Expression of hepatitis B surface antigen gene in yeast.

The DNA sequence coding for hepatitis B virus surface antigen (HBsAg) was placed under control of the repressible acid phosphatase promoter of the yeast Saccharomyces cerevisiae in a plasmid capable of autonomous replication in both yeast and Escherichia coli. Yeast transformed by this plasmid synthesized up to 5 X 10(5) molecules per cell of immunologically active HBsAg polypeptide in phosphate-free medium. The HBsAg polypeptides produced in the yeast cells were assembled into 20- to 22-nm spherical or oval particles and were immunogenic.     

Recombinant adenovirus induces antibody response to hepatitis B virus surface antigen in hamsters.

Recombinant adenoviruses carrying the hepatitis B virus surface antigen coding sequence in the adenovirus E3 region were constructed using DNA from either adenovirus type 5 or an adenovirus type 5 E3-region deletion mutant. Both of these recombinant adenoviruses replicated as efficiently as wild-type adenovirus in all human cells tested, including the human diploid cell strain WI-38. This indicates that insertion of the hepatitis B virus surface antigen gene into the E3 region does not significantly affect viral replication. Human cells infected with these recombinant adenoviruses secreted immunoreactive hepatitis B virus surface antigen. Since a practical small animal model for human adenoviruses was lacking, a hamster model was developed to evaluate the immunogenic potential of these recombinant adenoviruses. Upon intranasal inoculation, both wild-type adenovirus and the adenovirus E3-region deletion mutant replicated in the lungs of these animals and induced an antibody response against adenovirus. Hamsters similarly immunized with the live recombinant adenoviruses produced antibody against both adenovirus and hepatitis B virus surface antigen.     

乙型肝炎表面抗原基因在烟草中的表达

目的 建立植物中表达乙型肝炎表面抗原（HBsAg）的体系和方法,为研究植物乙型肝炎疫苗打下基础。方法 将HBsAg因基转入带有植物特异启动子及潮霉素筛选标记的双元载体pCAMBIA1301中,并将重组的载体转入农杆菌LBA4404,再通过农杆菌浸染的方法转化烟草。对筛选所得的转化株行鉴定。结果 转化株叶片进行β-葡萄糖苷酸酶（GUS）染色呈蓝色,表明靶基因被转入烟草并表达。应用HBsAg特异物引入转化株烟草的基因中扩增出678bp的条带。同时转化株叶片的粗提蛋白经酶联免疫吸附试验（ELISA）显示阳性结果,Western blot显示24000附近有明显条带。结论 烟草能够成功地表达乙型肝炎表面抗原。     

Treatment of hepatitis B surface antigen (HBsAg)-positive chronic hepatitis with recombinant alpha-A-interferon. Results of a phase II study

The HBsAg carrier state may present as chronic active hepatitis which may proceed to cirrhosis of the liver and to primary liver cell carcinoma. The large scale production of interferons made these substances available for long-term treatment. A deficiency in interferon production in chronic type B hepatitis presented the rational to treat this disease with interferon alpha-A. In this phase II-trial 3/31 patients eliminated HBsAg and 14/31 HBeAg. This was followed by normalisation of liver function tests and probably an improved prognosis. Efficiency of treatment was dependent on the interferon dose, the level of viral replication, the level of liver enzymes before treatment and concurrent infections (e. g. HIV infection). Reactivation occurred in five patients suggesting that the treatment period was too short in some individuals. Future studies will potentially improve efficiency by the modification of the interferon schedule and a better understanding of the mode of action of interferon.     

Change of hepatitis B surface antigen in patients with hepatitis during a five-year period.

Between January 1970 and December 1974, 122 cases of acute type B hepatitis were subtyped; 66 (54%) were of the ad type and 56 (46%) were of the ay type. "Cluster" cases (from a dialysis unit) were not considered. During the first period, subtype ad predominated, whereas in the second year there was a clear predominance of the ay subtype; the difference was statistically significant (P less than 0.02). In yearly periods the differences were significant between the years 1970 and 1974 (P less than 0.05), 1972 and 1974 (P less than 0.05), and 1972 and 1973 (P less than 0.05). If only patients without parenteral exposure are considered, there was clearly a shift between 1970-1972 and 1973-1974 in favor of the ay subtype (P less than 0.01). Since epidemiological factors such as injections and transfusions seem not to be responsible, it is suggested that a change of virus strain may be responsible for the different distribution of subtypes of hepatitis B surface antigen in the last year.     

慢性乙型肝炎肝组织内HBsAg、HBcAg的表达及临床研究进展

一直以来临床将血清乙型肝炎e抗原(HBeAg)、乙肝病毒DNA(HBV DNA)阳性作为乙肝病毒复制的标志,随着肝穿活检及抗病毒治疗的研究进展,肝活检组织中乙肝表面抗原(HBsAg)和乙肝核心抗原(HBcAg)的表达模式与血清乙型肝炎病毒(HBV)DNA定量、肝组织炎症活动度分级及纤维化分期之间关系的临床研究日益增多,本文就HBsAg和HBcAg在肝组织的表达模式及临床研究进展综述如下.     

Differential distribution of hepatitis B surface antigen and hepatitis B core antigen in the liver of hepatitis B patients.

One hundred liver biopsies from 100 patients with clinical presumptive diagnosis of hepatitis were examined by immunofluorescence for the presence of hepatitis B surface antigen (HBSAg) and hepatitis B core antigen (HBcAg). Of the 60 HBsAg-positive livers, 51 were diagnosed as chronic hepatitis on histological grounds, 6 as acute hepatitis, and 3 as "near-normal liver." From the 60 tissue-positive cases, 3 subjects were HBsAg seronegative. HBcAg was detected in 44 livers, all of which also had HBcAg in the localized in the cytoplasm and the membranes of the hepatocytes, and HBcAg in the nuclei and in 4 cases also in the cytoplasm. Predominant HBsAg expression in the cytoplasm was observed in near-normal liver, chronic persistent hepatitis, and cirrhosis with little activity. This correlated with the amount of ground glass hepatocytes in the biopsies. HBcAg and membrane-localized HBsAg were minimal in those conditions. HBcAg was most prevalent in patients with chronic aggressive hepatitis and active cirrhosis treated with immunosuppressive drugs, whereas the amounts of HBsAg and HBcAg in nontreated patients of those two groups and in acute hepatitis with signs of transition to chronicity were almost equal. HBsAg expression in liver cell membranes was most prominent in active forms of chronic hepatitis (chronic aggressive hepatitis and in active cirrhosis) and in acute hepatitis with signs of transition to chronicity. This observation correlated in the presence of HBcAg in the biopsies of those patients. In acute hepatitis both HBsAg and HBcAg were detected rarely and no membrane expression of HBsAg was observed. The over-all results show a significant relationship between the different degrees of accumulation of HBsAg and HBcAg in the liver and the various histological types of hepatitis and further suggest an interplay of both hepatitis B virus and host immune response in the development and pathogenesis of hepatitis B.     

A new hepatitis B virus surface antigen.

A new determinant of hepatitis B surface antigen (HBs Ag), g, which is distinct from the previously identified determinants a, d, y, w, and r, was studied. This new antigen tended to be associated with the d antigen; it was found in 97% of ad specimens and in 15% of ay specimens. With few exceptions, the atypical subtypes, adg- and ayg+, were identified in sera from blood donors, but not in sera from patients with acute viral hepatitis; this finding possibly reflects a reduced tendency of these viral strains to evoke a host immune response. The ability to identify g represents a refinement of HBs Ag serotyping that may be of value in the study of the epidemiology of hepatitis B virus.     

Hepatitis B surface antigen clearance in inactive hepatitis B surface antigen carriers treated with peginterferon alfa-2a

AIM: To examine the association between interferon(IFN) therapy and loss of hepatitis B surface antigen(HBs Ag) in inactive HBs Ag carriers. METHODS: This was a retrospective cohort study in inactive HBs Ag carriers, who were treatment-naive, with a serum HBs Ag level 100 IU/m L and an undetectable hepatitis B virus(HBV) DNA level( 100 IU/m L). All the 20 treated patients received subcutaneous PEG-IFN alfa-2a 180 μg/wk for 72 wk and were then followed for 24 wk. There were 40 untreated controls matched with 96 wk of observation. Serum HBs Ag, HBV DNA, and alanine aminotransferases were monitored every 3 mo in the treatment group and every 3-6 mo in the control group. RESULTS: Thirteen(65.0%) of 20 treated patients achieved HBs Ag loss, 12 of whom achieved HBs Ag seroconversion. Mean HBs Ag level in treated patients decreased to 6.69 ± 13.04 IU/m L after 24 wk of treatment from a baseline level of 26.22 ± 33.00 IU/m L. Serum HBV DNA level remained undetectable( 100 IU/m L) in all treated patients during the study. HBs Ag level of the control group decreased from 25.72 ± 25.58 IU/m L at baseline to 17.11 ± 21.62 IU/m L at week 96(P = 0.108). In the control group, no patient experienced HBs Ag loss/seroconversion, and two(5.0%) developed HBV reactivation.CONCLUSION: IFN treatment results in HBs Ag loss and seroconversion in a considerable proportion of inactive HBs Ag carriers with low HBs Ag concentrations.     

Alanine aminotransferase deficiency in a hepatitis B surface antigen positive patient presenting with acute hepatitis.

This report presents a hepatitis B surface antigen positive case presenting with acute hepatitis and with findings of low serum alanine aminotransferase in contrast to very high levels of aspartate aminotransferase. A 64 year-old female patient was admitted to our hospital with fatigue and jaundice. Hepatitis B surface antigen was positive. During follow up, aspartate aminotransferase levels remained very high, while alanine aminotransferase levels continued to be extremely low. Additionally, all of the patients five daughters had low alanine aminotransferase levels. The clinical importance of alanine aminotransferase deficiency is still unclear.     

Vertical transmission of the hepatitis B surface antigen.

Vertical transmission implies transmission of the hepatitis B virus from mother to infant. This occurs with great frequency (70 to 100 per cent) when the mother has acute hepatitis B near delivery. Studies indicate that transmission may occur transplacentally or during the birth process, Once infected, the infants apparently remain hepatitis B surface antigen carriers with persistent hepatitis indefinitely; The rate of transmission of the hepatitis B surface antigen from asymptomatic carrier mothers to their infants varies from 0 to 40 per cent in different areas of the world. The highest rate is in Taiwan where it was found that neonatal infection correlated with the height of complement fixation of surface antigen in the maternal serum. In our series of carrier mother-infant pairs, three infants became surface antigen positive as neonates; two became positive between three and six months; and one became positive between six and 12 months of age; Two babies developed extremely high titers of antibody to hepatitis B surface antigen the first year of life. In addition, 33 infants who were followed for from three to 42 months developed neither antigen nor antibody positivity.     

Immunogenicity of transgenic plant-derived hepatitis B surface antigen.

The focus of the Children's Vaccine Initiative is to encourage the discovery of technology that will make vaccines more readily available to developing countries. Our strategy has been to genetically engineer plants so that they can be used as inexpensive alternatives to fermentation systems for production of subunit antigens. In this paper we report on the immunological response elicited in vivo by using recombinant hepatitis B surface antigen (rHBsAg) purified from transgenic tobacco leaves. The anti-hepatitis B response to the tobacco-derived rHBsAg was qualitatively similar to that obtained by immunizing mice with yeast-derived rHBsAg (commercial vaccine). Additionally, T cells obtained from mice primed with the tobacco-derived rHBsAg could be stimulated in vitro by the tobacco-derived rHBsAg, yeast-derived rHBsAg, and by a synthetic peptide that represents part of the a determinant located in the S region (139-147) of HBsAg. Further support for the integrity of the T-cell epitope of the tobacco-derived rHBsAg was obtained by testing the ability of the primed T cells to proliferate in vitro after stimulation with a monoclonal anti-idiotype and an anti-idiotype-derived peptide, both of which mimic the group-specific a determinant of HBsAg. In total, we have conclusively demonstrated that both B- and T-cell epitopes of HBsAg are preserved when the antigen is expressed in a transgenic plant.     

HBsAg 及其与小、鼠白介素-18融合蛋白表达质粒的构建和DNA免疫

目的 观察乙型肝炎病毒表面抗原（HBsAg）基因表达质粒（pS)及其与鼠白细胞介素－18基因融合表达质粒（p18S)的构建及其诱导BALB／c鼠免疫应答的能力。方法 用聚合酶链反应（PCR）法获得S基因克隆到真核表达质粒pcDNA3.1 上,构建pS。另用三步PCR法将小鼠白细胞介素－18基因融合到S基因的5′端,再插入到pcDNA3.1 EcoRI位点上,构建了p18S。分别免疫10只和6只BALA/c小鼠,阴性对照组pcDNA3.1 免疫4只BALB／c小鼠。用酶联免疫方法检测每份血清的抗－HBs的效价。采用^3H标记法检测脾细胞HBsAg特异性细胞毒反应。结果 pS质粒免疫BALB／c小鼠多数于4周时出现抗－HBs,最高可达530mIU/ml,平均为135mIU/ml。p18S质粒也能诱导低水平的抗－HBs,平均为20mIU/ml。ps和p18S诱导的HBsAg特异性的细胞毒活性分别为37．1％和34％,而pcDNA3.1 载体质粒免疫小鼠血清未能检出抗－HBs,细胞毒活性为13．2％。结论 pS质粒能够有效地诱导BALB/c小鼠的体液和细胞免疫反应,白细胞介素－18基因与S基因融合表达质粒免疫似乎不能增强免疫应答,相反机体免疫应答下降。     

Expression of immunogenic epitopes of hepatitis B surface antigen with hybrid flagellin proteins by a vaccine strain of Salmonella.

A nonvirulent Salmonella dublin flagellin-negative, aromatic-dependent live vaccine strain has been used to express hepatitis B virus surface antigen epitopes in an immunogenic form. The envelope proteins of the virion are encoded by the S gene, which contains the pre-S1, pre-S2, and S coding regions. Synthetic oligonucleotides corresponding to amino acid residues S-(122-137) and pre-S2-(120-145) were inserted in-frame into the hypervariable region of a cloned Salmonella flagellin gene, and the recombinant plasmids were introduced into a flagellin-negative aroA mutant live vaccine strain of S. dublin, SL5928. The flagellin gene was expressed in bacteria carrying the plasmids as detected by immunoblotting with anti-flagellin (H1-d) serum. Both the S and pre-S2 epitopes were detected in bacteria carrying the relevant plasmid by immunoblotting with anti-HBs (antibody to hepatitis B virus surface antigen) and anti-peptide antisera. Animals immunized intramuscularly or orally with the live recombinant bacteria developed antibodies specific to these hepatitis B virus epitopes as detected by ELISA.     

Clinical utility of hepatitis B surface antigen quantitation in patients with chronic hepatitis B: a review

This clinically relevant review focuses on recent findings concerning hepatitis B surface antigen (HBsAg) quantitation in untreated patients and treated patients with chronic hepatitis B. Recent studies and emerging data have shown that both HBsAg and hepatitis B virus (HBV) DNA levels decline during the natural course of a chronic HBV infection; they are lowest in the inactive phase, which is also characterized by the highest HBsAg/HBV DNA ratio. It has been demonstrated that the combined use of HBsAg and HBV DNA levels might help in the identification of true inactive carriers with high accuracy. Retrospective analyses of HBsAg levels in patients undergoing therapy have suggested a role for HBsAg quantitation in monitoring the response to therapy. In comparison with nucleos(t)ide analogues (NAs), interferon-based therapy results in greater overall declines in serum HBsAg levels. A rapid on-treatment decline in HBsAg levels appears to be predictive of a sustained response. With the aid of HBsAg quantitation, it appears that we can anticipate an individualized approach to tailoring the treatment duration. The proposal of early stopping rules for patients not responding to pegylated interferon (according to a lack of any HBsAg decline) represents a step toward a response-guided approach. The development of stopping rules for patients treated with NAs is desirable for reducing the need for lifelong therapy. However, before stopping rules for antiviral therapy can be applied, we need to learn more about the kinetics of HBsAg declines during the natural history of the infection and as a response to therapy so that we can better define the best timing, the relevant HBsAg cutoff levels, and the best ways to apply these rules in clinical practice.