Fas/FasL在丁型肝炎发病机理中的作用

目的检测丁型肝炎病人肝组织Fas/FasL和HDAg表达,了解Fas/FasL在丁型肝炎致病机理中的作用。方法采用免疫组化单、双标记技术,检测48例丁型肝炎病人肝组织Fas、FasL和HDAg表达,以54例乙型肝炎作疾病对照。结果 Fas以肝细胞浆表达为主,HDAg以肝细胞核和胞浆表达为主,二抗原表达及分布呈一致性。FasL主要表达在浸润淋巴细胞浆、肝细胞核,与HDAg表达及分布不全一致,丁型肝炎组Fas、FasL检出率及表达强度较对照组高,三成分在各型肝炎中的表达强度有显著性差别意义。结论 Fas、FasL和HDAg均与肝组织炎症活动和病理损害相关,HDV感染可诱导肝细胞大量表达Fas,CTL通过Fas/FasL途径介导的肝细胞凋亡与HDV致病可能有关。     

丁型肝炎病人肝组织HDAg和 Fas表达及其相互关系

目的 研究丁型肝炎病人肝组织Fas和HDAg表达及其相互关系,了解Fas在HDV致病机制中的作用。方法 应用免疫组化单、双标记技术,检测48例丁型肝炎病人肝组织Fas、HDAg表达,以54例乙型肝炎病人肝组织Fas表达作对照。结果 Fas以肝细胞浆表达为主,HDAg以肝细胞核和胞浆表达为主,二抗原表达及分布呈一致性。各型肝炎中Fas和HDAg表达有统计学差别,两种抗原的分布和表达强度与肝细胞炎症活动和病理损害程度相关。结论 Fax和HDAg表达及分布密切相关,与肝组织炎症活动和病理损害相关,此提示HDV感染可诱导肝细胞表达Fas,Fas及其所介导的肝细胞凋亡在HDV致病机制中可能有重要作用。     

国内丁型肝炎流行概况

本文就全国各地报告的丁型肝炎从血清学标志和乙肝肝组织活检资料方面对各地ＨＤＶ感染情况、ＨＢＶ标志阴性中检出ＨＤＶ标志阳性以及在各型乙肝、肝硬化、肝癌和ＨＢｓＡｇ携带者ＨＤＶ感染的检出率进行了综合分析。据不完全统计，截至１９９２年底，全国有６７７３例乙肝患者检测了血清ＨＤＶ感染标志，阳性６０２例，各地检出率１．７３￣３７．５％，平均为８．８９％；检查乙肝肝组织中ＨＤＡｇ２７９７例，阳性者２３３例，各     

丁型肝炎

丁型肝炎病毒抗原的原核表达及抗原性分析——江永珍等（中国疾病预防控制中心病毒病预防控制所北京100052）；《中华实验和临床病毒学杂志》，2006，20（2）：38—41[目的：利用含T7启动子和HiS纯化标签pRSETB质粒构建丁型肝炎病毒抗原（HDAg）重组表达质粒（pRSETB—HDAg），转化宿主菌，表达并纯化，获得生物活性高抗原性强的基因工程重组抗原。方法：将中国河南株HDAg基因片段插入pRSETB表达质粒，转化BL21宿主菌，经IPTG诱导表达。表达产物经Chelating亲和层析纯化后，采用EIA方法分析HDAg的抗原性。结果：重组HDAg稀释1000倍（10ng／ml）仍与抗体有较强的反应。     

HDAg与HBsAg／HBcAg在丁型肝炎病人肝组织中的表达

研究HDAg、HBsAg和HBcAg在丁型肝炎病人肝组织中表达及关系 ,探讨HDV致病机理。应用免疫组化双重染色及连续切片技术 ,检测 79例丁型肝炎病人肝组织HDAg、HBsAg和HBcAg表达 ,以 5 2例乙型肝炎作对照。丁型肝炎HBsAg、HBcAg检出率 (81%、71% )较乙型肝炎 (94%、92 % )低 (P <0 0 5或 0 0 1)。HDAg以肝细胞核表达为主 ,HBsAg以肝细胞浆表达为主 ,HDAg和HBsAg表达强度及阳性细胞分布呈一致性 ,均与肝组织炎症活动和病理损害程度相关 (P <0 0 1)。HBcAg以肝细胞核表达为主 ,HBcAg阳性细胞明显少于HDAg阳性细胞。HDV感染可抑制HBV抗原表达 ;在HDV致病机制中既有HDV的直接细胞毒性作用 ,也有HBV和HDV的协同作用。     

乙丁型肝炎病毒感染树鼩肝组织Fas和ICE的表达

目的探讨乙型并丁型肝炎病毒感染树鼩肝组织中Ｆａｓ和ＩＣＥ表达与ＨＤＶ感染之间的关系。方法采用免疫组织化学技术对４５份ＨＤＶ／ＨＢＶ感染树鼩肝组织中ＨＤＡｇ、Ｆａｓ和ＩＣＥ的表达进行了检测。结果４５份肝组织中有 ３９份可检出 Ｆａｓ（阳性率 ８６．７％）,有 ４３份可检出 ＩＣＥ（阳性率 ９５．６％）。 Ｆａｓ和 ＩＣＥ在肝细胞质和（或）膜上表达,以肝细胞质内表达为主。 ＨＤＡｇ表达与 Ｆａｓ和 ＩＣＥ表达之间有显著相关性（ ｘ２值为 ２９．２和 ３６．２,Ｐ＜ ００１）, ＨＤＡｇ表达越强, Ｆａｓ和 ＩＣＥ表达也越强。结论肝细胞内的 ＨＤＡｇ表达可诱导Ｆａｓ和 ＩＣＥ的表达。     

乙丁型肝炎病毒感染树鼠句肝组织Fas和ICE的表达

目的探讨乙型并丁型肝炎病毒感染树鼠句肝组织中Fas和ICE表达与HDV感染之间的关系。方法采用免疫组织化学技术对45份HDV/HBV感染树鼠句肝组织中HDAg、Fas和ICE的表达进行了检测。结果45份肝组织中有39份可检出Fas(阳性率86.7%),有43份可检出ICE(阳性率95.6%)。Fas和ICE在肝细胞质和(或)膜上表达,以肝细胞质内表达为主。HDAg表达与Fas和ICE表达之间有显著相关性(χ2值为29.2和36.2,P＜0.01),HDAg表达越强,Fas和ICE表达也越强。结论肝细胞内的HDAg表达可诱导Fas和ICE的表达。     

Fas抗原在乙型病毒性肝炎肝组织中表达的研究

目的 检测乙型病毒性肝炎肝组织中Fas抗原,以探讨Fas抗原的表达与肝细胞损伤之间的关系。方法 以免抗—Fas多克隆抗体,采用免疫组织化学技术检测33例急性重型肝炎,23例慢性迁延型肝炎,32例慢性活动性肝炎,21例活动性肝硬化和13例慢性重型肝炎肝组织中Fas抗原表达情况。结果 上述5组分型中Fas抗原检出率分别为90.9％、43.5％、78.1％、85.7％和92.3％。急性重型肝炎Fas抗原分布于大片坏死区残留肝细胞内,慢性肝炎和活动性肝硬化Fas抗原阳性细胞分布于碎屑状坏死灶周围或假小叶周边部。肝组织病理改变越重,Fas表达越强。结论 Fas抗原的表达与肝细胞损伤密切相关。     

HDV感染后乙型肝炎患者血清肝炎病毒标志物的变化

目的　分析HDV感染患者血清病毒性肝炎标志物的变化和意义 ,探讨HDV致病机理。方法　对 469例HDV阳性乙型肝炎患者常见各类型病毒性肝炎血清标志物的变化等作统计分析 ,以 2 13例HDV( -)乙型肝炎患者作对照。结果　HDV感染后血清HBeAg检出率降低 (P <0 .0 1)。在HDV ( +)HBVDNA( -)组 ,HBeAg( -)的机会大 (P <0 .0 1)。在急性肝炎、重型肝炎和肝硬化患者HDAg( +)HBeAg( -)为主要血清病毒表现形式 (P <0 .0 1或 0 .0 5 )。HDV感染后合并其它肝炎病毒感染率高于乙型肝炎组。结论　HDV感染可抑制HBV复制或HBeAg表达 ,混合感染HDV的乙型肝炎中HDV的直接细胞毒性作用可能起主要致病作用。重叠感染HDV的乙型肝炎患者其病情重、病死率高和容易慢性化。     

“Defective” mutations of hepatitis D viruses in chronic hepatitis D patients

AIM: To verify whether "defective" mutations existed in hepatitis D virus (HDV). METHODS: Hepatitis delta antigen (HDAg)-coding sequences were amplified using Pfu DNA polymerases with proof-reading activities from sera of five patients with chronic hepatitis D. Multiple colonies were sequenced for each patient. Pfu analyzed a total of 270 HDV clones. Three representative defective HDV clones were constructed in expression plasmids and transfected into a human hepatoma cell line. Cellular proteins were extracted and analyzed by Western blot. RESULTS: Four of five cases (80%) showed defective HDV genomes in their sera. The percentage of defective genomes was 3.7% (10/270). The majority (90%) of the defective mutations were insertions or deletions that resulted in frameshift and abnormal stop translation of the HDAg. The predicted mutated HDAg ranged from 45 amino acids to >214 amino acids in length. Various domains of HDAg associated with viral replication or packaging were affected in different HDV isolates. Western blot analysis showed defected HDAg in predicted positions. CONCLUSION: "Defective" viruses do exist in chronic HDV infected patients, but represented as minor strains. The clinical significance of the "defected" HDV needs further study to evaluate.     

慢性乙型肝炎患者血清前S1抗原检测的意义

目的探讨PreS1抗原检测的临床价值。方法采用酶联免疫吸附试验法检测421例慢性乙型肝炎患者血清PreS1抗原和HBV标记物;采用荧光定量PCR法检测HBVDNA。结果在421例慢性乙型肝炎患者中,HBVDNA阳性者367例,其中PreS1Ag阳性者188例(51.2%),HBeAg阳性者119例(32.4%),后两者有显著性差异(P0.01);在高HBVDNA载量(105～107copies/ml和107copies/ml)组患者中,PreS1Ag阳性率(60.2%,60.0%)显著高于HBVDNA阴性组(33.3%)和低载量(103～105copies/ml)组(41.9%,P0.01);但在421例患者中,PreS1Ag阳性率(48.9%)低于HBVDNA(87.2%,P0.01)。结论 PreS1Ag能够较HBeAg更好地反映HBV在体内的复制状态,但尚不能代替HBVDNA的检测。     

Apoptosis mediated by the Fas system in the fulminant hepatitis by hepatitis B virus

The pathogenesis of the fulminant hepatitis B is poorly understood and both viral factors and the hosts immune response play a role. Previous studies in liver tissues of patients with chronic hepatitis B showed overexpression of Fas antigen and this was correlated with the activity of the hepatitis. The present study was done to determine the role of Fas in fulminant hepatitis B and the virological characteristics of hepatitis B infection.
We studied three patients with fulminant hepatitis B. HBV-DNA was detected by dot-blot hybridization and polymerase chain reaction. The S and C gene were sequenced. Levels of serum soluble Fas antigen were detected by enzymoimmunoassays procedure. Apoptosis was determined by the TUNEL technique. Fas antigen expression was evaluated by a immunoperoxidase method. Ten healthy subjects acted as controls.
The three patients showed a high expression of Fas antigen particularly among infiltrating lymphocytes; in these areas we also found many cells with in situ DNA nick labelling signals in the nuclei of most viable hepatocytes. Serum levels of soluble Fas antigen were higher in patients with fulminant hepatitis B than in controls. No specific genome mutations of hepatitis B virus were found.
These data suggest that the Fas system involved in the liver injury of patients with fulminant hepatitis B.     

Significance of hepatitis B virus surface antigen, hepatitis C virus expression in hepatocellular carcinoma and pericarcinomatous tissues

AIM： To investigate the correlation between hepatitis B virus surface antigen （HBsAg）, hepatitis C virus （HCV） expression in hepatocellular carcinoma （HCC）, the HAI score of the noncancerous region of the liver and the serum Alpha fetoprotein （AFP） level.
METHODS： The patterns of HBsAg and HCV in 100 cases of HCC and their surrounding liver tissues were studied on paraffin-embedded sections with immunohistochemistry, the histological status was determined by one pathologist and one surgeon simultaneously using the hepatitis activity index （HAIl score, and AFP was detected by radioimmunity. The study included 100 consecutive patients who underwent curative resection for HCC. Based on HBsAg and HCV expression, the patients were classified into 4 groups： patients positive for HBsAg （HBsAg group）, patients positive for HCV （HCV group）, patients negative for both HCV and HBsAg （NBNC group） and patients positive for both HBsAg and HCV （BC group）.
RESULTS： The BC group had significantly higher HAI scores than the other three groups. （BC 〉 HCV 〉 HBsAg 〉 NBNC）. HBV and HCV virus infection was positively correlated with HAI （rs = 0.39, P = 0.00011. The positive rate of AFP （85.7%） and the value of AFP （541.2 ng/mL） in the group with HBV and HCV co-infection were the highest among the four groups. The positive rate （53.3%） of AFP and the value of AFP （ 53.3 ng/mL） in the group with none-infection of HBV and HCV were the lowest. HBV and HCV virus infection was positively correlated with AFP（rs = 0.38, P = 0.0001）.
CONCLUSION： The AFP increase in patients with liver cancer was positively correlated with the infection of HBV and HCV. The-serum AFP elevation by the infection of HBV and HCV is one of mechanisms which lead to hepatocarcinogenesis, and the antivirus intervening treatment of hepatitis is significant for the prognosis of liver cancer. From our Spearman＇s rank correlation analysis, we can conclude that the severity of virally induced     

丁型肝炎病毒核酶结构改建及其对丙型肝炎病毒RNA的剪切活性

目的 观察经过结构改建的丁型肝炎病毒核酶（HDV核酶）对丙型肝炎病毒（HCV）基因组RNA是否存在剪切作用。方法 对HDV基因组核酶的茎IV区和底物结合区进行改建,获得3种针对HCV RNA的HDV核梅RzCI、RzC2和RzC3。体外转录制备含HCV RNA 5′－非编码区（5′-noncoding region,5′-NCR)及部分C区的底物RNA（HCV RNA5′-NCR-C）,进行5′端放射性标记。在一定的pH、温度、Mg^2 浓度和有或无去离子甲酰胺存在等条件下,将核酶和底物按摩尔比100：1混合,反应2h后聚丙烯酰胺凝电泳,放射自显影,推算剪切百分率。结果 RzCI和RzCI2可剪切HCV RNA5′-NCR-C,在适宜浓度的去离子甲酰胺存在时剪切效率较高,RzC3对底物几乎无剪切活性。结果 设计得当的HDV基因组核酶可以剪切异源性RNA分子HCV RNA。     

乙型肝炎病毒表面抗原一级结构多态性的初步研究

目的 研究慢性乙型肝炎患者体内乙型肝炎病毒（HBV）表面抗原一级结构的多态性。方法 设计特异性引物，自7例慢性乙型肝炎患者血清中扩增S基因全长或全基因组片段，TA克隆法克隆到T载体中，随机选择克隆测序。结果 共20个克隆被测序。20个克隆全S蛋白的总一致率仅为32．0％，13株全长为401氨基酸残基的克隆氨基酸一致效率为82．5％。患者血清中发现2株克隆编码截短型表面抗原中蛋白，在前S1或前S2的免疫决定区或可能的肝细胞结合部位均发现缺失突变。结论 慢性乙肝患者体内存在HBV准种群，病毒编码的截短型表面抗原中蛋白可为HBV诱导原发性肝癌提供一种途径，应加强对HBsAg一级结构多态性的研究。     

Measurement of hepatitis B virus core-related antigen as predicting factor for relapse after cessation of lamivudine therapy for chronic hepatitis B virus infection

Background

Prolonged lamivudine therapy has two major problems: breakthrough hepatitis during treatment and relapse of aminotransferase (ALT) after cessation of the therapy. The aim of this study was to examine factors that could predict ALT flare after stopping lamivudine therapy.

Methods

We analyzed 22 Japanese patients with chronic hepatitis B infection, in whom lamivudine therapy was stopped after HBV DNA level had been gone undetectable (<3.7 LGE/ml) during at least six consecutive months. The post-treatment followed up was carried for 28 months in median (range 9–41). HBV core-related antigen (HBcrAg) assay was assessed using newly developed assay.

Results

After cessation of lamivudine therapy, 11 patients (50%) had relapsed (reactivation of serum ALT >80 IU/l, relapsers) and remaining 11 (50%) did not relapse (non-relapsers). In the univariate comparison of relapsers versus non-relapsers, HBcrAg level at lamivudine cessation point (4.5 ± 1.0 versus 3.4 ± 0.9; p = 0.0145) has been shown as a significant predictive factor for non-relapse. All patients with HBcrAg <3.0 log U/ml at the cessation point had no ALT flares. Multivariate analysis on effects of 10 factors (age, sex, cirrhosis, pretreatment ALT level, HBV DNA level, HBcrAg level, mean months till undetectable HBV DNA, duration of undetectable HBV DNA and HBcrAg level at lamivudine cessation point), indicated that HBcrAg level at lamivudine cessation point <3.4 log U/ml was the only independent predictive factor for absence of the post-treatment relapse.

Conclusions

HBcrAg level at lamivudine cessation point might be useful as a prognostic predictor of response to lamivudine therapy cessation. The measurement of HBcrAg is a useful additional test for monitoring chronic HBV infection.