HBV HBx 蛋白调控肝癌细胞增殖的机制研究

目的 探讨 HBV HBx 蛋白调控肝癌细胞增殖的潜在机制。 方法 过表达 HBx 后, 检测肝癌细胞HepG2 的增殖水平。 在过表达 Flag-HBx 的肝癌细胞 HepG2 中免疫共沉淀 HBx 后进行蛋白质质谱鉴定,Score 阈值设定为大于 50, Coverage 阈值设定为大于 10。 过表达 HBx 的肝癌细胞 HepG2 中, 使用 siRNA 敲低质谱鉴定的相应基因, 检测 HepG2 的增殖水平。 结果 过表达 Flag-HBx 后, 肝癌细胞 HepG2 的增殖水平上升 (P< 0. 05)。 与对照组比较, 敲低 MCUR1 后肝癌细胞 HepG2 的增殖水平下降 (P< 0. 05)。 与对照组比较, 同时过表达 MCUR1 和 HBx 能够促进肝癌细胞 HepG2 的增殖 (P< 0. 05), 而仅过表达 MCUR1 对肝癌细胞 HepG2 的增殖无显著影响。 与过表达 HBx 组比较, 过表达 HBx 的同时敲低 MCUR1 时肝癌细胞HepG2 的增殖水平下降 (P< 0. 05)。 与对照组比较, 过表达 HBx 后肝癌细胞 HepG2 的 ROS 水平上升 (P<0. 05); 与对照组比较, 过表达 HBx 的同时敲低 MCUR1 时肝癌细胞 HepG2 的 ROS 水平无显著变化。 与对照组或过表达 HBx 组比较, 同时过表达 MCUR1 和 HBx 能够增加肝癌细胞 HepG2 的 ROS 水平 (P< 0. 05)。与过表达 HBx 组比较, 过表达 HBx 的同时敲低 MCUR1 时肝癌细胞 HepG2 的增殖水平下降 (P< 0. 05), 但这种下降能被 H2O2恢复。 与对照组比较, 过表达 HBx 后肝癌细胞 HepG2 中转录因子 Nrf2 在细胞核中的定位增加 (P< 0. 05), 而过表达 HBx 的同时敲低 MCRU1 后 Nrf2 在细胞核中的定位无显著变化。 与对照组比较, 过表达 HBx 后肝癌细胞 HepG2 中转录因子 Notch 的激活形式 NICD1 在细胞核中的定位增加 ( P<0. 05), 而过表达 HBx 的同时 Nrf2 抑制剂 ML385 处理后 NICD1 在细胞核中的定位无显著变化。 与过表达HBx 组比较, 过表达 HBx 的同时 ML385 处理后肝癌细胞 HepG2 的增殖水平下降 (P< 0. 05)。 与过表达HBx 组比较, 过表达 HBx 的同时 Notch 抑制剂 IMR-1 处理后肝癌细胞 HepG2 的增殖水平下降 (P< 0. 05)。 结论 HBx 促进 HepG2 细胞增殖与 MCUR1 / ROS / Nrf2 / Notch 轴有关。     

Notch信号通路与肝癌的研究进展

Notch信号通路是一种高度保守的信号通路,在调节细胞分化、增殖和凋亡等一系列生理病理过程中都起着关键性的作用。Notch信号在肝癌中频繁发生异常表达及激活突变,其通过多种机制促进肝癌的发生发展,且与肝癌的侵袭、转移和预后密切相关。因此,Notch信号可作为肝癌治疗的一个靶标,为肝癌的治疗提供新的方向和机遇。     

Notch通路介导的microRNAs对肝癌干细胞的调节作用

肝癌干细胞(hepatic cancer stem cells,HSCs)是存在于肝细胞癌中的一类具有干细胞特性的细胞,与肝癌的形成、 生长、 转移、 药物耐受和复发有密切关系.microRNAs(miRNAs)是一种长约19~22 nt的短链非编码RNA,在细胞众多的生化活动中起着重要的调节作用,通过作用于细胞中关键信号通路的重要节点分子调控肿瘤的发生发展.而Notch信号通路是调控肿瘤干细胞自我更新、 增殖和分化过程的最重要的信号通路之一.miRNAs在Notch信号通路中的作用如何?miRNAs通过靶向作用于Notch信号通路进而发挥调控肝癌干细胞的自我更新、 分化及致癌性作用的机制是什么?这均是本文重点阐述的问题.     

URG4在肝癌组织中的表达及其与HBx的关系

目的:检测URG4在肝癌组织中的表达及意义。探讨URG4与HBx在肝癌组织表达之间的关系。方法:免疫组化方法检测URG4和HBx在正常组织、肝癌组织及癌旁组织中的表达分布,并分析其表达的意义及表达之间的相关性。结果:URG4在肝癌组织及肝癌细胞系中的阳性表达率分别为48%和54%,而在正常组织中仅22.2%弱表达,其与HBx表达的相关系数分别为0.38和0.45(P0.05)。结论:URG4在肝癌组织及癌旁组织中的表达高于正常组织,且与HBx密切相关。     

施旺细胞发育及其调控的信号通路

施旺细胞是由神经嵴细胞分化而来的周围神经系统中特有的成髓鞘细胞,它的发育过程分为施旺细胞前体、不成熟施旺细胞、成熟施旺细胞三个阶段。此过程受到很多细胞因子的影响,如轴突的神经调节蛋白、自分泌因子等。这些因子共同调节施旺细胞的分化、增殖、成熟以及迁移。本文阐述了施旺细胞发育、细胞因子的作用及其作用的受体和信号通路,并展望施旺细胞未来的研究方向和临床应用的潜在价值。     

肝癌新型预后分子ERα与乙型肝炎病毒X蛋白相互作用研究进展

HBV相关原发性肝癌发病率存在显著性别差异，雌激素在其中发挥重要作用。HBx直接参与肝癌的发生，其与雌激素受体间的相互作用可能是导致肝癌发生的一条重要信号通路。     

与过敏性哮喘发生有关的细胞信号转导通路

过敏性哮喘,是一类包括气道平滑肌异常,气道炎症以及气道可逆性狭窄改变的慢性炎症性疾病,以血清中IgE水平升高、气道周围嗜酸性粒细胞(EOS)浸润、支气管黏膜损害以及气道高反应性等为特征.     

JAK/STAT信号通路与肝细胞性肝癌的肿瘤进展和预后的相关性研究

目的:探讨JAK/STAT信号通路在肝细胞性肝癌组织中的表达及其与预后的关系。方法:选取196例肝细胞性肝癌组织标本,以正常肝脏组织标本20例作为对照,应用SABC免疫组化方法检测JAK-1蛋白和STAT-3蛋白的表达,分析二者与肝细胞性肝癌的病理分级、临床分期及预后的关系。结果:JAK-1蛋白和STAT-3蛋白在肝细胞性肝癌组织中的阳性表达率均高于正常肝脏组织(P=0.02,0.01);肝细胞性肝癌中JAK-1蛋白和STAT-3蛋白的表达与患者性别、年龄、肿瘤大小、有无肝硬化均无显著相关;包膜不完整(P=0.01,0.008)、出现门静脉癌栓(P=0.02)、分化未成熟(P=0.01,0.009)、临床III～IV期(P=0.02,0.008)的肝细胞性肝癌组织中JAK-1蛋白和STAT-3蛋白的表达明显高于有完整包膜、未出现门静脉癌栓、分化较成熟及临床I～II期的组织。Cox比例风险回归模型分析表明JAK-1蛋白和STAT-3蛋白均为影响肝细胞性肝癌预后的危险因素。结论:JAK/STAT信号通路的过度活化在肝细胞性肝癌的发生和发展过程中起重要作用,可以作为评价肝细胞性肝癌恶性程度及预后的重要生物学指标。     

HBx抗原对DNA损伤反应的影响与肝细胞癌发生

肝细胞癌(HCC)是世界范围内发病率非常高的恶性肿瘤之一。HBV和HCV感染是肝细胞癌发生的主要致病因素。乙型肝炎病毒x蛋白(hepatitis B virus x protein,HBx)是HBV基因组x区编码的一种多功能蛋白,在肝细胞癌发生中起重要作用。HBV抗原表达可引起氧化应激(oxidative stress,OS)致DNA损伤(DNA damage)。研究发现DNA损伤反应的异常是肿瘤发生早期的重要机制。本文重点就HBx对DNA损伤反应的影响及这一影响作用与肝细胞癌发生关系的研究进展做一综述。     

巨噬细胞极化信号通路及其调控机制研究进展

巨噬细胞是一类具有异质性和可塑性的免疫细胞,由单核细胞分化而来,在不同环境下可极化为不同的表型,分化中的巨噬细胞可位于极端分化的两型中的任意阶段。巨噬细胞在此极化过程中涉及较多的信号通路和转录调控,故本文将介绍具体巨噬细胞极化的通路及调控因子。     

乙型肝炎病毒变异与肝细胞癌发生关系

乙型肝炎病毒(hepatitis B virus,HBV)基因组复制时,以病毒前基因组RNA作为模板合成子代病毒DNA,催化该过程的逆转录酶缺乏校对功能,所以HBV易出现变异.近年来,各国学者通过比较肝细胞癌(hepatocellular carcinoma,HCC)患者和非HCC患者的HBV基因序列,发现HBV基本核...     

Hepatitis B Virus X Protein: The X Factor in Chronic Hepatitis B Virus Disease Progression

Introduction: Hepatitis B virus (HBV) is the most common aetiological factor causing hepatocellular carcinoma (HCC). HBx gene plays an enigmatic role in HBV-related HCC. In this study we have analysed amino acid substitutions in HBx from HBV-infected individuals of different clinical stages. Materials and Methods: HBV-infected individuals (n = 93) were recruited in the study. DNA was extracted from plasma, amplified, and DNA sequencing was performed using specific primers targeting HBx gene (540 bp). Results: Among the study participants, 57% had chronic HBV infection, 30% had chronic liver disease (CLD) and 13% had HBV related HCC. Genotypes such as D1, D2, D3, A1, C2 and B2 were identified of which genotype D2 was predominant (78%). HBxC-terminal deletion was observed in four hepatitis B e antigen (HBeAg) negative participants with CLD. The frequency of aminoacid substitution in proapoptotic domain was higher in HBeAg negative participants including I127V (34%), K130M (34%), V131I (40%). The frequency of double mutation (K130M+V131I) and triple mutation (I127V+K130M+V131I) were found to be higher (32% and 36%) in HBeAg negative participants. Also, we identified L5M substitution (4.3%) in HBeAg positive participants with advanced liver disease. Conclusion: In HBx gene, aminoacid substitutions at positions 127, 130, 131 are associated with poor expression of HBeAg. We suggest screening for HBx aminoacid substitutions especially in patients with HBeAg negative chronic HBV infection to predict the clinical outcome and enable early treatment to prevent disease progression.     

Stimulation of Cellular Proliferation by Hepatitis B Virus X Protein

Chronic infection with the hepatitis B virus (HBV) is a known risk factor in the development of human hepatocellular carcinoma (HCC). The HBV-encoded X protein, HBx, has been investigated for properties that may explain its cancer cofactor role in transgenic mouse lines. We discuss here recent data showing that HBx is able to induce hepatocellular proliferation in vitro and in vivo. This property of HBx is predicted to sensitize hepatocytes to other HCC cofactors, including exposure to carcinogens and to other hepatitis viruses. Cellular proliferation is intimately linked to the mechanism(s) by which most tumor-associated viruses transform virus-infected cells. The HBx alteration of the cell cycle provides an additional mechanism by which chronic HBV infection may contribute to HCC.     

Evaluation of Impact of Serial Hepatitis B Virus DNA Levels on Development of Hepatocellular Carcinoma

We aimed to investigate the impact of hepatitis B virus (HBV) DNA on the development of hepatocellular carcinoma (HCC). We conducted a case/control study based on 506 chronic HBV patients followed up since 1997. Forty-one patients developed HCC, and each of them was age and gender matched with two simultaneously recruited controls without HCC. HBV DNA was measured at the initial visit, at yearly intervals, and at the last visit. Patient age at the time of HCC development was 55 ± 9 years. Forty-nine (40%) patients experienced antiviral treatment. The median time from diagnosis to the development of HCC was 17 months, and the control patients were followed for 92 months. At the trough level (defined as lowest level among all studied visits), more (27 patients; 66%) HCC patients had HBV DNA levels of >10,000 copies/ml than the controls (17 patients; 21%). The area under the receiver operating characteristic curve of the trough log HBV DNA level for HCC was 0.79 (95% confidence interval [CI], 0.69 to 0.89). Trough log HBV DNA (odds ratio, 11.4; 95% CI, 3.6 to 37.6; P < 0.0001) and liver cirrhosis (odds ratio, 11.4; 95% CI, 3.6 to 36.2; P < 0.0001) levels were independently associated with HCC after an adjustment for age, gender, antiviral treatment, and HBV genotype. The difference in the trough HBV DNA level was more obvious among untreated patients (5.7 ± 1.4 log copies/ml in HCC patients versus 3.2 ± 1.3 log copies/ml in control patients; P < 0.0001) than among those who had received antiviral treatment (3.0 ± 1.4 log copies/ml in HCC patients versus 2.5 ± 0.9 log copies/ml in control patients; P = 0.38). A high trough HBV DNA level was associated with a higher risk of HCC. Whether antiviral treatment could prevent HCC was uncertain.Chronic hepatitis B virus (HBV) infection is the commonest cause of hepatocellular carcinoma (HCC) in Southeast Asia (8). In addition to the host factors, including an older age and male gender, viral factors such as being positive for HBV e antigen (HBeAg) and having high HBV DNA levels, genotype C HBV (particularly subgenotype Ce), and basal core promoter mutations have been suggested to associate with a higher risk of HCC (7, 9, 10, 11, 14, 28). In several cohort studies in Taiwan, China, and Hong Kong, a single high HBV DNA level higher than 4 log to 4.5 log copies/ml was associated with an increased HCC risk in the subsequent 10 years (7, 9, 10, 11). In addition, patients who had persistently high HBV DNA levels at the last follow-up visit also had an increased risk of developing HCC (10). In all previous studies, there was no information on the HBV DNA levels during the years between the first and last follow-up visits. As HBV DNA levels tend to fluctuate, particularly in HBeAg-negative patients, an assessment at a single time point cannot accurately reflect the severity of liver disease (6, 12, 13).In recently published management guidelines, antiviral treatment is recommended in patients with liver cirrhosis when the HBV DNA level is higher than 4 log copies/ml (15, 21, 23). This recommendation was based primarily on a multicentered, placebo-controlled trial among Asian chronic HBV patients who suffered from severe liver fibrosis (20). In this study, lamivudine treatment was demonstrated to reduce the risk of liver-related complications in 3 years. However, the benefit of prevention of HCC development among lamivudine-treated patients became dubious after excluding patients who developed HCC within the first year. In an Italian series, HBV DNA suppression by lamivudine and adefovir could not prevent the development of HCC among patients with decompensated liver cirrhosis (18).In this study, we aimed to investigate the impact of changes in HBV DNA on the development of HCC, either in the natural course of disease or as a result of antiviral treatment. To address this question, we studied the HBV DNA levels at different time intervals of follow-up among a cohort of chronic HBV patients before the development of HCC compared to those of simultaneously recruited controls.     

Detection of Hepatitis B Virus Surface Antigen Mutants in Paraffin-Embedded Hepatocellular Carcinoma Tissues

The antigenic a determinant of the human hepatitis B virus surface antigen (HBsAg) is highly conserved and involved in inducing neutralizing antibody. Mutations on this determinant have been associated with the hepatitis B virus (HBV) escape from vaccination but are also found in chronic HBV carriers, but their involvement in liver diseases including hepatocellular carcinoma (HCC) is unclear. To investigate the possible liver disease-associated role of HBsAg mutants, their incidence was analyzed in 11 paraffin embedded HCC tissues using an improved DNA extraction method. Mutations on the a determinant (Thr126Ser, Gly145Arg, a double mutant Thr126Ser/Gln129Asn, Met133Leu and Thr140Ile) were identified in 5 samples while the wild type sequence was found in 2 others. Future characterization of these HCC-associated HBsAg mutants should provide new insights on their role in the pathogenesis of HCC.     

Association of Polymorphism in MicroRNA 604 with Susceptibility to Persistent Hepatitis B Virus Infection and Development of Hepatocellular Carcinoma

MicroRNA polymorphisms may be associated with carcinogenesis or immunopathogenesis of infection. We evaluated whether the mircoRNA-604 (miR-604) polymorphism can affect the persistence of hepatitis B virus (HBV) infection, and the development to hepatocellular carcinoma (HCC) in patients with chronic HBV infection. A total of 1,439 subjects, who have either past or present HBV infection, were enrolled and divided into four groups (spontaneous recovery, chronic HBV carrier without cirrhosis, liver cirrhosis and HCC). We genotyped the precursor miR-604 genome region polymorphism. The CC genotype of miR-604 rs2368392 was most frequently observed and T allele frequency was 0.326 in all study subjects. The HBV persistence after infection was higher in those subjects with miR-604 T allele (P=0.05 in a co-dominant and dominant model), which implied that the patients with miR-604 T allele may have a higher risk for HBV chronicity. In contrast, there was a higher rate of the miR-604 T allele in the chronic carrier without HCC patients, compared to those of the HCC patients (P=0.03 in a co-dominant model, P=0.02 in a recessive model). The T allele at miR-604 rs2368392 may be a risk allele for the chronicity of HBV infection, but may be a protective allele for the progression to HCC in chronic HBV carriers.

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