Antibody responses in mice stimulated by various doses of the potato-derived major surface antigen of hepatitis B virus.

The ability of potato-derived major surface antigen of hepatitis B virus (P-HBsAg) to elicit antibody responses to different dosages of P-HBsAg ranging from 0.02 to 30 µg administered orally in mice was examined. All immunized groups produced specific serum IgG and fecal IgA antibodies against P-HBsAg, even at low levels (<5 µg), after administration of a 0.5-µg yeast-derived HBsAg (Y-HBsAg; LG Life Sciences, Republic of Korea) booster.

Oral immunogenicity of potato-derived HBsAg middle protein in BALB/c mice.

The antibodies to preS2 synthetic peptides have been probed to neutralize hepatitis B virus (HBV), and also the addition of preS2 sequence could enhance the antibody response compared with a conventional vaccine in the non- and low responders. Previously, we generated transgenic potatoes expressing middle protein, which contains additional 55 amino acid preS2 region at the N-terminus of the S protein, of HBV to determine the feasibility of developing a plant-delivered HBV vaccine. In this study, we monitored the immune response after induction of immunoglobulin by boosting and assessed the efficacy of the mucosal immune response with regard to generate IgA antibodies. The HBsAg middle protein expressed in our transgenic potatoes was well immunized at low antigenic quantities in mice and the induced anti-S or anti-preS2 antibodies were sustained for the whole period without decrease. Orally delivery of plant-derived HBsAg middle protein to mice resulted in fecal anti-S or anti-preS2 as well as serum IgG. In addition, we used antibodies induced from the immunized mice with the potato-derived rHBsAg in competition assay as competitors to confirm the binding ability of preS2 antibodies to surface antigen of hepatitis virus. Anti-preS2 antibodies induced from immunized mice with transgenic potatoes effectively competed with anti-preS2 murine antibody H8 as expected. From these results, the inclusion of preS2 antigen to HBV plant vaccine may provide additional protective immunity in the HBV prevention.

Danshao huaxian capsule in treatment of decompensated cirrhosis resulting from chronic hepatitis B.

BACKGROUND: The prognosis of decompensated cirrhosis resulting from chronic hepatitis B is poor, and the benefits of treatment with interferon are outweight serious side-effects and the risk of fatal exacerbation of disease. Danshao huaxian capsule rapidly reduces hepatitis B virus (HBV)-DNA in serum to undetectable levels. METHODS: A total of 35 patients with chronic hepatitis B and decompensated cirrhosis were treated with danshao huaxian 1.2 g. p.o. tid daily. Before the treatment, HBV-DNA in serum was positive in all patients. Ten patients had Child-Pugh class B and 25, class C hepatitis B. Seven patients underwent liver transplantation within 6 months of initial treatment. Of the 10 patients of class B, 5 died within 6 months, and the other 5 did not complete the treatment for some reasons; the 25 patients of class C were treated for at least 6 months (mean=19 months). RESULTS: In most of the 25 patients, liver function was improved slowly but markedly after 9 months of treatment, showing a decreased level of serum bilirubin from 67+/-13 to 30+/-4 micromol/L (P<0.05, baseline vs. 6 months), an increased level of serum albumin from 27+/-1 to 34+/-1 g/L (P<0.05) and a decreased level of Child-Pugh score from 10.3+/-0.4 to 7.5+/-0.5 (P<0.05). Three patients developed resistance to danshao huaxian because of a mutation in the YMDD motif, but liver function was not deteriorated. Inhibition of viral replication with danshao huaxian resulted in a significant improvement of liver function in patients with decompensated HBV cirrhosis, but the long-term results remain uncertain. CONCLUSION: Danshao huaxian capsule is effective in inhibiting viral DNA replication in patients with decompensated cirrhosis and making clinical improvement.

Standardization and validation of an alkaline phosphatase-linked immunoassay to quantify a plant-derived antibody directed against the hepatitis B virus surface antigen.

Immunopurification is one of the most effective chromatography steps to purify the hepatitis B surface antigen, which have successfully been used as an active pharmaceutical ingredient of hepatitis B vaccines. Plant-derived antibodies could be an appropriated ligand for such purposes because plants are the most cost-effective production systems and have the additional advantage that plant viruses cannot infect humans. In this work, a polyclonal antibody alkaline phosphatase-linked immunoassay was standardized and validated to quantify a plant-derived antibody directed against the HBsAg. The validation of an immunoassay to quantify plantibodies is a relatively complex task due to the complexity of the plant extract, the low level of expression of this molecule, and the potential interferences of endogenous peroxidases contributed by plants. These results allow estimating the plant-derived antibody concentration up to 3.81 ng/mL with high specificity, precision, and repeatability. The working range of the standard curve was between 3.81 and 60 ng/mL, and the intra- and inter-variation coefficients were between 10% and 20% in a production process's sample dependent way. This enzyme-linked immunosorbent assay is considered valuable to improve the design of the purification process and also to obtain a better estimation of the antibody expression level and process's recovery.

Thymosin alpha1 accelerates restoration of T cell-mediated neutralizing antibody response in immunocompromised hosts.

Thymosin alpha1 is a biological response modifier that has been used clinically for the treatment of chronic hepatitis B viral infection. Both immunomodulatory and immediate intracellular mechanisms have been postulated to explain the effect of thymosin alpha1 on hepatocytes infected with hepatitis B virus (HBV). Here, we established a new animal model and the related suitable conditions to access the thymosin activity by means of measuring the production of neutralizing antibody against hepatitis B surface antigen (HBsAg). We proved that chemically synthesized thymosin alpha1 restored the T cell-mediated antibody production following its suppression in mice by 5-fluorouracil (5-FU), and found that thymosin alpha1 showed activity at a low dose of 30 microg/kg. Further studies utilizing the flowcytometric analysis showed that thymosin alpha1 at this dose accelerated the replenishment and maturation of thymocytes while the expression of Smoothened (Smo) of the Hedgehog (Hh)-signaling in CD4-CD8- thymocytes, the potent negative regulator of proliferative responses, was not affected. The restoration of some of the defects in the host defense systems may facilitate elimination of infectious agents, and the present study provides a novel model to define the restoration of T cell-mediated immune responses to hepatitis B virus in vivo.

[Immunomodulation by herbal agents. A double-blind study in a medical university hospital involving a hepatitis B vaccine adjuvant model]. / Immunmodulation durch pflanzliche Wirkstoffe. Doppelblindstudie an einer medizinischen Universitätsklinik am Adjuvansmodell zur Hepatitis-B-Impfung.

AIMS: Using the hepatitis B vaccination as a model, to investigate the extent to which the herbal immunomodulator, Esberitox N, supports seroconversion. METHOD: 346 medical students participated in the placebo-controlled, randomized double-blind study. They took 3 x 2 tablets of the test substances daily, beginning 3 days prior to the injection and ending two weeks after it. The target outcomes were seroconversion and the level of the anti-HBs titer. RESULTS: The data of 157 volunteers treated with the test substance, and 161 treated with placebo were analysed. After the first injection, the seroconversion rate was 22% in both test substance and placebo groups, and showed no advantage for the volunteers receiving the test substance. After the second injection, 89% of all members of each group revealed seroconversion. After the first injection, anti-HBs titers were appreciably higher in the test substance group (n = 34) than in the placebo group (n = 36; PWilcoxon = 0.003). The respective median values were 37.0 IU/L (95% CI: 18-68) and 15.5 IU/L (95% CI: 8-30). CONCLUSION: The immunomodulator tested has negligible influence on seroconversion, but does enhance the immune response of subjects experiencing seroconversion.

[Breast-feeding by mothers with positive serum hepatitis B virus test].

Sixty one samples of colostrum from mothers with positive serum Hepatitis B virus (HBV) test were collected and determined for serial HBV markers along with HBV DNA. Results showed that HBsAg, HBeAg and anti-HBc were most likely transferred by milk, while anti-HBs and anti-HBe had less possibility to enter the colostrum (P < 0.01). The overall HBV DNA positive rate was 42.6% in the 61 samples. The various HBV markers detected in mothers' serum presented a high consistency with those discovered in the corresponding milk samples. There were significantly higher rate of milk HBV transmission from mothers with serum positive HBsAg, HBeAg and anti-HBc or either one of them than those with positive for anti-HBs and/or anti-HBe.

Influence of perceived psychological stress and distress on antibody response to low dose rDNA hepatitis B vaccine.

The present study focused on the relationship between psychological stress and immune reaction to a novel antigen. Participants completed questionnaires on daily hassles, psychoneurotic complaints, coping style, and loneliness, 2 and 6 months after the first of a series injections with a low dose recombinant DNA hepatitis B vaccine. Antibody response was determined 7 months after the first vaccination. Based on the psychological questionnaires two different stress measures were calculated: a Stress Index score-month-2 and a Stress Index score-month-6 indicating stress levels experienced at the beginning and at the end of the study respectively. Antibody levels were found to be negatively related with the Stress Index score-month-2. Although the influence of psychological stress reported on month 6 tended to be in the same direction, this effect was not significant. Coping styles and loneliness were not associated with antibody formation. These results suggest that antibody formation to rDNA hepatitis B vaccine is negatively influenced by psychological stress.

Reversal of the immunosenescent phenotype by dehydroepiandrosterone: hormone treatment provides an adjuvant effect on the immunization of aged mice with recombinant hepatitis B surface antigen.

This study sought to establish whether administration of dehydroepiandrosterone (DHEA) or its sulfate derivative to aged mice could effectively correct the immunosenescent phenotype. Supplemental DHEA sulfate and topical DHEA fully corrected the age-associated dysregulated production of T cell lymphokines by cells from all of the different lymphoid organs tested. Either DHEA or DHEA sulfate supplementation promoted enhanced antibody responses against recombinant hepatitis B surface antigen (rHBsAg) by the aged recipients when incorporated directly into the vaccine. When DHEA was provided either topically or was incorporated directly into vaccine, vigorous primary and secondary antibody responses were detected in the aged mice given a single administration of DHEA, regardless of the mode of administration. It was also established that DHEA treatment could enhance specific antibody responses to rHBsAg in aged animals that had previously not been effectively immunized by conventional vaccination procedures.

Effects of zinc supplementation on zinc status and immunity in haemodialysis patients.

The depression of immunity to various antigens in chronic uremia is a frequently encountered phenomenon. Zinc deficiency might well be an important factor in its genesis. The aim of this study was to investigate the role of zinc deficiency in this reduced immune response. Two groups of 7 patients on haemodialysis who had failed to respond with seroconversion to an earlier vaccination against hepatitis B were revaccinated. One group received zinc by the addition of zinc chloride to the dialysate. Before initiation of the study zinc in plasma and leucocytes was measured. No difference in plasma and leucocyte zinc was observed between the two groups. Zinc in leucocytes was lower in patients than in a group of healthy volunteers (61.5 pmol/10E6 cells +/- 4.6 versus 73.8 +/- 5.6, p less than 0.005). Plasma zinc showed no difference between patients and healthy volunteers. During zinc supplementation zinc in plasma rose in the patient group receiving zinc (10.4 mmol/L +/- 1.5 to 14.2 +/- 1.9, p less than 0.005). However, no rise in leucocyte zinc was seen. At the end of the trial seroconversion had occurred in 2 patients in each group. It is concluded that zinc supplementation in haemodialysis patients does not lead to the restoration of leucocyte zinc to normal levels. Neither did it lead to an enhanced antibody response in our population after revaccination of haemodialysis patients against hepatitis B.

Hepatitis B-like infection in a Pacific white-sided dolphin (Lagenorhynchus obliquidens).

A Pacific white-sided dolphin (Lagenorhynchus obliquidens) developed clinical signs, serum biochemical values, and serologic viral markers consistent with chronic persistent hepatitis caused by a hepatitis B-like virus. The hepatitis had a sporadic cyclical pattern of lethargy, inappetance, and icterus, with leukocytosis and increased serum activities of alanine transaminase, aspartate transaminase, and gamma-glutamyltransferase. The serum from this dolphin contained hepatitis B virus core antibodies, hepatitis B surface antibodies, and hepatitis B viral DNA. Supportive treatment consisted of administration of antibiotics, cimetidine, menadiol sodium diphosphate, and vitamin/dextrose supplementation. A clinically normal killer whale (Orcinus orca) housed in the same pool had serum hepatitis B surface antibodies, suggesting immunologic responsiveness and that this disease was not species-specific.

A large outbreak of acupuncture-associated hepatitis B.

An epidemiologic investigation of an acupuncturist's practice in Rhode Island identified 35 patients who were infected with hepatitis B virus during 1984. Of 366 patients seen by the acupuncturist during 1984, 316 (86%) completed questionnaires and submitted serum for hepatitis B serology. Use of tests for immunoglobulin M antibody to hepatitis B core antigen (IgM anti-HBc) identified 17 case-patients who otherwise may have gone undetected. Thirty-four of the 35 case-patients were treated in only one of the two clinics run by the acupuncturist. Patients who received a greater number of acupuncture needles during their treatment course were more likely to have been infected; the attack rate for patients who received less than 150 needles was 9%, compared with 33% for patients who received greater than or equal to 450 needles (p less than 0.001). Attack rates were higher during a one-month period when the index case-patient was more likely to have been viremic than during any other period in 1984 (relative risk = 4.1, 95% confidence interval = 2.3-7.3). While observing the acupuncturist's technique, the investigators noted several potential mechanisms for needle contamination. This study highlights the potential for transmission of hepatitis B in situations of repeated needle use.

Hepatocellular carcinoma, hepatic cirrhosis, and hepatitis B virus infection in Nigeria.

A total of 200 consecutive patients bearing histologically confirmed primary hepatocellular carcinoma (PHC) were studied at University of Calabar Teaching Hospital (CTH), Calabar, Nigeria in 5 years and compared with 150 patients with metastatic hepatic carcinoma (MHC). Four hundred symptomless non-icteric nontumor-bearing individuals were noncontemporaneous controls. Sera (a total of 750) were assayed for markers of hepatitis B virus (HBV) infection; hepatitis B surface antigen (HBsAg), hepatitis B core antibody (anti-HBc), and hepatitis B surface antibody (anti-HBs). Specimens which were positive for HBsAg also were examined for hepatitis E antigen (HBeAg) and antibody (anti-HBe). The results show that PHC was associated with HBV seropositivity in 80% of patients, and postnecrotic (macronodular) cirrhosis of the liver in 90% clearly indicating a strong association between primary liver cancer and HBV infections and liver cirrhosis. The main factor associated with seropositivity among normal controls was a large number of therapeutic injections. Seropositives received over twice as many injections as seronegatives. Public health measures are urgently required to prevent parenteral transmission of HBV in Nigeria.

Aflatoxin exposure, hepatitis B virus infection and liver cancer in Swaziland.

A study was carried out in Swaziland to assess the relationship between aflatoxin exposure, hepatitis B infection, and the incidence of liver-cell carcinoma, which is the most commonly occurring malignancy among males in Swaziland. Levels of aflatoxin intake were evaluated in dietary samples from households across the country, and crop samples taken from representative farms. Prevalence of hepatitis B markers was estimated from the serum of blood donors, and liver cancer incidence was recorded for the years 1979-83 through a national system of cancer registration. Across 4 broad geographic regions, there was a more than 5-fold variation in the estimated daily intake of aflatoxin, ranging from 3.1 to 17.5 micrograms. The proportion of HBV-exposed individuals was very high (86% in men), but varied relatively little by geographic region; the prevalence of carriers of the surface antigen was 23% in men, and varied from 21 to 28%. Liver cancer incidence varied over a 5-fold range, and was strongly associated with estimated levels of aflatoxin. In an analysis involving 10 smaller subregions, aflatoxin exposure emerged as a more important determinant of the variation in liver cancer incidence than the prevalence of hepatitis infection. Aflatoxin estimates from crop samples appeared to be a reasonable surrogate for dietary measurements. A comparison with dietary aflatoxin levels measured in an earlier survey in Swaziland suggested that programmes aimed at reducing contamination levels had had some success.

Outbreak of hepatitis B associated with acupuncture.

In the period February to May 1980, acute hepatitis B occurred in six patients who had received acupuncture at a chiropractic clinic. Of persons attending the clinic (October 1979 to April 1980), hepatitis B occurred more frequently among those who had received acupuncture (6 of 103) than among those who had not (0 of 408; P less than .0001). Hepatitis patients had received acupuncture during two exposure periods: November 27-28, 1979, and February 19-20, 1980. During these periods the clinic reused acupuncture needles after immersing them overnight in a 1:750 solution of benzalkonium chloride. The sequence of acupuncture sessions suggests that hepatitis B virus remained infectious on the acupuncture needles for 20 to 27 hours despite this procedure. It is recommended that only either disposable needles or reuseable needles that have been physically cleaned and autoclaved after each use be used in acupuncture procedures.

[Clinical and immunological studies of an experimental series of vaccines against hepatitis B]. / Klinicheskie i immunologicheskie issledovaniia éksperimental'nykh serii vaktsiny protiv gepatita B.

Two experimental lots of hepatitis B vaccine were prepared by purification of HBsAg from human plasma, inactivation at 100 degrees C for 2 min and at 37 degrees C for 72 hours with formalin in a concentration of 1:4000. The former lot comprises purified HBsAg (40 mg/ml) adsorbed with 0.32 mg % Al(OH)3, the latter consists of purified HBsAg stabilized with 0.1% human albumin and adsorbed with 1 mg% Al(OH)3. The immune response after vaccination with the first lot was observed in 54.54% of the vaccinees. The second vaccine after 3 administrations of 80 micrograms/ml each produced a very good primary and a very good secondary immune response. Both vaccines are nonreactogenic and well tolerated by the vaccinees.

Strategies for the prevention of post-transfusion hepatitis.

Hepatitis is a common and potentially serious adverse effect of blood transfusion. A large number of strategies have been developed or proposed to reduce the incidence of post-transfusion hepatitis (PTH). The education of physicians regarding the risks of hemotherapy and the judicious use of blood must be the cornerstone of any substantial reduction in PTH. The use of volunteer rather than paid donors is associated with a much reduced incidence of PTH. Deferral of donors implicated in PTH is also helpful. Other proposed strategies include donor alanine amino-transferase levels, donor anti-HBc testing, the provision of immune globulin to recipients, and the inactivation, removal, or immune neutralization of the virus from blood products. In the absence of a blood substitute, autologous transfusion is an excellent means of improving transfusion safety. The incidence of PTH type B should decrease as an increasing proportion of donors and recipients are immunized by vaccine and as increasingly sensitive tests for HBsAg become available. The development of a serologic test and vaccine for non-A, non-B hepatitis would be outstanding accomplishments, but their absence underscores the need to pursue vigorously other means of reducing the incidence of the disease.

Prevalence of syphilis and hepatitis B among homosexual men in two saunas in Amsterdam.

In a pilot study by field-screening in two different sauna baths, a day-sauna and a night-sauna, 318 homosexual men were investigated for syphilis and hepatitis B surface antigen (HBsAg) and its antibody (anti-HBs). Of the 134 visitors to the day-sauna 35.1% were seroreactive for syphilis, four (3%) carried HBsAg, and 95 (70.9%) anti-HBs. Of the 184 visitors to the night-sauna 33.2% were seroreactive for syphilis, 16 (8.7%), carried HBsAg, and 97 (52.7%) anti-HBs. The correlation between seroreactivity for syphilis and the presence of HBsAg amd anti-HBs was statistically significant.