乙型肝炎病毒preS1基因对肝癌干细胞的发生和发展的促进作用研究
发布时间：2018-08-19 17:14
【摘要】：肝细胞癌是一种常见的原发性恶性肿瘤。乙型肝炎病毒是造成肝细胞癌发生的一个主要原因,但是以往的研究都是从间接的方式去阐述其致癌的机理,其致癌方式并没有被清楚地说明。我们从文献得知癌症干细胞作为一种癌组织产生的源头,能直接促进癌症的发生,因此我们想从癌症干细胞方面研究乙型肝炎病毒介导的肝细胞癌形成原因。为了研究肝癌细胞与普通肝细胞之间的区别,我们检测肝癌细胞系HepG2与正常肝细胞系L02的干细胞相关因子(Klf4、c-Myc、Nanog、Oct4和Sox2)的mRNA表达水平以及肝癌干细胞表面标志物(CD90、CD117和CD133)的表达水平。我们发现HepG2细胞系的干细胞相关因子的mRNA表达水平和肝癌干细胞表面标志物的表达水平都比L02细胞系高。这个发现告诉了我们肝细胞向肝癌细胞转化的过程与细胞的去分化和癌干细胞表面标志物表达紧密相关。紧接着我们对肝癌细胞系HepG2以及其转入乙型肝炎病毒基因组的细胞系HepG2.2.15进行研究,我们发现在HepG2.2.15细胞系的干细胞相关因子mRNA表达水平上明显比HepG2细胞系高,癌干细胞表面标志物的表达水平也显著比HepG2高。随后我们在细胞生长实验上发现HepG2.2.15细胞系的增殖能力和迁移能力比HepG2细胞系更强。以上的实验结果证实HepG2.2.15细胞系在乙型肝炎病毒基因组的影响下,其癌干细胞化的水平比HepG2细胞系高。在以往的文献报道当中,我们发现乙型肝炎病毒导致肝细胞癌的主要因素可能是病毒生命周期中产生的HBx蛋白和preS/S蛋白部分,我们在相关文献中也发现HBx、preS1和preS2与肝癌的发生发展可能相关。因此我们设计了一些实验检测相关蛋白HBx、preS1和preS2对于肝细胞转化成癌干细胞的能力。我们在肝细胞系L02细胞和肝癌细胞系HepG2细胞中分别稳定表达了 HBx、preS1和preS2基因。在我们检测各个构建好的细胞系的癌干细胞相关指标之前,我们惊奇地发现稳定表达preS1的L02细胞系的生长情况和细胞形态发生了巨大的改变,随后我们深入检测preS1相关细胞系的癌干细胞水平。我们发现了 preS1能强烈地促进相应细胞系的癌干细胞化,使其干细胞相关因子mRNA表达水平显著提高,癌干细胞标志物的表达水平也部分提高了。随后通过一系列细胞生长实验确定了 preS1基因能促进肝细胞系L02的自我更新和增殖能力,揭示乙肝病毒的PreS1蛋白是导致肝癌干细胞发生发展的重要因素。
[Abstract]:Hepatocellular carcinoma is a common primary malignant tumor. Hepatitis B virus is one of the main causes of hepatocellular carcinoma, but previous studies have explained the mechanism of its carcinogenesis in an indirect way, and its carcinogenesis has not been clearly explained. We know from the literature that cancer stem cells as a source of cancer tissue can directly promote the occurrence of cancer, so we want to study the pathogenesis of Hepatitis B virus mediated hepatocellular carcinoma from the aspect of cancer stem cells. In order to study the difference between hepatoma cells and normal hepatocytes, we detected the mRNA expression level of stem cell related factors (Klf4C- Myct4 and Sox2) in HepG2 and L02, and the expression of CD90 CD117 and CD133 on the surface of hepatoma stem cells. We found that the mRNA expression level of stem cell related factors and the expression level of hepatoma stem cell surface marker in HepG2 cell line were higher than those in L02 cell line. This discovery tells us that the process of transforming hepatocytes into hepatoma cells is closely related to cell dedifferentiation and expression of tumor stem cell surface markers. Then we studied the hepatoma cell line HepG2 and the cell line HepG2.2.15 which was transferred to the genome of hepatitis B virus. We found that the mRNA expression level of stem cell related factors in HepG2.2.15 cell line was significantly higher than that in HepG2 cell line. The expression of tumor stem cell surface markers was also significantly higher than that of HepG2. Then we found that the proliferation and migration ability of HepG2.2.15 cell line was stronger than that of HepG2 cell line. The above results confirmed that the level of cancer stem cell transformation of HepG2.2.15 cell line was higher than that of HepG2 cell line under the influence of hepatitis B virus genome. In previous literature, we have found that the main factors of hepatitis B virus leading to hepatocellular carcinoma may be the HBx protein and preS/S protein part of the virus life cycle. We also found that HBX preS1 and preS2 may be associated with the occurrence and development of liver cancer. Therefore, we designed some experiments to detect the ability of HBxP preS1 and preS2 to transform hepatocytes into cancer stem cells. We stably expressed HBX preS1 and preS2 genes in L02 and HepG2 cells, respectively. We were surprised to find that the growth and morphology of the L02 cell line, which expressed preS1 stably, had changed dramatically before we examined the relevant markers of cancer stem cells in each constructed cell line. We then examined the level of cancer stem cells in preS1-associated cell lines. We found that preS1 could strongly promote the carcinomatosis of the corresponding cell lines and increase the mRNA expression level of stem cell related factors and the expression level of cancer stem cell markers. Then a series of cell growth experiments confirmed that preS1 gene can promote the self-renewal and proliferation of liver cell line L02. It is revealed that the PreS1 protein of hepatitis B virus is an important factor leading to the development of liver cancer stem cells.
【学位授予单位】：武汉大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R735.7
本文编号：2192304
[Abstract]:Hepatocellular carcinoma is a common primary malignant tumor. Hepatitis B virus is one of the main causes of hepatocellular carcinoma, but previous studies have explained the mechanism of its carcinogenesis in an indirect way, and its carcinogenesis has not been clearly explained. We know from the literature that cancer stem cells as a source of cancer tissue can directly promote the occurrence of cancer, so we want to study the pathogenesis of Hepatitis B virus mediated hepatocellular carcinoma from the aspect of cancer stem cells. In order to study the difference between hepatoma cells and normal hepatocytes, we detected the mRNA expression level of stem cell related factors (Klf4C- Myct4 and Sox2) in HepG2 and L02, and the expression of CD90 CD117 and CD133 on the surface of hepatoma stem cells. We found that the mRNA expression level of stem cell related factors and the expression level of hepatoma stem cell surface marker in HepG2 cell line were higher than those in L02 cell line. This discovery tells us that the process of transforming hepatocytes into hepatoma cells is closely related to cell dedifferentiation and expression of tumor stem cell surface markers. Then we studied the hepatoma cell line HepG2 and the cell line HepG2.2.15 which was transferred to the genome of hepatitis B virus. We found that the mRNA expression level of stem cell related factors in HepG2.2.15 cell line was significantly higher than that in HepG2 cell line. The expression of tumor stem cell surface markers was also significantly higher than that of HepG2. Then we found that the proliferation and migration ability of HepG2.2.15 cell line was stronger than that of HepG2 cell line. The above results confirmed that the level of cancer stem cell transformation of HepG2.2.15 cell line was higher than that of HepG2 cell line under the influence of hepatitis B virus genome. In previous literature, we have found that the main factors of hepatitis B virus leading to hepatocellular carcinoma may be the HBx protein and preS/S protein part of the virus life cycle. We also found that HBX preS1 and preS2 may be associated with the occurrence and development of liver cancer. Therefore, we designed some experiments to detect the ability of HBxP preS1 and preS2 to transform hepatocytes into cancer stem cells. We stably expressed HBX preS1 and preS2 genes in L02 and HepG2 cells, respectively. We were surprised to find that the growth and morphology of the L02 cell line, which expressed preS1 stably, had changed dramatically before we examined the relevant markers of cancer stem cells in each constructed cell line. We then examined the level of cancer stem cells in preS1-associated cell lines. We found that preS1 could strongly promote the carcinomatosis of the corresponding cell lines and increase the mRNA expression level of stem cell related factors and the expression level of cancer stem cell markers. Then a series of cell growth experiments confirmed that preS1 gene can promote the self-renewal and proliferation of liver cell line L02. It is revealed that the PreS1 protein of hepatitis B virus is an important factor leading to the development of liver cancer stem cells.
【学位授予单位】：武汉大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R735.7
【参考文献】
相关期刊论文 前1条
1 Mustafa Sunbul;;Hepatitis B virus genotypes:Global distribution and clinical importance[J];World Journal of Gastroenterology;2014年18期
，本文编号：2192304
本文链接：https://www.wllwen.com/kejilunwen/jiyingongcheng/2192304.html
最近更新
教材专著
