肝癌患者C型乙型肝炎病毒前S基因突变研究

目的探讨肝癌患者C型乙型肝炎病毒(HBV)前S(pre-S)基因的突变情况。方法自肝癌患者血清中提取HBV基因组DNA,扩增其pre-S基因片段并进行序列分析,用MEGA3.1和C lustalx1.8作为所测基因进行分型,并对比pre-S基因序列。结果所测样品中,C型占56.0%,B型占20.0%,B/C混合感染型占24.0%。肝癌患者C型HBV前S区普遍存在基因突变。肝癌患者C型HBV pre-S1区域以个体化的点突变为主,而pre-S2区域则存在大量的缺失突变。结论 Pre-S基因是肝癌患者C型HBV的高变区。     

肝细胞癌患者B和C基因型乙型肝炎病毒X蛋白的变异特点

背景与目的：X蛋白是乙型肝炎病毒(hepatitis B virus,HBV)最重要的致病因子之一,它在HBV相关性肝细胞癌(hepatocellular carcinoma,HCC)的发生发展中起着重要作用。目前已知B、C基因型HBV相关性HCC的临床及病理表现不尽相同,但目前尚不清楚这种差别是否与B、C基因型HBV X基因之间的差别有关。因此本实验拟研究B、C基因型间HBV X蛋白氨基酸差异及其在HCC患者中的变异及特点,初步探讨其与HCC发生、发展的关系。方法：PCR扩增22份乙型肝炎病毒表面抗原(HBsAg)阳性HCC患者血清HBV X基因,克隆、测序并以Vector NTI6．0软件分析其基因型。以DNAMAN软件对标准HBV及HCC来源的HBV X蛋白进行氨基酸同源分析。结果：检测的22个HBV X基因片段均属于B或C基因型。B、C基因型HBV X蛋白之间存在14个氨基酸的差异,HCC患者来源的B、C型HBV X蛋白存在4个氨基酸的共有变异,C型HBV X蛋白尚有6个型特异性变异。这些差异或变异氨基酸均位于X蛋白B、T细胞表位或反式激活区及调节区内。结论：B、C基因型HBV X蛋白之间存在氨基酸差异,且在HCC中发生基因型特异性变异,这些差异或变异氨基酸可能导致X蛋白免疫学功能及反式激活功能的不同。     

乙型肝炎病毒的致癌机理

大约90％的肝癌与乙型肝炎或丙型肝炎病毒感染有关,这些病毒的感染引起急性或慢性肝炎,慢性肝炎中的部分患者逐渐发展为肝细胞癌。从免疫学角度来看,血液中S抗原(表面抗原,HBsAg)为阳性的持续携带者比非HBsAg携带者其肝癌发生率高100倍。从流行病统计学角度,乙型肝炎感染的地理分布与肝癌(HCC)的发生率也非常一致。 肝癌通常在患慢性肝炎20～50年后才会发病,现在,人们普遍认为肝癌的发生过程是由于存在于感染细胞内的乙型肝炎病毒基因诱发细胞癌基因的活化与抑癌基因的失活造成的。本文拟就存在于感染细胞内HBV基因与c-myc、p53等细胞基因的变异作一论述。 慢性肝炎及肝癌组织中几乎都能发现HBV DNA的多克隆整合,一般位于HBV DR_1或DR_2的近旁。即     

鸭肝癌实验模型研究——鸭乙型肝炎病毒对鸭胚的感染

人体肝癌的发生,虽然在流行病学方面提示肝癌与乙型肝炎病毒(HBV)和黄曲霉毒素B_1(AFB_1)等因素有关,但迄今为止,尚无证明HBV和AFB_1引发人体肝癌的直接证据。其中重要原因是缺少直接研究人体肝癌癌变的模型和手段。鸭乙型肝炎病毒(DHBV)与人乙型肝炎病毒(HBV)、土拨鼠乙型肝炎病毒(WHV)以及地松鼠乙型肝炎病(GSHV)同属嗜肝DNA病毒科(Hepadra-     

反向杂交检测肝癌相关乙型肝炎病毒前C区突变方法的建立和应用

背景与目的：日益增多的研究表明,乙型肝炎病毒(hepatitis B virus, HBV)DNA前C区G1896A和G1899A突变是肝癌发生的危险因素。本研究旨在建立简单、快速、灵敏和准确的检测HBV前C区突变的反向杂交方法,并应用于检测江苏省启东地区HBV DNA前C区突变与肝癌发生的关系。方法：设计并合成HBV DNA前C区1896和1899位点的特异性探针,通过优化条件建立特异、敏感的杂交体系,并与直接测序检测结果进行比较。将该方法应用于检测启东100例肝癌和100例慢性HBV携带者(对照组),分析HBV DNA前C区突变与肝癌的关系。结果：反向杂交对血清样本的最低检测下限为HBV DNA 103 copy/mL,检测混合感染时比直接测序更占优势,混合株中10％以上的突变株均可被检测。启东地区HBV DNA前C区G1899A突变与肝癌高发具有相关性(P=0.000,OR=4.846, 95%CI：2.240～10.485),而G1896A突变未见其相关性。结论：反向杂交检测HBV DNA前C区突变方便、快速、准确,可有效监控肝癌的发生,适合临床大规模推广应用。     

启东肝癌高发区乙肝病毒流行株全基因分析

目的：通过对江苏启东地区乙型肝炎病毒（HBV）全基因序列的分析，探讨该地区肝癌高发的分子病毒学病因。方法：以蛋白酶K消化后，酚／氯仿抽提7例肝炎和7例肝癌患者血清中DNA。应用聚合酶链反应（PCR），扩增血清中HBV基因全长，克隆至T载体后行全自动测序。以PHYLIP软件构建的系统进化树判断HBV的基因型；以Clastal W软件对序列进行突变分析。结果：14例标本中有12例为C基因型，2例为B基因型。肝癌和肝炎组中HBV基因型类别分布无差异。有5例HBV发生了PreS2的缺失突变，其中肝癌标本4例（57．1％），肝炎标本仅1例（14．3％）。HBV基因组中常见的点突变为PreS1区的nt．3116及核心启动子区的nt．1762／1764，发生率高达78．6％（11／14）。与肝炎组相比，肝癌中点突变发生率显著增高的位点为前C区nt．1899G→A的突变（P=0．01）及核心启动子区nt．1653C→T的突变（P=0．05）。结论：启东地区HBV以C基因型为主；启东HBV基因组中可能存在着与肝癌相关的点突变和缺失突变。     

乙型肝炎病毒前S2区缺失突变与肝癌相关性研究

目的研究肝细胞肝癌（hepatocarcinoma,HCC）患者乙型肝炎病毒（Hepatitis B virus,HBV）前S2区缺失突变情况。方法应用PCR法扩增HBV前S区基因,应用DNAMAN软件对所测基因进行分析。结果 23.08%（6/26）的肝癌患者发现存在preS2区的缺失突变,缺失位点为nt 3223～nt 3268,缺失长度为30～45 bp。肝癌患者缺失突变发生率显著高于HBV无症状携带者。结论 HBV preS2区的缺失突变可能与肝癌的发生密切相关,该突变的发生机制仍有待进一步研究。     

抑癌基因p16与乙肝病毒相关性肝癌研究进展

p16蛋白是细胞周期素依赖性蛋白激酶CDK4特异性的抑制子，通过抑制CDK4的活性，使细胞分裂停滞于G1-S期而阻止细胞的异常增殖，因而p16基因为重要的抑癌基因。乙型肝炎病毒(HBV)感染后，其DNA可整合入宿主肝细胞基因组中，并引起抑癌基因的改变，HBV与p16的改变有相关性。p16基因的改变在HBV相关性肝癌的发病机制中占重要地位。     

乙型肝炎病毒复制周期

慢性乙型肝炎病毒(HBV)感染是致癌性的。已对HBV基因组与肿瘤DNA之间关系进行过许多研究,但人类和动物慢性乙肝转变成肝细胞癌(HCC)的机理尚未清楚。持续感染是一个重要危险因素,可以相信如能治疗慢性感染就可防止发生肝癌,这促使许多人研究HBV在感染的肝细胞内复制和持续产生病毒的机制。     

广西肝癌高发区原发性肝细胞癌 HBV X 基因的整合及影响因素分析

目的 了解广西肝癌高发区乙型肝炎病毒（hepatitis B virus,HBV）X区基因在肝细胞癌（hepatocellular carcinoma,HCC）染色体中的整合及影响因素。方法 以30例与HBV相关的原发性肝细胞癌患者为研究对象。提取HCC组织及癌旁组织标本DNA作为模板,以HBV X基因上游序列和人类基因组Alu重复序列为引物,应用重复序列-多聚酶链反应（Alu-PCR）扩增整合的HBV X片段及两侧的人类基因组DNA片段。扩增产物进行测序,计算目的片段整合率并分析相关的影响因素。结果 18例HCC组织检测到HBV X基因的整合片段,整合率为60.00%（18/30）;26例癌旁组织检测到HBV X基因的整合片段,整合率为86.67%（26/30）。癌旁组织HBV病毒整合率高于HCC组织,差异有统计学意义（χ²=5.445,P=0.020）。不同性别、年龄、HBeAg、HBV DNA、ALT、AST的HCC癌组织及其癌旁组织HBV X基因整合率比较差异均无统计学意义（P＞0.05）。结论 广西肝癌高发区癌旁组织比HCC组织HBV整合率高,说明HBV整合发生在感染早期。HBV X基因整合与HCC患者性别、年龄、HBeAg、HBV DNA、ALT、AST无明显关系。     

Replication efficiency and sequence analysis of full-length hepatitis B virus isolates from hepatocellular carcinoma tissues

Prolonged replication of hepatitis B virus (HBV) in liver tissues of hepatitis B patients has been considered as an important risk factor for the development of malignancy. Few studies on full-length HBV sequencing in association with the replication efficiency of isolates from HCC tissues have been reported. To study the structural and functional genomics of HBV isolates from Chinese hepatocellular carcinoma (HCC) patients, full-length HBV genomes were amplified from 6 HBV-marker positive HCC tissues and used to transfect HepG2 cells. Five of 6 isolates showed high replicative efficiency. All isolates were of genotype C and "hot-spots" mutations were detected in the B cell and T helper (Th) cell epitopes of the envelope and the core region. In addition, the X region of 2 isolates contained a stop-codon mutation that was predicted to result in a truncated X protein. High replicative HBV immune escape mutants that persist in infected hepatocytes could be 1 of the important factors to initiate pathological processes for the development of HCC in Chinese patients.     

Analysis of the complete hepatitis B virus genome in patients with genotype C chronic hepatitis and hepatocellular carcinoma

Hepatitis B virus (HBV) genotype C and the basic core promoter (BCP) mutations were reported to be associated with the development of hepatocellular carcinoma (HCC). In this study the full sequences of HBV genomes were analyzed in order to find the other predictors of HCC development. We determined the full sequences of HBV genomes in 24 genotype C carriers who developed HCC (HCC group) at the beginning of follow-up and at the time of HCC diagnosis, and 20 patients who did not develop HCC (non-HCC group) served as a control. The number of nucleotide and amino acid substitutions in most regions was higher in the HCC group than in the non-HCC group, and the following substitutions and deletions were found more frequently in the HCC group than in the non-HCC group: G1317A and T1341C/A/G in the X promoter region were detected in 13 and six of the HCC cases, four and none of the non-HCC cases, respectively; and pre-S2 deletion was detected in eight HCC and none of the non-HCC cases. Compared with the wild type X promoter, the mutant type X promoters, M1 (G1317A), M2 (T1341C), and M4 (T1341G) showed increases in activity of 2.3, 3.8, and 1.4 times, respectively, in HepG2 cells. Substitutions and deletion of nucleotides of the HBV genome, especially the pre-S2 deletion and G1317A and T1341C/A/G mutations may be useful markers for predicting the development of HCC. ( Cancer Sci 2007; 98: 1921–1929)     

Occult hepatitis B virus infection is associated with the development of hepatocellular carcinoma in chronic hepatitis C patients

BACKGROUND: Occult hepatitis B virus (HBV) infection frequently occurs in patients with HBV surface antigen (HBsAg)-negative chronic liver disease, and much evidence suggests that it is a risk factor for hepatocellular carcinoma (HCC) development. However, to the authors' knowledge, no follow-up study has been performed to date evaluating HCC occurrence over time in chronic hepatitis patients with or without occult HBV infection. METHODS: A cohort of the 380 HBsAg-negative chronic hepatitis patients attending the study institution between 1991-2000 were evaluated and tested for occult HBV DNA by analysis of liver biopsy specimens. RESULTS: There were 135 patients (35.5%) with occult HBV and 245 patients (64.5%) without occult HBV. Cirrhosis was significantly associated with occult HBV infection (P = 0.01). One hundred thirty-four of these patients were followed for a minimum of 50 months (median, 82.8 +/- 32.6 mos). Fifty-three patients (39%) were occult HBV carriers and 81 (61%) were not. Nine patients developed HCC during the follow-up; eight were positive and one was negative for occult HBV (P = 0.002). CONCLUSIONS: The current observational cohort study showed that, among the HBsAg-negative patients with chronic hepatitis, HCC develops for the most part in carriers of occult HBV. Therefore, the evaluation of HBV genomes in chronic hepatitis patients appears to be a powerful tool for the identification of individuals at higher risk of HCC development.     

A study on sequence variations in pre‐S/surface,X and enhancer II/core promoter/precore regions of occult hepatitis B virus in non‐B,non‐C hepatocellular carcinoma patients in Taiwan

This study was to investigate the clinical significance and virologic factors of occult hepatitis B virus (HBV) infection in hepatocellular carcinoma (HCC) patients without hepatitis B surface antigen (HBsAg) or anti‐hepatitis C virus (non‐B, non‐C) in Taiwan. Serum HBV DNA (occult HBV) was detected in 90 of 222 non‐B, non‐C HCC patients and 24 of 300 non‐B, non‐C controls without HCC. Of 90 occult HBV‐infected HCC patients, the sequences of HBV pre‐S/surface, X and enhancer II/core promoter/precore genes were analyzed from 40 patients. Direct sequencing of such genes was also performed in 24 non‐B, non‐C controls without HCC and 40 HBsAg‐positive HCC controls. Compared with non‐B, non‐C controls without HCC, non‐B, non‐C subjects with HCC had significantly higher prevalence of occult HBV (p < 0.0001). Moreover, M1I and Q2K in pre‐S2 gene and G1721A were more common in occult HBV‐infected patients with HCC than in those without HCC. Compared with the HBsAg‐positive HCC controls, occult HBV‐infected HCC patients had higher frequencies of M1I and Q2K in pre‐S2 gene, G185R and S210N in surface gene, A36T and A44L in X gene, and G1721A in enhancer II gene, and had lower rates of pre‐S deletions and A1762T/G1764A, A1846T, G1896A and G1899A in core promoter/precore genes. Multivariate analysis showed Q2K in pre‐S2 gene, G1721A and A1846T were independent factors for occult HBV‐infected HCC. Our study suggested that the virological factors of HBV related to HCC were different between occult HBV‐infected and HBsAg‐positive patients. The G1721A, M1I and Q2K in pre‐S2 gene may be useful viral markers for HCC in occult HBV carriers. © 2009 UICC     

Past exposure to hepatitis B virus as a risk factor for hepatocellular carcinoma in patients with chronic liver disease.

The aim of the study was to determine whether past exposure to hepatitis B virus (HBV) influences the risk of the development of hepatocellular carcinoma (HCC) in Japanese patients with chronic liver disease (CLD). We conducted a hospital-based case-control study of 141 HCC patients with CLD and 151 controls with CLD but without HCC. Past exposure to HBV was assessed by antibody to hepatitis B core antigen (anti-HBc) positivity. Ninety-two patients (65%) with HCC were anti-HBc positive compared with 65 patients (43%) with CLD alone (P < 0.01). A multivariate analysis using logistic regression modelling revealed that anti-HBc positivity significantly increased the risk of the development of HCC [odds ratio (OR) 2.0, P = 0.01]. In the anti-HBc-positive patients, a significantly increased risk of HCC was seen among the patients positive for anti-HBc alone (OR, 2.6; P < 0.01). However, a significant OR was not obtained among the patients with a transient HBV infection implied by positivity for both antibody to hepatitis B surface antigen and anti-HBc (OR, 1.5; P = 0.48). These results indicate that past exposure to HBV is a risk factor for HCC in Japanese CLD patients, especially when they have no serological evidence of immunity to HBV.     

Hepatitis B virus reactivation in hepatocellular carcinoma patients undergoing transcatheter arterial chemoembolization therapy

Aim: The effect of transcatheter arterial chemoembolization (TACE) therapy on hepatitis B virus (HBV) reactivation in hepatocellular carcinoma (HCC) patients with prior resolved hepatitis B is not fully understood. Methods: From January 2006 to December 2010, 43 hepatitis B surface antigen (HBsAg)‐negative/anti‐hepatitis B core antigen (HBc) positive patients with newly diagnosed unresectable HCC were enrolled in the study. All underwent TACE therapy. Results: Four patients (9.3%) developed HBV reactivation with mild/moderate hepatitis. The median number of TACE cycles received was 3.5 (range 3–4 cycles). The median time interval between the occurrence of HBV reactivation and the completion of TACE therapy was 3 months (range 1–5 months) and their median HBV DNA level was 1.58 × 10⁴ IU/mL (range, 1.65 × 10³–6.42 × 10⁴ IU/mL). After the introduction of lamivudine at the occurrence of HBV reactivation, all had resolution of hepatitis. An exploratory analysis indicated that significant predictors of HBV reactivation included increased serum total bilirubin coexisting with cirrhosis and the total number of cycles of TACE received. Conclusion: The administration of TACE therapy may increase the risk of HBV reactivation in HBsAg‐negative/anti‐HBc‐positive patients diagnosed with unresectable HCC. Further studies are warranted to explore the optimal management of HBV reactivation in patients with prior resolved hepatitis B.     

Difference in histological activity of non-tumorous liver tissue between hepatitis B virus- and hepatitis C virus-associated hepatocellular carcinoma without cirrhosis

To elucidate the difference in the liver carcinogenetic process during hepatitis B virus (HBV) and hepatitis C virus (HCV) infections, non-tumorous liver tissues obtained from 10 patients who developed HBV-associated hepatocellular carcinoma (HCC) without cirrhosis were compared with those obtained from 26 patients who developed HCV-associated HCC without cirrhosis. The extent of fibrosis was similar in both groups. In contrast, necroinflammatory activities were significantly higher in patients with HCV than in patients with HBV. These results indicate that ongoing liver inflammation mediates the hepatocarcinogenesis more pronouncedly in HCV infection than in HBV infection.     

Role of hepatitis B virus genotype mixture, subgenotypes C2 and B2 on hepatocellular carcinoma: compared with chronic hepatitis B and asymptomatic carrier state in the same area

The role of genotype mixture and subgenotypes remains controversial in determining the clinical outcome of chronic hepatitis B virus (HBV) infection. We aimed to determine their role on the development and the recurrence of hepatocellular carcinoma (HCC). HBV genotypes, serum viral load and hepatitis B e antigen (HBeAg) seroconversion were determined in 462 HCC patients, 234 chronic hepatitis patients and 425 asymptomatic carriers born in Eastern China. In the 462 HCC patients, 62 (13.4%), 337 (72.9%) and 49 (10.6%) had HBV subgenotype B2, C2 and genotype mixture, respectively. Genotype mixture in HCC patients and hepatitis patients was associated with higher viral load than HBV C2 (P = 0.012, P = 0.000) and more frequent than asymptomatic carriers (P = 0.005, P = 0.000). HBV C2 was more prevalent in HCC patients compared with controls. Proportion of HBV B2 in HCC patients decreased consecutively from <30 to 50-59 years group (P = 0.024). Age-related changes of HBeAg seroconversion were not consistent with serum viral load in HCC patients with HBV B2 and genotype mixture, quite in contrast to hepatitis patients. By multivariate regression analysis, age >or=40 years and serum viral load (>or=10 000 copies/ml) were independently associated with hepatocarcinogenesis, whereas age 相似文献