10 pairs of gastric cancer and adjacent normal tissues and the microarray data of 10 pairs of micro RNA expression profiles. We screened 809 up-regulated and 92 down-regulated differentially expressed genes, 58 up-regulated and 8 down-regulated differentially expressed micro RNA.. A total of 32 differential genes were related to the regulation of E2F family by comprehensive TRED database screening. The functional significance analysis and signal pathway analysis were carried out with the difference gene as the research object. This part of the study provides data information and theoretical basis for searching for gastric cancer related pathogenic genes. Combined with the prediction of micro RNA database such as Targetscan, 10 micro RNA, such as miR-320a,miR-200,miR-150,miR-125a-5p,miR-92a,miR-31,miR-30,miR-17,let-7d,let-7i, are selected to be related to E2F3. After a large number of literature enrichment analysis, we speculate that miR-320a and let-7d may affect the metastasis and differentiation of gastric cancer by regulating E2F3. The second part is the study on the regulatory effect of miR-320a and let-7d on E2F3. The expression of transcription factors E2F3 and miR-320a,let-7d in gastric cancer was detected by qRT-PCR and Western blot. It was found that the expression of E2F3 was increased in gastric cancer. The expression level of gastric cancer was significantly correlated with the degree of differentiation and clinical stage of gastric cancer. The expression of miR-320a,let-7d was significantly low in gastric cancer tissues and cells, and its expression level was significantly correlated with the expression of E2F3. SGC-7901 and MGC-803 cell lines were transformed with miR-320a and let-7d mimics to verify the effect of overexpression of miR-320a and let-7d on E2F3. The results showed that the expression level of E2F3 in miR-320a and let-7d overexpressed cells was significantly decreased. The results showed that the expression of E2F3 was negatively correlated with the expression of miR-320a,let-7d. It can be seen that miR-320a and let-7d may affect the metastasis and differentiation of gastric cancer by regulating the expression of transcription factor E2F3.
【學(xué)位授予單位】：吉林大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2017
【分類號(hào)】：R735.2
【參考文獻(xiàn)】
相關(guān)期刊論文 前2條
1 尹娜,王新娟,尚永豐;中介因子復(fù)合體在基因轉(zhuǎn)錄調(diào)控中的作用[J];生理科學(xué)進(jìn)展;2003年04期
2 ;Gene expression profiles of hepatoma cell line HLE[J];World Journal of Gastroenterology;2003年04期
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