J.A. Madrigal

Analysis of the cytokine production by cord and adult blood

To date, over 400 human umbilical cord blood cord blood (CB) transplants have been reported from different centres world-wide and it is generally agreed that CB represents an encouraging alternative to bone marrow (BM) transplantation. There are a variety of reasons for this which include the wider availability and easier access of CB compared to BM. In addition it has been suggested that there is a reduced graft-versus-host-disease (GvHD) with CB compared to BM transplantation. The explanations for this implied benefit are numerous, but research into this area is only just beginning. Nevertheless, it is clear that both T cells and natural killer (NK) cells have reduced function when isolated from CB compared to adult and both these cell types have been implicated in GvHD pathogenesis. How and why the function is reduced is yet to be determined. Many laboratories have tried to answer these questions and the majority have done this by comparing the function of lymphocytes obtained from adult blood with those compared with CB. Since cytokine production by a cell is an indication of the cells function it is important to determine the differences between adult and CB with respect to production of these soluble factors. Here, we have reviewed the current research regarding these CB and adult cell comparisons with an emphasise on cytokine production. Our aim is to obtain a clearer understanding of the mechanisms which may be involved in causing a reduced GvHD in CB compared to BM transplantation.

Human leukocyte antigens influence the immune response to a pre-S/S hepatitis B vaccine

In this study we investigated the effects of a single pre-S/S (Hepagene) revaccination in a large population of multiple S vaccinated anti-HBs antibody nonresponder individuals (< 3 IU/l). We investigate the influence of vaccine dose (5, 10, 20 and 40 micrograms/ml), number of previous S containing vaccinations and the individuals HLA genotype on both B- and T-cell responses. We show that 76% of persistently nonresponder individuals produce anti-HBs antibody (> 3 IU/l) following a single revaccination with Hepagene. This anti-HBs antibody response was dose dependent. The group that received 5 micrograms/ml of Hepagene vaccine produced significantly less anti-HBs antibody than those receiving 10, 20 and 40 micrograms/ml doses (p < 0.05 in all cases). Individuals homozygous for HLA-DRB1*0701; DQB1*0202 failed to produce > 100 IU/l of anti-HBs antibody, whereas, heterozygous individuals required > 10 micrograms/ml Hepagene vaccine. The T-cell responses to Hepagene were exclusive of the dose and magnitude of anti-HBs antibody responses. There was a trend towards increased stimulation indices in those individuals who received repeated S containing vaccines. We have clearly shown that the immune response to Hepagene is influenced by the HLA genotype of the individual. However, further investigation is required to determine the specific role of these molecules in hepatitis B vaccine nonresponse. Hepagene is a registered trademark of Hedeva Pharma Ltd.

The influence of host factors and immunogenetics on lymphocyte responses to Hepagene® vaccination

We have shown that both demographic and immunogenetic factors are involved in the immune responses of Hepagene vaccinated individuals who were persistent nonresponders to S containing hepatitis B vaccines. The HLA-DRB1 0701; DQB1 0202 genotype was found to be associated with a decline of anti-HBs antibodies (anti-HBs) and were frequent in those individuals who remained nonresponders following booster vaccination. Contrary to previously published S vaccination data, Hepagene stimulated T-cell responses showed a lack of correlation with the humoral responses. Limiting dilution analysis demonstrated that the cellular immune response is associated with the kinetics of exposure to Hepagene rather than magnitude of the anti-HBs response. It remains that despite the inclusion of the pre-S proteins 74% nonresponder vaccinated individuals failed to produce > 100 IU/l of anti-HBs. However, these were persistent nonresponders and it was therefore encouraging that two doses of Hepagene did seroconvert (> 10 IU/L) 61% of this difficult group.

Hepatitis B third‐generation vaccines: improved response and conventional vaccine non‐response – evidence for genetic basis in humans

The lack of response to hepatitis B vaccination remains a problem for those individuals directly at risk of hepatitis B infection, particularly those who work in the health care industry. The factors associated with non‐response to hepatitis B vaccination have been investigated in 86 non‐responder health care workers who had received multiple ‘S’ vaccinations without sustained production of anti‐HBs. This group received a recently developed hepatitis B vaccine, HepageneTM, which included proteins derived from the envelope region of HBV, not present in currently licensed vaccines. The pre‐S1 and pre‐S2 proteins were included in HepageneTM in order to circumvent anti‐HBs non‐responsiveness which had previously been demonstrated in the inbred mouse model. The inclusion of these additional proteins in HepageneTM enabled some seroconverion, from non‐responder to responder; however, a proportion of the vaccinees remained non‐responders and the reasons for this have been investigated here, with reference to HLA alleles and the demographic predisposition. Here the mechanisms that underlie hepatitis B vaccine non‐response have considered the distribution of HLA alleles, age, sex, height and weight in addition to the T‐cell responses to HepageneTM derived antigens.

HLA-A typing by reference strand-mediated conformation analysis (RSCA) using a capillary-based semi-automated genetic analyser

HLA typing of class I loci by reference strand-mediated conformation analysis (RSCA) using a slab gel genetic analyser has been described. This study adapted the method for use in the capillary based ABI PRISM 310. Control DNA samples were used to create a database of mobility values for 37 HLA-A alleles. The technique was validated by comparing RSCA and sequence-specific oligonucleotide probe (SSOP)/sequence-specific primer amplification (SSP) HLA-A locus typing results from 214 cord blood samples. Of the samples tested, 6.5% required confirmatory typing by SSP, compared with a repeat rate of 10-40% for SSOP. In 200 samples where no SSP was necessary, there was 100% concordance between RSCA and previous results. The ABI PRISM 310 RSCA method defines HLA-A types at medium resolution and is quick and easy to implement.