Induction of peripheral blood T follicular helper cells expressing ICOS correlates with antibody response to hepatitis B vaccination

T follicular helper (TFH) cells, a critical subset of CD4⁺ T cells, provide help to B cells during the procession of the humoral immune response in the germinal center (GC) and extrafollicular sites. CXCR5⁺CD4⁺ T cells in human circulating blood, referred to herein as peripheral TFH (pTFH) cells, share phenotypes and functional properties with TFH cells in GC. Hepatitis B vaccine protects about 60% of the chronic hepatitis C patients from hepatitis B. The immunological bases that lead to the induction of protective antibody response is not well understood. In the present study, the pTFH cells subsets were determined in 18 healthy controls (anti-HBs ≥ 100 mIU/mL; HC), 21 nonresponders (anti-HBs < 10 mIU/mL; NR), and 23 weak responders (10 mIU/mL ≤ anti-HBs < 100 mIU/mL; WR) of chronic hepatitis patients upon routine hepatitis B vaccination. Though the frequency of the pTFH cell was equivalent in HC, WR, and NR, ICOS⁺pTFH cells in HC underwent expansion with increased IL-21 secretion and production of serum anti-HBs response at 4 weeks after a full course of hepatitis B vaccination. These changes were not shown in both NR and WR. Analysis of ICOS⁺pTFH cells represents a novel cellular determinant of the hepatitis B vaccine–induced humoral immune response, which may have relevance for design of hepatitis B vaccine.     

The spot-ELISA: a sensitive in vitro method to study the immune response to hepatitis B surface antigen.

We present our experience with a new sensitive in vitro method for the study of the immune response to hepatitis B surface antigen (HBsAg). This immunoenzymatic technique, called spot-ELISA, detects specific immunoglobulin production (e.g. IgG- or IgM-anti-HBs) by stimulated individual B cells in vitro. We have studied the immune response against HBsAg by mitogenic stimulation and subsequent spot-ELISA assay in 24 well-documented subjects with known immune status after either natural infection or vaccination with plasma-derived vaccine. Results indicate a good correlation between in vitro IgG anti-HBs spots and immune 'memory.' A possible predictive value in non-immunized and non-responder subjects awaits further confirmation.     

Absence of anti-hepatitis B surface antibody after vaccination does not necessarily mean absence of immune response

A small number of subjects vaccinated against hepatitis B do not produce anti-hepatitis B surface (HBs) antibody levels detectable by commercial assays. Others lose detectable anti-HBs at some time after vaccination. The absence of clinical hepatitis despite potential exposure to hepatitis B virus (HBV) in both kinds of subjects suggests that they might be protected by low antibody levels. However, besides anti-HBs, T helper response and memory cells which may be induced by the vaccine are certainly also important for immunity against HBV. In the present study, samples from vaccinated subjects, found to be anti-HBs negative in an initial assay, subsequently showed positive results in, respectively, 25%, 36% and 38% of the cases, when a second, third and fourth assay was used. In addition, 360 samples from “nonresponders” and from vaccinees who had lost anti-HBs, the reactivity of which was under the enzyme-linked immunoassay-cut-off value were compared to that of nonvaccinated controls. The absorbances were found to be significantly higher in the nonresponders (0.038) and in the vaccinees having lost anti-HBs (0.041), than in the controls (0.025). Such findings contribute to explaining why so-called nonresponders as well as vaccinees who have lost anti-HBs nevertheless appear to be protected. Received: 16 June 2000     

The immune response of healthy Nigerian adults to small doses of hepatitis B vaccine: comparison of 10- and 20-micrograms doses

The immunogenic effect of hepatitis B vaccine (H-B-vax) was evaluated in 120 seronegative healthy Nigerians. Three doses of the vaccine were given at 0, 1, and 6 months. Serial blood samples were tested 1 month after each vaccination for hepatitis B surface antibody (anti-HBs) and antibody to hepatitis B core antigen (anti-HBc). Of 60 vaccines given 20 micrograms of the vaccine, 40% had significant anti-HBs response 1 month after the first dose, 70% after the second dose, and 91.7% after the third dose. In the 60 vaccines given 10-micrograms doses, the seroconversion rates were 35, 73.3, and 90%, respectively. It is concluded that this vaccine in 10-micrograms doses is as effective as the larger doses in producing anti-HBs. The administration of small doses would reduce the cost of large-scale vaccination programs in developing countries.     

Active immunization of homosexual men using a recombinant hepatitis B vaccine

Twenty homosexual men [13 anti-human immunodeficiency virus (HIV)-positive, seven anti-HIV negative] without HBsAg, anti-HBs, and anti-HBc were vaccinated with three 20 micrograms doses of a recombinant hepatitis B vaccine. All anti-HIV-positive homosexuals were nonresponders independent of the initial number of CD4-positive cells. Among seven anti-HIV-negative individuals, five responded. After three doses of the vaccine, CD4-positive cells fell in anti-HIV positive individuals by 22.4%. A similar fall in CD4-positive cells of an average 24.9% was noted in 17 matching, but nonvaccinated, anti-HIV-positive homosexuals. The study indicates that the efficacy of vaccination in anti-HIV-positive individuals is questionable. There is, however, no evidence that vaccination against hepatitis B might be harmful to anti-HIV-positive subjects.     

Immunogenicity in chimpanzees of experimental hepatitis B vaccines prepared from intact hepatitis B virus,purified polypeptides,or polypeptide micelles

The immunogenicity of three experimental hepatitis B vaccines was evaluated in chimpanzees. Although no antibody to hepatitis B surface antigen (anti-HBs) was detected in two chimpanzees that received an aqueous polypeptide vaccine subcutaneously, a strong anti-HBs response was observed two and ten weeks, respectively, following challenge with hepatitis B virus. Inoculation of two additional chimpanzees with a micellar preparation of these polypeptides by the intravenous route resulted in anti-HBs production in one of the chimpanzees. Two chimpanzees inoculated subcutaneously with an aqueous vaccine of formalin-inactivated intact hepatitis B virus developed anti-HBs in low titers, but the development of antibody to the hepatitis B core antigen following challenge inoculations suggested that subclinical HBV infections may have occurred despite prior vaccination.     

Immune response to hepatitis B revaccination

Two hundred and twelve individuals who failed to respond or responded poorly to hepatitis B vaccination or whose anti-HBs levels had dropped below 10 IU/L at follow-up were revaccinated. Only 18% of initial nonresponders, but 96% of those who responded initially, developed anti-HBs above 10 IU/L after revaccination. In 85% of vaccinees, anti-HBs titres after a fourth immunization were significantly higher than after completion of the first full immunization course, and despite a steeper decline of antibody levels after revaccination, anti-HBs seemed to persist better than after the initial course of three inoculations. All individuals who responded initially and in whom anti-HBs had become undetectable developed antibodies. Response to booster doses was seen after three to five days, and a fifth inoculation given to 23 individuals induced even better responses.     

Detection of hepatitis B surface antigen (HBsAg) in peripheral blood mononuclear cells of patients with acute and chronic active hepatitis B

Seventy five patients with acute and chronic active hepatitis (CAH) were studied by indirect immunofluorescence with monoclonal antibodies for the presence of hepatitis B surface antigen (HBsAg) on peripheral blood mononuclear cells (PBMC). The viral surface antigen was detected in the PBMC of all the patients with hepatitis B virus (HBV)-induced CAH and in acute patients with more than 2 months of evolution. No HBsAg was detected in the samples obtained from 12 normal controls or from 14 non-A, non-B CAH patients. Analysis of PBMC subsets revealed that HBsAg was present in non-T cells; dual fluorescence studies showed HBsAg on surface Ig-positive lymphocytes. The binding of anti-HBs monoclonal antibodies was higher than that of a goat anti-HBs serum, and the highest reactivity was observed with an antibody against the pre-S(2)-region sequence. Both HBsAg and hepatitis B core antigen (HBcAg) were also detected in lysates of PBMC by dot blot analysis.     

High rate of seroconversion following administration of a single supplementary dose of recombinant hepatitis B vaccine in Iranian healthy nonresponder neonates

Forty-nine Iranian neonates who failed to develop a protective anti hepatitis B surface antigen (HBs) response following primary vaccination with triple doses of recombinant hepatitis B vaccine, were classified as hypo-responders (anti-HBs > 1 < 10 IU/l) or non-responders (anti-HBs < 1 IU/l) and subsequently challenged with a single supplementary vaccine dose. A protective and anamnestic type of response was observed in 90% (44/49) of the neonates. The mean titer of anti-HBs antibody was significantly higher following supplementary vaccination, compared to that achieved after primary vaccination. This was more evident in the primary hypo-responder neonates (P < 0.00001) than the non-responder group (P < 0.002). The results indicate that a significant proportion of the non-responder neonates to HBs antigen can be induced to develop a protective and long-lasting antibody response by administration of a single additional vaccine dose. Received: 24 September 1996     

慢性乙型肝炎患者B细胞功能的研究

对24例乙型慢迁肝患者进行体外B细胞功能研究.结果显示慢性迁延性肝炎(慢迁肝)患者外周血单个核细胞自发产生或经美洲商陆(PWM)刺激后产生的IgG、IgM量与正常人相比无显著差异.乙型慢活肝患者自发产生的IgG、IgM量比正常人明显升高,但经PWM刺激后,IgG、IgM的产生量反而比对照组明显减低.慢活肝患者血清IgG量与B细胞体外培养产生的IgG量有较明显的相关性.     

German experimental hepatitis B vaccine — Influence of variation of dosage schedule,sex and age differences on immunogenicity in health care workers

Summary Of the medical staff of our hospital 217 members at high risk for hepatitis B were immunized with an experimental hepatitis B vaccine and anti-HBs titers used to study the influence of two dosage schedules, age, and sex on immunogenicity. Participants were 34 years of age (mean; range, 20–61); they were divided into two groups and vaccinated three times. Group A received 42 µg HBsAg for each vaccination. Group B received 84 µg for the first and 21 µg for the second and third vaccinations. The seroconversion rate was 32.7% after the first, 78.8% after the second, and 95.7% after the third vaccination. The participants who failed to produce anti-HBs titer (3 IU/l;n=9) or whose anti-HBs titers were below 50 IU/l (n=31) were vaccinated a fourth time. Only mild side effects of injections were observed in a third of all participants, usually in the form of a sore arm.Between groups A and B there were no significant differences as far as the seroconversion rate and anti-HBs titer were concerned. Nonresponders plus low-responders accounted for 19%. Female participants produced a markedly higher anti-HBs titer than males, and the female/male ratio among non- and low-responders was 1:2; among nonresponders, 1:2.5. There was a negative correlation of the anti-HBs titer with the age of the participants. These results not only have practical consequences for revaccination policy, but also offer the opportunity to further study the genetic regulation of the immune response to a complex peptide antigen in man.Abbreviations anti-HBc antibody to hepatitis B core antigen - anti-HBe antibody to hepatitis B e antigen - anti-HBs antibody to hepatitis B surface antigen - HBsAg hepatitis B surface antigen - IU/l international units per liter - MSD Merck, Sharp, and Dohme     

Dissociated antibody responses to the S and pre-S2 regions of the hepatitis B virus after vaccination in hemophiliacs

The membranes of hepatocytes and the pre-S2 envelope protein of the hepatitis B virus (HBV) contain binding sites for polymerized human albumin, which is thought to act as a link between HBV and hepatocytes. Hence, anti-pre-S2 antibodies should prevent HBV uptake by the liver, and there is indeed preliminary evidence that they protect chimpanzees from HBV infection. To evaluate whether a plasma-derived vaccine containing the pre-S2 sequence induced an anti-pre-S2 response in 105 vaccinated hemophiliacs, anti-pre-S2 was measured in parallel with antibody to hepatitis B surface antigen (anti-HBs). Eighty-five percent of the hemophiliacs had both anti-pre-S2 and anti-HBs when vaccination was completed, 13% had anti-HBs alone, and 2% (two cases) had anti-pre-S2 alone. Eighty-seven percent of anti-pre-S2-positive hemophiliacs compared with only 50% of anti-pre-S2-negative hemophiliacs (P less than 0.001) developed high anti-HBs titers (greater than or equal to 1,000 mlU/ml). This study demonstrates, therefore, that the antibody responses to the S and pre-S2 regions of HBV may be dissociated after vaccination in hemophiliacs and that higher anti-HBs titers are attained in anti-pre-S2-positive hemophiliacs.     

Immunoregulation of the in vitro anti-HBs antibody synthesis in chronic HBsAg carriers and in recently boosted anti-hepatitis B vaccine recipients.

We report a study on immunoregulation of in vitro antibody to hepatitis B surface antigen (anti-HBs) synthesis induced by pokeweed mitogen (PWM) from peripheral blood mononuclear cells (PBMC) in chronic hepatitis B surface antigen (HBsAg) carriers and in 'high responders', (anti-HBs RIA ratio greater than or equal to 20 in serum), recently boosted with anti-hepatitis B vaccine. Anti-HBs was detected in 11 days PBMC supernatants (SN) from 24 out of 36 'high responders', but in none from 31 chronic HBsAg carriers, despite detectable amounts of polyclonal IgG and antibody to hepatitis B core antigen (anti-HBc) were produced. The lack of anti-HBs production by chronic HBsAg carriers did not seem to be determined by suppressor influences because T lymphocytes from the majority of chronic HBsAg carriers, co-cultured with 'high responders' PBMC did not suppress anti-HBs production. Co-cultures between HBsAg carriers T4 positive (helper/inducer) cells and allogenic 'high responder' non-T cells produced anti-HBs antibody, indicating that HBsAg carrier T cells are not deficient in this allogenic helper function under PWM stimulation. Allogenic cocultures between HBsAg carrier non-T cells and 'high responder' T4 positive cells failed in anti-HBs production: a specific B lymphocyte defect might be involved in the lacking anti-HBs synthesis in chronic HBV patients. Antigen-induced specific anti-HBs synthesis experiments indicate that B cells themselves seem to be the target for HBsAg-induced suppression of anti-HBs antibody response.     

Acute hepatitis B viral infection in a patient with anti-HBs.

We report a patient with antibody to hepatitis B surface antigen (anti-HBs) but no antibodies to other hepatitis B virus components, who developed acute symptomatic type B hepatitis. The possible explanations for this unusual serological pattern are 1) the antibody-positive status, which developed against only a subdeterminant of hepatitis B surface antigen (HBsAg), arose naturally or as the result of cross-reaction with a variety of antigens; and 2) seroconversion to anti-HBs occurred in response to surface antigen of a mutant strain of hepatitis B virus (HBV). This anti-HBs positivity, in the absence of antibody to hepatitis B core antigen, does not provide natural immunization against HBV infection, and so is not protective. Individuals who are positive to anti-HBs antibody alone which is not elicited by HBV vaccine, should be vaccinated against possible HBV infection.     

Correlation between HBV DNA detection by polymerase chain reaction and Pre-S1 antigenemia in symptomatic and asymptomatic hepatitis B virus infections

The presence of hepatitis B virus (HBV) genome in sera from 73 symptomatic and asymptomatic HBsAg carriers was studied by the polymerase chain reaction (PCR) with primers specific for the S and C regions. Pre-S proteins of the HBV envelope were detected in serum by a specific monoclonal antibody in a double immunoradiometric assay. Out of twenty-five symptomatic patients with chronic active hepatitis (14 with HBeAg and 11 with anti-HBe), all were positive for HBV DNA by PCR, while 14/14 HBeAg and 2/11 (18%) of the anti-HBe patients were positive by dot blot hybridization. All but one anti-HBe patient (96%) carried Pre-S1 proteins. Among the asymptomatic HBsAg carriers, HBV DNA was detected by PCR in 14/14 (100%) HBeAg positive patients and in 25/34 (73%) anti-HBe positive patients. Pre-S1 proteins were found, respectively, in 14/14 (100%) and 11/22 (50%) of the same cases tested in parallel. The 20 healthy blood donors devoid of HBV markers and with normal transaminases tested were found negative for HBV DNA using PCR. Out of 12 patients who recovered from acute hepatitis B, all were found negative by PCR analysis after a mean follow up of 1 year after seroconversion to anti-HBs. When serial samples from 2 patients (one with acute hepatitis B, the other with chronic hepatitis B) were tested for the presence of HBV DNA and of Pre-S1 proteins, both markers showed parallel development.(ABSTRACT TRUNCATED AT 250 WORDS)     

Serologic markers of viral hepatitis A,B, C,and D in patients with hemophilia

Forty-one patients with hemophilia A were studied for the prevalence of serological markers for hepatitis A, hepatitis B, hepatitis C (non-A and non-B hepatitis), and delta hepatitis (hepatitis D). Ten of 41 (24.4%) patients demonstrated hepatitis A antibody and 31 of 41 (75.6%) patients had a serologic marker for previous hepatitis B infection; four of these 31 patients (13%) also demonstrated antibody to delta agent (hepatitis D). Thirty-seven of 41 (90.2%) patients demonstrated antibody for hepatitis C. Nine of 31 (29%) patients with a hepatitis B marker (no hepatitis B vaccinees) were negative for anti-HBc but positive for anti-HBs; all of these nine patients were HIV antibody positive, although they had no overt immunodeficiency. Twenty-six of 41 (63.5%) patients were HIV antibody positive. Of HIV antibody positive patients, 27%, 88%, and 100% demonstrated evidence of a previous hepatitis A, hepatitis B, or hepatitis C, respectively. Of HIV antibody negative patients; 20%, 53%, and 73% of the patients demonstrated evidence of a previous hepatitis A, hepatitis B, or hepatitis C infections, respectively. The difference between HIV antibody positive and HIV antibody negative groups was not significant for hepatitis A but was significant for hepatitis B (P < 0.001) and hepatitis C (P < .001). Of the 31 patients with a hepatitis B serologic marker, all had antibody to hepatitis C. Of 10 patients, without a hepatitis B serologic marker, only 6 (60%) had antibody to hepatitis C. Clinically, none of our patients demonstrated any evidence of liver disease; however, 10 patients reported a previous history of hepatitis. These data suggest that patients with hemophilia in the United States, particularly those with HIV antibody, have high prevalence of hepatitis B and C infections.     

大剂量HBsAg疫苗对小鼠细胞免疫应答的调节作用

目的:探讨大剂量HBsAg疫苗对机体细胞免疫应答的影响。方法:不同剂量HBsAg疫苗两次免疫小鼠后, 用氚-胸腺嘧啶核苷(-[H³]TdR)掺入法, ELISA方法分别测定免疫鼠T淋巴细胞增殖能力, 细胞培养上清中白细胞介素-2(IL-2), γ干扰素(IFN-γ)含量及血清中抗HBsIgG2a阳转率。结果:大剂量HBsAg疫苗单次免疫后, 小鼠T淋巴细胞增殖能力显著强于对照组(P＜0.05);Th1型细胞因子IL-2、IFN-γ显著多于对照组(P<0.05);抗HBsIgG2a阳转率显著高于小剂量组(P＜0.01)。加强免疫后, 各项指标增高更显著, 小剂量组才呈现显著性增高。结论:大剂量HBsAg疫苗可诱生特异性细胞免疫应答, 并使细胞免疫趋向Th1型。     

Co-occurrence of HBsAg and anti-HBs: Two consecutive infections or a sign of advanced chronic liver disease?

Simultaneous presence of hepatitis B surface antigen (HBsAg) and antibodies to the surface antigen (anti-HBs) was detected in 32 out of 89 Dutch chronic hepatitis patients of Caucasian race. HBsAg was subtyped ad in 28 and ay in four cases. Anti-HBs could be subtyped in 25 cases using reference antigens discriminating between d, y, and w1-4 subdeterminants. In 20 patients HBsAg subtype ad (HBsAg/ ad) was accompanied by antibody to subdeterminant y (anti-y), whereas HBsAg/ ay and anti-d were simultaneously detected in the serum of one patient. The antibody pattern in sera from the remaining patients was complex. Eighteen anti-HBs-positive patients were matched for age, histology, and hepatitis B e antigen (HBeAg) status with 18 anti-HBs-negative patients. Differences in risk factors for acquiring a hepatitis B infection were not found. These results do not support the hypothesis that co-occurrence of HBsAg and anti-HBs is due to two consecutive infections with hepatitis B virus. The frequency of the co-occurrence of HBsAg and anti-HBs was found to be related to the degree of progressive liver disease, since anti-HBs was found in three out of 23 asymptomatic carriers, in four out of 20 chronic persistent hepatitis patients, in 20 out of 41 chronic active hepatitis patients, and in all five patients with chronic active hepatitis and cirrhosis. The high frequency of anti-HBs in advanced liver disease may be the result of a disturbed immunologic response mechanism.     

Modelling long-term persistence of hepatitis B antibodies after vaccination

Antibody response to hepatitis B vaccination was measured in 97 institutionalized, mentally handicapped patients after a 0-, 1-, 6-month vaccination schedule. Two groups were formed at month 7 according to the antibody response, which determined whether they needed an additional vaccine dose at month 12, to achieve an antibody titre of 100 IU/liter. All residents were followed up yearly for the first 5 years, after which, they received another booster dose. Another blood sample was taken 11 years after the start of the program. A linear mixed-regression model was used to analyze the data. Random and fixed effects were included to determine the generally known risk factors and the still unknown individual characteristics that influence the titre of hepatitis B surface antibodies (anti-HBs). The mean anti-HBs titre was a function of time, type of mental retardation (Down's syndrome or other types of mental retardation), the use of antiepileptic drugs, and the additional booster at month 12. The immediate and vigorous response of the immune system to booster vaccination shows that the immunologic memory is good after primary vaccination. For the maintenance of protection, the recommendation for mentally retarded patients in institutions is vaccination of all seronegative residents as well as new entrants, after which, no additional boosters will be necessary. J. Med. Virol. 57:100–103, 1999. © 1999 Wiley-Liss, Inc.     

Low-dose vaccination against hepatitis B in children: one-year follow-up

Six hundred forty-three children, negative for markers of hepatitis B virus (HBV) infections, were given three X 2-micrograms doses of Merck, Sharp and Dohme (MSD) plasma derived hepatitis B vaccine (H-B-Vax) at monthly intervals. Twelve months after the first dose of vaccine, antibody to hepatitis B surface antigen (anti-HBs) was detected in 89% of children by radioimmunoassay (RIA) and in 83% by enzyme immunoassay (EIA). Seroconversion rates and anti-HBs titres were significantly greater in 1-4-year-olds than in older children (p less than 0.01). Eighteen children with no anti-HBs or other markers of HBV at this time were given 10 micrograms of vaccine and tested one month later. Seventeen developed anti-HBs, 12 at levels consistent with an anamnestic response. Forty-nine HBV-marker-negative children seroconverted for antibody to hepatitis B core antigen (anti-HBc) in the 8-month period before or the 12-month period following vaccination. Forty-six of these children were positive for anti-HBs, and one has been confirmed as a chronic carrier of hepatitis B surface antigen (HBsAg). Three cases of clinical hepatitis B in children have been seen in the community since the vaccination programme began. Two of these were amongst the estimated 5% of children who were not vaccinated. The third was in a vaccinee and occurred 4 1/2 months after the last dose of vaccine.(ABSTRACT TRUNCATED AT 250 WORDS)