ILF2在肝細(xì)胞肝癌中的生物學(xué)功能及分子機(jī)制研究
發(fā)布時間：2018-07-27 12:23
【摘要】：背景在我國,肝細(xì)胞肝癌(Hepatocellular carcinoma,HCC)是一種高發(fā)的惡性腫瘤,90%的患者都有乙型肝炎的病史。中國每年死于肝癌約11萬人,占全世界肝癌死亡人數(shù)的45%。肝癌患者初期癥狀并不明顯,然而一旦出現(xiàn)癥狀,其病程大多已進(jìn)入晚期,如果惡性程度高則其預(yù)后更差。雖然近幾十年在肝癌切除、放射療法、化學(xué)療法及肝臟移植等治療手段取得諸多進(jìn)展,但是肝癌患者的長期預(yù)后仍然不理想。預(yù)后差的主要原因是肝癌的有效治療靶點(diǎn)未找到以及肝癌發(fā)生發(fā)展的分子機(jī)制不明了。因此,尋找更具敏感性和特異性的肝癌的分子標(biāo)志物和探究其在肝癌發(fā)生發(fā)展中的作用機(jī)制,將是進(jìn)一步提高肝癌生存率的關(guān)鍵,對肝癌今后的早期診斷和早期治療有著深遠(yuǎn)影響。目的本研究目的是探討Interleukin enhancer binding factor 2(ILF2)在我院進(jìn)行肝癌肝切除患者的肝癌組織和癌旁組織,以及肝癌細(xì)胞株中的表達(dá)水平,進(jìn)而探索其與患者相關(guān)的各病理指數(shù)之間的聯(lián)系,在TCGA數(shù)據(jù)庫中分析其與肝癌患者預(yù)后的關(guān)系。用蛋白酶體抑制劑MG132分別在0,2,4,8,24h五個不同時間段處理肝癌細(xì)胞系,驗(yàn)證ILF2在肝癌組織及肝癌細(xì)胞中的高表達(dá)是通過轉(zhuǎn)錄水平引起。利用細(xì)胞學(xué)實(shí)驗(yàn)和體內(nèi)成瘤技術(shù),深度闡明ILF2在肝癌肝癌發(fā)生發(fā)展過程中所發(fā)揮的作用,以及其在肝癌形成中存在的信號通路。通過干擾ILF2的表達(dá)以及對肝癌細(xì)胞的培養(yǎng)基中添加OμM,5μM,1OμM,20μM不同濃度的人表皮生長因子受體(EGFR)的抑制劑厄洛替尼,探討兩種處理對細(xì)胞活性和增殖能力的協(xié)同作用。基于表達(dá)譜芯片的數(shù)據(jù),對下游差異基因進(jìn)行GO分析,利用KEGG數(shù)據(jù)庫對差異基因進(jìn)行Pathway分析,從而對ILF2下游分子或下游通路進(jìn)行深一步分析和研究。方法1.提取27例HCC患者的肝癌肝切腫瘤組織及配對癌旁組織的mRNA,收集另外72例肝癌病人的腫瘤及癌旁石蠟組織,提取L02,Huh7,Bel-7402,MHCC-LM3,SMCC-7721和SK-HEP-1細(xì)胞系中的mRNA和8對肝癌組織及癌旁組織的蛋白質(zhì),利用Western免疫印跡、實(shí)時熒光定量PCR、免疫組化等技術(shù)驗(yàn)證白介素增強(qiáng)結(jié)合因子2(ILF2)在HCC腫瘤組織及各肝癌細(xì)胞系中的表達(dá)情況。2.根據(jù)肝癌組織的免疫組化結(jié)果,探究ILF2與肝癌病人臨床病理參數(shù)之間的關(guān)系。3.利用TCGA數(shù)據(jù)庫分析ILF2與肝癌患者生存率的關(guān)系。4.通過蛋白酶體抑制劑MG132處理肝癌細(xì)胞系,探索ILF2的表達(dá)升高是否通過轉(zhuǎn)錄水平的上調(diào)引起。5.通過轉(zhuǎn)染ILF2小干擾RNA及慢病毒穩(wěn)定轉(zhuǎn)染過表達(dá)ILF2的方式,分別干擾或過表達(dá)ILF2,通過細(xì)胞學(xué)實(shí)驗(yàn),如細(xì)胞克隆積聚實(shí)驗(yàn)、細(xì)胞凋亡實(shí)驗(yàn)和Cell Counting Kit-8(CCK-8)等,探索在肝癌細(xì)胞增殖和細(xì)胞凋亡的調(diào)控過程中ILF2所發(fā)揮的作用。6.為了探究ILF2在體內(nèi)成瘤過程中的作用和影響,我們在裸鼠體內(nèi)進(jìn)行腫瘤細(xì)胞瘤體生成實(shí)驗(yàn)。7.運(yùn)用Western blot技術(shù)尋找ILF2調(diào)控的下游分子,來探索ILF2在肝癌中發(fā)揮作用的分子機(jī)制。8.通過敲低ILF2的表達(dá)以及不同濃度的厄洛替尼處理肝癌細(xì)胞,運(yùn)用Cell CountingKit-8(CCK-8)和克隆形成實(shí)驗(yàn),驗(yàn)證不同處理后對腫瘤細(xì)胞活性和增殖能力的影響。9.基于表達(dá)譜芯片數(shù)據(jù),對下游的顯著性差異基因進(jìn)行GO分析從而了解ILF2的生物學(xué)功能,從Biological Process,Cellular Component和Molecular Function 三個方面進(jìn)行闡釋,可以得到更加全面的信息。利用KEGG數(shù)據(jù)庫對這些差異基因進(jìn)行Pathway分析,從而有利于我們判斷差異基因與哪些信號通路相關(guān),全面了解ILF2的分子機(jī)制。結(jié)果1.在27對和另外72對肝癌病例中,我們發(fā)現(xiàn),和正常癌旁組織相比,在肝癌組織中ILF2的RNA和蛋白質(zhì)表達(dá)量明顯升高。ILF2表達(dá)量與患者年齡、性別、TNM分期、腫瘤分級、血管侵襲和肝硬化沒有相關(guān)性,與腫瘤大小具有密切關(guān)聯(lián)性(p=0.043)。相對于正常肝永生化細(xì)胞L02細(xì)胞,其余5株細(xì)胞系的ILF2的表達(dá)升高。TCGA數(shù)據(jù)庫對ILF2表達(dá)與肝癌患者預(yù)后進(jìn)行分析,結(jié)果顯示ILF2表達(dá)量升高與病人低生存率明顯關(guān)聯(lián)(p=0.0135)。2.在0,2,4,8,24h不同時間段用蛋白酶體抑制劑MG132處理肝癌細(xì)胞系,運(yùn)用免疫印跡法檢測不同時間處理細(xì)胞中的ILF2的表達(dá)。MG132處理后ILF2的表達(dá)沒有明顯變化。3.在體外功能實(shí)驗(yàn)中,我們發(fā)現(xiàn)ILF2過表達(dá)之后,可以促進(jìn)肝癌細(xì)胞的克隆形成和增殖能力,而抑制ILF2的表達(dá)之后,肝癌細(xì)胞的克隆形成和增殖能力明顯下降。4.流式凋亡實(shí)驗(yàn)結(jié)果發(fā)現(xiàn),ILF2過表達(dá)之后,與對照組相比,肝癌細(xì)胞的凋亡比例減少,干擾ILF2后結(jié)果則相反。裸鼠皮下成瘤組織進(jìn)行TUNEL實(shí)驗(yàn),結(jié)果顯示與對照組相比,過表達(dá)ILF2組的凋亡細(xì)胞明顯減少。5.在裸鼠體內(nèi)的瘤體生成實(shí)驗(yàn)中,Lenti-ILF2組得到的瘤體在重量和體積上均明顯要高于對照組,免疫組化分析兩組腫瘤組織的Ki67存在表達(dá)差異,并且過表達(dá)組的Ki67明顯高表達(dá)。6.分子機(jī)制研究表明ILF2影響凋亡蛋白的表達(dá),如BOK,BAX,BCL-2和cIAPl。ILF2升高導(dǎo)致的BCL-2和cIAP1的升高而BOK和BAX表達(dá)降低。而干擾ILF2后,下游蛋白的結(jié)果則相反。而對同為凋亡蛋白BAK來說,無論ILF2過表達(dá)或者干擾,BAK的表達(dá)均無變化。裸鼠皮下成瘤組織進(jìn)行免疫組化實(shí)驗(yàn),結(jié)果顯示,與對照組相比,過表達(dá)ILF2組的BCL-2和cIAP1明顯升高。7.與si-NC對照組相比,敲低ILF2的表達(dá)確實(shí)能增強(qiáng)厄洛替尼對肝癌細(xì)胞的活性抑制,兩種處理同時進(jìn)行能夠發(fā)揮協(xié)同抑制作用。同樣的濃度梯度處理細(xì)胞,在體外進(jìn)行克隆實(shí)驗(yàn),和CCK8結(jié)果一致的是,si-ILF2聯(lián)合厄洛替尼處理明顯能提高厄洛替尼對肝癌細(xì)胞的生長或增殖抑制作用。8.GO分析中,在Biological Process,ILF2與NF-KB、JNK和G1/S等信號通路密切相關(guān);從Cellular Component發(fā)現(xiàn),ILF2參與細(xì)胞外泌體、mRNA加工以及核蛋白復(fù)合物的組成;而通過Molecular Function的信息可知,其參與離子通道、RNA和DNA的結(jié)合。Pathway分析結(jié)果顯示,脂肪酸代謝、糖代謝和AMPK信號通路與其密切相關(guān)。結(jié)論1.和肝癌鄰近正常組織與正常肝細(xì)胞相比,在肝癌組織和肝癌細(xì)胞中,ILF2的mRNA和蛋白水平明顯升高,肝癌患者中ILF2的高表達(dá)與腫瘤大小及低生存率密切相關(guān),而且ILF2的表達(dá)上調(diào)主要是通過轉(zhuǎn)錄水平的調(diào)節(jié)。2.ILF2增強(qiáng)肝癌細(xì)胞的活性和增殖能力、抑制腫瘤細(xì)胞的凋亡,并且促進(jìn)裸鼠瘤體的生長和增殖能力。3.在細(xì)胞水平上,BOK,BAX,BCL-2和cIAP1被證明由ILF2的調(diào)節(jié),并且在體內(nèi)成瘤腫瘤組織中證實(shí)ILF2過表達(dá)組出現(xiàn)BCL-2和cIAP1明顯升高。4:體外克隆形成實(shí)驗(yàn)和細(xì)胞活性實(shí)驗(yàn)的研究結(jié)果均說明,si-ILF2聯(lián)合不同濃度的EGFR抑制劑厄洛替尼在肝癌細(xì)胞的增殖能力和細(xì)胞活性抑制上有可能存在聯(lián)合或協(xié)同作用。5.表達(dá)譜芯片結(jié)果顯示,ILF2與mRNA加工、RNA/DNA相互結(jié)合、代謝過程以及AMPK信號通路等密切相關(guān)。
[Abstract]:Background in our country, Hepatocellular carcinoma (HCC) is a high incidence of malignant tumor. 90% of the patients have the history of hepatitis B. In China, about 110 thousand people died of liver cancer every year, and the initial symptoms of 45%. cancer patients in the death toll of all the world's liver cancer are not obvious. However, once the symptoms appear, the course of the disease is mostly in the late stage. If the malignancy is high, the prognosis is worse. Although many advances have been made in the past few decades in the treatment of liver cancer, radiotherapy, chemotherapy and liver transplantation, the long-term prognosis of the patients with liver cancer is still not ideal. The main reason for the poor prognosis is that the effective therapeutic targets for liver cancer have not been found and the molecular mechanism of the development of liver cancer. Therefore, the search for more sensitive and specific molecular markers of liver cancer and the mechanism of its role in the development of liver cancer will be the key to further improving the survival rate of HCC, and have a profound impact on the early diagnosis and early treatment of HCC. The purpose of this study is to explore the Interleukin enhancer binding f Actor 2 (ILF2) was used in the liver cancer tissues and para cancer tissues of the patients with hepatectomy and the expression level of the hepatoma cell lines in our hospital, and then the relationship between the pathological indexes related to the patients was explored, and the relationship with the prognosis of the patients with liver cancer was analyzed in the TCGA database. The proteasome inhibitor MG132 was in the 0,2,4,8,24h, respectively, in the 0,2,4,8,24h five. The high expression of ILF2 in liver cancer tissues and hepatoma cells is caused by the transcriptional level in different time periods. Using cytological experiments and in vivo tumorigenesis, the role of ILF2 in the development of hepatocellular carcinoma and the signaling pathway in the formation of liver cancer is deeply elucidated. By interfering with ILF2 The expression of O mu M, 5 mu M, 1O mu M, and 20 micron M of the human epidermal growth factor receptor (EGFR) inhibitor erlotinib were added to the hepatoma cells, and the synergistic effect of two treatments on the cell activity and proliferation ability was discussed. Based on the data of the expression chip, the downstream differential gene was analyzed by GO, and the KEGG database was used. The differential gene was analyzed by Pathway to carry out a deep analysis and Study on the downstream molecules or downstream pathways of the ILF2. Method 1. the mRNA of hepatoma liver resection and paracancerous tissue in 27 cases of HCC patients was extracted, and 72 other cancer patients and paraffin paraffin tissues were collected, and L02, Huh7, Bel-7402, MHCC-LM3, SMCC-7721 and SK-HEP- were extracted. The protein of mRNA and 8 in the 1 cell lines, using Western immunoblotting, real-time fluorescence quantitative PCR, immunohistochemistry and other techniques to verify the expression of interleukin enhanced binding factor 2 (ILF2) in HCC tumor tissue and all hepatoma cell lines.2. based on the immunohistochemical results of liver cancer tissue, ILF2 and liver cancer were explored. The relationship between human clinicopathological parameters.3. using TCGA database to analyze the relationship between ILF2 and the survival rate of patients with liver cancer.4. through the proteasome inhibitor MG132 treatment of liver cancer cell lines, explore whether the increase of ILF2 expression through the up regulation of transcriptional level causes.5. through the transfection of ILF2 small interference RNA and lentivirus stable transfection of ILF2. Interference or overexpression of ILF2, through cytological experiments, such as cell clone accumulation, apoptosis and Cell Counting Kit-8 (CCK-8), explore the role of ILF2 in the regulation of proliferation and apoptosis of hepatoma cells,.6. in order to explore the role and effect of ILF2 in the process of tumor formation in vivo, and we enter the body in nude mice. The tumor cell tumor formation experiment.7. uses Western blot technology to find the downstream molecules regulated by ILF2 to explore the molecular mechanism of ILF2 in the liver cancer,.8. by knocking low ILF2 expression and treating hepatoma cells with different concentrations of erlotinib, and using Cell CountingKit-8 (CCK-8) and clone formation experiments to verify the different treatment after treatment. The effect of.9. on the activity and proliferation of tumor cells is based on the expression spectrum chip data, and the GO analysis of the significant differentially differentially downstream genes is carried out to understand the biological function of ILF2, and the three aspects of Biological Process, Cellular Component and Molecular Function can be explained in order to get more comprehensive information. The use of KEGG database Pathway analysis of these differentially differentiable genes helps us to determine which signaling pathways are associated with the molecular mechanisms of ILF2. Results 1. in 27 pairs and 72 other cases of liver cancer, we found that the RNA and protein expression of ILF2 in the liver cancer tissues is significantly higher in.ILF2 expression than in normal para cancerous tissues. The volume was not associated with age, sex, TNM staging, tumor classification, vascular invasion and cirrhosis, and was closely related to the size of the tumor (p=0.043). Compared with normal liver immortalized L02 cells, the expression of ILF2 in the other 5 cell lines increased by.TCGA database to analyze the expression of ILF2 and the prognosis of liver cancer patients, and the results showed the ILF2 table. A significant association between the increase of amount and the low survival rate of the patient (p=0.0135).2. was used to treat the hepatocellular carcinoma cell lines with proteasome inhibitor MG132 at different time periods of 0,2,4,8,24h. The expression of ILF2 in the treated cells at different time was detected by Western blot, and the expression of ILF2 was not obviously changed after.MG132 treatment, and.3. in the function experiment in vitro, we found ILF2. After overexpression, it can promote the cloning and proliferation of hepatoma cells, and after inhibiting the expression of ILF2, the clone formation and proliferation ability of hepatoma cells decreased obviously by.4. flow apoptosis experiment. After ILF2 overexpression, the apoptosis ratio of hepatoma cells decreased compared with the control group, and the result of interference ILF2 was the opposite. Nude mice skin was the opposite. The results of TUNEL test in the lower tumor tissue showed that compared with the control group, the apoptotic cells overexpressed in the ILF2 group significantly reduced the tumor formation of.5. in the nude mice. The weight and volume of the Lenti-ILF2 group were significantly higher than those of the control group. The expression of Ki67 in the two groups of tumor tissues in the group of immuno histochemical analysis was different. The.6. molecular mechanism of the expression of Ki67 in the expression group indicates that ILF2 affects the expression of apoptotic proteins, such as BOK, BAX, BCL-2 and cIAPl.ILF2, which leads to the increase of BCL-2 and cIAP1, while BOK and BAX are reduced. The expression of BCL-2 and cIAP1 in the subcutaneous tissue of nude mice showed that compared with the control group, the expression of BCL-2 and cIAP1 in the overexpressed ILF2 group was significantly higher than that of the si-NC control group. The expression of low ILF2 could indeed enhance the inhibition of erlotinib on the activity of hepatoma cells, and the two treatments could play a synergistic inhibition at the same time. The same concentration gradient treatment cells were cloned in vitro, and the CCK8 results were consistent with that si-ILF2 combined with erlotinib significantly enhanced the.8.GO analysis of erlotinib's inhibitory effect on the growth or proliferation of hepatoma cells, which was closely related to Biological Process, ILF2 and NF-KB, JNK, and G1/S; from Cellular C. Omponent found that ILF2 participates in the extracellular secretory, mRNA processing and the composition of the nucleoprotein complex; and through the Molecular Function information, it is found that its involvement in ion channels, RNA and DNA binding.Pathway analysis results show that fatty acid metabolism, sugar metabolism and AMPK signaling pathway are closely related to it. Conclusion 1. and hepatocellular carcinoma are adjacent to normal tissues and positive cells. The mRNA and protein levels of ILF2 increased significantly in hepatoma and hepatoma cells. The high expression of ILF2 was closely related to the tumor size and low survival rate in HCC patients, and the up-regulated expression of ILF2 mainly enhanced the activity and proliferation of hepatoma cells through the regulation of.2.ILF2 at the transcriptional level and inhibited the tumor cells. Apoptosis, and promote the growth and proliferation of nude mice,.3. at the cell level, BOK, BAX, BCL-2 and cIAP1 have been proved to be regulated by ILF2, and in the tumor tissue of the body, it is confirmed that BCL-2 and cIAP1 in the ILF2 overexpression group are obviously elevated in the presence of.4: in vitro and in cell activity experiments. Different concentrations of EGFR inhibitor erlotinib may have joint or synergistic.5. expression profiles on the proliferation ability and cell activity inhibition of hepatoma cells. The results of ILF2 and mRNA processing, RNA/DNA binding, metabolic process and AMPK signaling pathway are closely related.
【學(xué)位授予單位】：浙江大學(xué)
【學(xué)位級別】：博士
【學(xué)位授予年份】：2017
【分類號】：R735.7
本文編號：2147812
[Abstract]:Background in our country, Hepatocellular carcinoma (HCC) is a high incidence of malignant tumor. 90% of the patients have the history of hepatitis B. In China, about 110 thousand people died of liver cancer every year, and the initial symptoms of 45%. cancer patients in the death toll of all the world's liver cancer are not obvious. However, once the symptoms appear, the course of the disease is mostly in the late stage. If the malignancy is high, the prognosis is worse. Although many advances have been made in the past few decades in the treatment of liver cancer, radiotherapy, chemotherapy and liver transplantation, the long-term prognosis of the patients with liver cancer is still not ideal. The main reason for the poor prognosis is that the effective therapeutic targets for liver cancer have not been found and the molecular mechanism of the development of liver cancer. Therefore, the search for more sensitive and specific molecular markers of liver cancer and the mechanism of its role in the development of liver cancer will be the key to further improving the survival rate of HCC, and have a profound impact on the early diagnosis and early treatment of HCC. The purpose of this study is to explore the Interleukin enhancer binding f Actor 2 (ILF2) was used in the liver cancer tissues and para cancer tissues of the patients with hepatectomy and the expression level of the hepatoma cell lines in our hospital, and then the relationship between the pathological indexes related to the patients was explored, and the relationship with the prognosis of the patients with liver cancer was analyzed in the TCGA database. The proteasome inhibitor MG132 was in the 0,2,4,8,24h, respectively, in the 0,2,4,8,24h five. The high expression of ILF2 in liver cancer tissues and hepatoma cells is caused by the transcriptional level in different time periods. Using cytological experiments and in vivo tumorigenesis, the role of ILF2 in the development of hepatocellular carcinoma and the signaling pathway in the formation of liver cancer is deeply elucidated. By interfering with ILF2 The expression of O mu M, 5 mu M, 1O mu M, and 20 micron M of the human epidermal growth factor receptor (EGFR) inhibitor erlotinib were added to the hepatoma cells, and the synergistic effect of two treatments on the cell activity and proliferation ability was discussed. Based on the data of the expression chip, the downstream differential gene was analyzed by GO, and the KEGG database was used. The differential gene was analyzed by Pathway to carry out a deep analysis and Study on the downstream molecules or downstream pathways of the ILF2. Method 1. the mRNA of hepatoma liver resection and paracancerous tissue in 27 cases of HCC patients was extracted, and 72 other cancer patients and paraffin paraffin tissues were collected, and L02, Huh7, Bel-7402, MHCC-LM3, SMCC-7721 and SK-HEP- were extracted. The protein of mRNA and 8 in the 1 cell lines, using Western immunoblotting, real-time fluorescence quantitative PCR, immunohistochemistry and other techniques to verify the expression of interleukin enhanced binding factor 2 (ILF2) in HCC tumor tissue and all hepatoma cell lines.2. based on the immunohistochemical results of liver cancer tissue, ILF2 and liver cancer were explored. The relationship between human clinicopathological parameters.3. using TCGA database to analyze the relationship between ILF2 and the survival rate of patients with liver cancer.4. through the proteasome inhibitor MG132 treatment of liver cancer cell lines, explore whether the increase of ILF2 expression through the up regulation of transcriptional level causes.5. through the transfection of ILF2 small interference RNA and lentivirus stable transfection of ILF2. Interference or overexpression of ILF2, through cytological experiments, such as cell clone accumulation, apoptosis and Cell Counting Kit-8 (CCK-8), explore the role of ILF2 in the regulation of proliferation and apoptosis of hepatoma cells,.6. in order to explore the role and effect of ILF2 in the process of tumor formation in vivo, and we enter the body in nude mice. The tumor cell tumor formation experiment.7. uses Western blot technology to find the downstream molecules regulated by ILF2 to explore the molecular mechanism of ILF2 in the liver cancer,.8. by knocking low ILF2 expression and treating hepatoma cells with different concentrations of erlotinib, and using Cell CountingKit-8 (CCK-8) and clone formation experiments to verify the different treatment after treatment. The effect of.9. on the activity and proliferation of tumor cells is based on the expression spectrum chip data, and the GO analysis of the significant differentially differentially downstream genes is carried out to understand the biological function of ILF2, and the three aspects of Biological Process, Cellular Component and Molecular Function can be explained in order to get more comprehensive information. The use of KEGG database Pathway analysis of these differentially differentiable genes helps us to determine which signaling pathways are associated with the molecular mechanisms of ILF2. Results 1. in 27 pairs and 72 other cases of liver cancer, we found that the RNA and protein expression of ILF2 in the liver cancer tissues is significantly higher in.ILF2 expression than in normal para cancerous tissues. The volume was not associated with age, sex, TNM staging, tumor classification, vascular invasion and cirrhosis, and was closely related to the size of the tumor (p=0.043). Compared with normal liver immortalized L02 cells, the expression of ILF2 in the other 5 cell lines increased by.TCGA database to analyze the expression of ILF2 and the prognosis of liver cancer patients, and the results showed the ILF2 table. A significant association between the increase of amount and the low survival rate of the patient (p=0.0135).2. was used to treat the hepatocellular carcinoma cell lines with proteasome inhibitor MG132 at different time periods of 0,2,4,8,24h. The expression of ILF2 in the treated cells at different time was detected by Western blot, and the expression of ILF2 was not obviously changed after.MG132 treatment, and.3. in the function experiment in vitro, we found ILF2. After overexpression, it can promote the cloning and proliferation of hepatoma cells, and after inhibiting the expression of ILF2, the clone formation and proliferation ability of hepatoma cells decreased obviously by.4. flow apoptosis experiment. After ILF2 overexpression, the apoptosis ratio of hepatoma cells decreased compared with the control group, and the result of interference ILF2 was the opposite. Nude mice skin was the opposite. The results of TUNEL test in the lower tumor tissue showed that compared with the control group, the apoptotic cells overexpressed in the ILF2 group significantly reduced the tumor formation of.5. in the nude mice. The weight and volume of the Lenti-ILF2 group were significantly higher than those of the control group. The expression of Ki67 in the two groups of tumor tissues in the group of immuno histochemical analysis was different. The.6. molecular mechanism of the expression of Ki67 in the expression group indicates that ILF2 affects the expression of apoptotic proteins, such as BOK, BAX, BCL-2 and cIAPl.ILF2, which leads to the increase of BCL-2 and cIAP1, while BOK and BAX are reduced. The expression of BCL-2 and cIAP1 in the subcutaneous tissue of nude mice showed that compared with the control group, the expression of BCL-2 and cIAP1 in the overexpressed ILF2 group was significantly higher than that of the si-NC control group. The expression of low ILF2 could indeed enhance the inhibition of erlotinib on the activity of hepatoma cells, and the two treatments could play a synergistic inhibition at the same time. The same concentration gradient treatment cells were cloned in vitro, and the CCK8 results were consistent with that si-ILF2 combined with erlotinib significantly enhanced the.8.GO analysis of erlotinib's inhibitory effect on the growth or proliferation of hepatoma cells, which was closely related to Biological Process, ILF2 and NF-KB, JNK, and G1/S; from Cellular C. Omponent found that ILF2 participates in the extracellular secretory, mRNA processing and the composition of the nucleoprotein complex; and through the Molecular Function information, it is found that its involvement in ion channels, RNA and DNA binding.Pathway analysis results show that fatty acid metabolism, sugar metabolism and AMPK signaling pathway are closely related to it. Conclusion 1. and hepatocellular carcinoma are adjacent to normal tissues and positive cells. The mRNA and protein levels of ILF2 increased significantly in hepatoma and hepatoma cells. The high expression of ILF2 was closely related to the tumor size and low survival rate in HCC patients, and the up-regulated expression of ILF2 mainly enhanced the activity and proliferation of hepatoma cells through the regulation of.2.ILF2 at the transcriptional level and inhibited the tumor cells. Apoptosis, and promote the growth and proliferation of nude mice,.3. at the cell level, BOK, BAX, BCL-2 and cIAP1 have been proved to be regulated by ILF2, and in the tumor tissue of the body, it is confirmed that BCL-2 and cIAP1 in the ILF2 overexpression group are obviously elevated in the presence of.4: in vitro and in cell activity experiments. Different concentrations of EGFR inhibitor erlotinib may have joint or synergistic.5. expression profiles on the proliferation ability and cell activity inhibition of hepatoma cells. The results of ILF2 and mRNA processing, RNA/DNA binding, metabolic process and AMPK signaling pathway are closely related.
【學(xué)位授予單位】：浙江大學(xué)
【學(xué)位級別】：博士
【學(xué)位授予年份】：2017
【分類號】：R735.7
【參考文獻(xiàn)】
相關(guān)期刊論文 前1條
1 Mitsuro Kanda;Hiroyuki Sugimoto;Yasuhiro Kodera;;Genetic and epigenetic aspects of initiation and progression of hepatocellular carcinoma[J];World Journal of Gastroenterology;2015年37期
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