Comparison of HLA-DR1-restricted T cell response induced in HLA-DR1 transgenic mice deficient for murine MHC class II and HLA-DR1 transgenic mice expressing endogenous murine MHC class II molecules

Anthony Pajot1, Véronique Pancré4, Nicolas Fazilleau2, Marie-Louise Michel3, Gerhild Angyalosi4, David M. Ojcius5, Claude Auriault4, François A. Lemonnier1 and Yu-Chun Lone1

1 Unité d'Immunité Cellulaire Antivirale, 2 Unité Biologie Moleculaire du Gène and 3 Unité de carcinogenese hépatique et Virologie Moléculaire, INSERM U370, Institut Pasteur, Paris, France
4 Laboratoire d'Immunopathologie Cellulaire des maladies infectieuses, CNRS l'UMR 8527 de l'Institut de Biologie, Lille, France
5 School of Natural Sciences, University of California, Merced, CA 95344, USA

Correspondence to: Y.-C. Lone; E-mail: lone{at}pasteur.fr


    Abstract
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 Abstract
 Introduction
 Methods
 Results
 Discussion
 References
 
Transgenic mice expressing human HLA class II molecules provide a useful model for identifying HLA-restricted CD4+ epitopes. However, the influence of endogenous murine H-2-restricted T cell responses on HLA-restricted responses is not known. In the present study, we show that HLA-DR1 transgenic mice deficient for H-2 class II expression (HLA-DR1+/+/IAß0/0) exhibit an equivalent expression level of the transgene HLA-DR1 and a similar diversity in the TCR repertoire, but a slightly different number of CD4+ peripheral T cells, when compared to HLA-DR1 transgenic mice in which H-2 class II molecules were retained (HLA-DR1+/+/IAß+/+). More importantly, a strong antigen-specific HLA-DR1-restricted response was observed in nearly all HLA-DR1+/+/IAß0/0 mice immunized with HBV envelope protein (HBs) or capsid protein (HBc), whereas weak HBs- or HBc-specific HLA-DR1-restricted responses were detected in half of the immunized HLA-DR1+/+/IAß+/+ mice. Conversely, strong HBs- or HBc-specific H-2-restricted T cell responses were detected in HLA-DR1+/+/IAß+/+ mice but not in HLA-DR1+/+/IAß0/0 mice. Our results indicate that the coexpression of endogenous H-2 class II molecules reduces the intensity of HLA-DR1-restricted antigen-specific responses in transgenic mice, by favoring murine over human MHC recognition and education. Thus, HLA-DR1+/+/IAß0/0 mice represent a better model for identifying and characterizing HLA-DR1-restricted epitopes relevant for human disease.

Keywords: antigen, major histocompatibility complex, transgenic mice, vaccine


    Introduction
 Top
 Abstract
 Introduction
 Methods
 Results
 Discussion
 References
 
The development of protective T cell immunity against infectious pathogens involves major histocompatibility complex (MHC) class I-restricted cytotoxic CD8+ T cells (CTL) and MHC class II-restricted helper CD4+ T cells. Antigen-specific CD8+ T cell immunity primarily involves direct elimination of antigen-expressing cells (13) while antigen-specific CD4+ T cells promote optimal CD8+ T cell (4,5) and B cell activation (68).

The T cell-mediated response to complex antigens involves recognition of selected peptide epitopes presented in the context of MHC molecules expressed on antigen-presenting cells (9). The identification of immunogenic epitopes among the hundreds or thousands of potential epitopes present in an antigenic protein is thus a critical step in attempts to optimize T cell-mediated immunity, and is important for the development of efficient vaccines. However, the identification of specific epitopes from complex antigens can be a cumbersome and difficult process, due to the multiplicity of MHC molecules encoded by distinct gene loci in both mice and humans. Moreover, using T cell clones isolated from whole antigen-stimulated cells it's not obvious to identify the specificity of the T cell epitopes and their restriction alleles. To address this problem, we and others have developed transgenic (Tg) mice expressing human leukocyte antigen (HLA) class I complexes to facilitate identification of immunogenic HLA class I-restricted CTL epitopes (10,11). This experimental animal model is also useful for in vivo studies of cellular immune responses to various candidate vaccines, and it represents an attractive vertebrate system with immunological relevance for the human immune system. However, currently available mice often fail to develop responses against CTL epitopes known to elicit HLA-class I restricted responses in humans. To increase the abundance of murine CD8+ T cells restricted by human HLA class I molecules, it is essential to eliminate endogenous expression of murine H-2 class I molecules in such a way that only the Tg HLA alleles act as restriction elements in the mice (1214). A number of novel CTL epitopes for various antigens have already been identified with these mice, and synthetic peptides representing these epitopes have been used to prepare therapeutic vaccines to treat cancer patients. But the therapeutic effects of these vaccines are still far from optimal. One of the reasons may lie in the absence of concomitant tumor-specific CD4+ T cell responses. Indeed, recent studies have underlined the importance of CD4+ helper T (TH) lymphocytes in both the initiation and maintenance of CD8+ immune responses (1518). Despite the accepted requirement for antigen-specific CD4+ T cells, only a limited number of helper T cell epitopes (TH epitopes) restricted by specific human alleles have been identified until now. We have therefore focused our efforts on developing an HLA-class II Tg animal model to facilitate identification of new immunogenic TH-epitopes. In the present study, we analyzed the HLA-DR1-restricted T cell responses elicited in HLA-DR1 Tg mice deficient for H-2 class II expression (HLA-DR1+/+/IAß0/0), compared to those induced in ‘classic’ HLA-DR1+/+/IAß+/+ Tg mice. Our results demonstrate that there is competition for immunodominance between H-2 class II- and HLA class II-restricted T cell responses, and suggest that the presence of endogenous H-2 class II molecules may introduce a bias in favor of H-2 class II-restricted over HLA-class II-restricted T cell responses in HLA-DR1+/+/IAß+/+mice. The data presented in this report are consistent with HLA-DR1+/+/IAß0/0 mice, providing a more sensitive and specific model for identifying and characterizing HLA-DR1-restricted epitopes for a variety of human disease-associated Ags.


    Methods
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 Abstract
 Introduction
 Methods
 Results
 Discussion
 References
 
Mice
The mice used in this study were bred and maintained in pathogen free conditions in the animal facilities of the Pasteur Institute, Paris. The HLA-DR1 IAß0/0 transgenic mice were obtained at the Pasteur Institute of Lille by back-crossing the original HLA-DR1 transgenic mice (19) with Aß0/0 mice (B6-IAß0/0 mice) (20).

Peptides
The HLA-DR1 binding peptides, HBs179–194 QAGFFLLTRILTIPQS, HBc111–125 GRETVIEYLVSFGVW, HIV-1 Gag263–277 KRWIILGLNKIVRMY, the H-2 IAb binding peptides HBs126–138 RGLYFPAGGSSSG and HBc128–140 TPPAYRPPNAPIL were synthesized by Primm (Milano, Italy) and dissolved in PBS–10% DMSO at a concentration of 1 mg/ml.

FACS analysis of peripheral B and T lymphocytes
B lymphocytes were purified on a mini-MACS column (Miltenyi Biotec, Bergish Gladbach, Germany) by positive selection. The expression of HLA-DR1 and H-2 IAb molecules was analyzed by immunofluorescence using FITC-labeled anti-HLA-DR (L243) and PE-labeled anti-IAß (AF6-120.1) (BD Biosciences, San Diego, CA). The percentage of single CD4+ and CD8+ T lymphocytes was determined by double staining, using PE-labeled anti-mouse CD4 and FITC-labeled anti-mouse CD8 (Caltag, South San Francisco, CA). The expression of the different BV TCR was analyzed using PE-labeled anti-mouse CD4 and the following biotinylated specific antibodies: BV 2 (B20.6), BV 3 (KJ25), BV 4 (KT10.4), BV 5.1.2 (MR.9.4), BV 6 (44.22), BV 7 (TR 130), BV 8.1.2.3 (F23.1), BV 9 (MR.10.2), BV 10 (B.21.5), BV 11 (RR.3.15), BV 12 (MR11.1), BV 13 (MR.12.4), BV 14 (1412) and BV 17 (KJ 23.1). Cells were incubated for 30 min on ice with the first antibody and, following two washes, incubated with the second antibody (4°C, 30 min), washed twice, fixed with paraformaldehyde and analyzed by a FACSCalibur (Becton Dickinson, Bedford, MA).

Immunoscope analysis
The TCR repertoire of CD4+ T cells from naive transgenic mice was analysed for BV usage using oligonucleotides specific for each family of BV segments, as described elswhere (21). In brief, spleen CD4+ T cells were positively selected on auto MACS (Miltenyi Biotec, Bergish Gladbach, Germany). RNA was prepared using the RNA easy Kit (Qiagen, Hilden, Germany) and reverse transcribed into cDNA using oligo (dT) and SuperScript II (Invitrogen, Carlsbad, CA). Half of the reaction product was then amplified by 40 cycles of PCR (annealing at 60°C) in 25 µl, in the presence of BV-specific primers and subjected to an elongation reaction (run-off) with internal BC or BJ fluorescent-tagged primers. The labeled products were then loaded on a 6% acrylamide/8 M urea gel and separated by a run of 7 h at 35 W on a 373 DNA sequencer (Applied Biosystems). Data were analyzed using immunoscope software (21) designed for this purpose.

Immunization with HBc and HBs antigen
The HBc antigen used in this study was produced in Escherichia coli and purchased at Diasorin (Baluggia, Italia). The recombinant preS2/HBs antigen (ayw HBV subtype) was derived from transfected mammalian cells (CHO) (22). For HBc immunization (4 µg of HBc in 50 µl alum adjuvant; Serva, Heidelberg, Germany), mice were injected twice at the base of the tail. Similarly, for HBs immunization, 2 µg of HBs was injected and the mice were also immunized twice. Before immunization, all mice were anesthetized intraperitoneally (i.p.) with 75 mg/kg of pentobarbital (Ceva, Santé Animale, Libourne, France).

T cell proliferation assay
Twelve days after immunization, the spleen was removed from animals and placed in RPMI serum-free medium. Splenocytes were RBC-depleted and submitted to a Ficoll gradient, then cultured in RPMI supplemented with 10% FCS, 10 mM HEPES, 1 mM sodium pyruvate, 5 x 10–5 M 2-mercaptoethanol. The spleen suspension was then adjusted to 10 x 106 cells/ml (5 x 105 cells/well) and incubated with 20 µg/ml, 6 µg/ml or 2 µg/ml of peptide. The cell suspension was incubated in 3% FCS in complete medium for 72 h at 37°C in 5% CO2. One µCi of [3H]thymidine was added to each well 16 h before the cells were harvested on a TOMTEC collector and the incorporated radioactivity was measured on a micro beta counter (Perkin Elmer, Courtaboeuf, France). Results are shown as SI (stimulation index) = (c.p.m. peptide)/(c.p.m. medium).

IL4 secretion assay
Twelve days after the last immunization with HBc Ag, the spleen suspension was stimulated with 20 µg/ml of HBc111–125 or irrelevant HIV-1 Gag263–277 peptide. After 4 h of incubation, IL4 secretion was determined by use of a cytokine secretion capture assay, following the protocol supplied by the manufacturer (Miltenyi Biotec, Bergish Gladbach, Germany) and adapted from Brosterhus et al. (23). Cells were then labeled with IL4 catch antibody and PE-detection antibody and counterstained with FITC-labelled CD4 antibody. After additional labeling with anti-FITC microbeads, the CD4+ cells were sorted and analyzed by FACS for IL4 secretion.


    Results
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 Abstract
 Introduction
 Methods
 Results
 Discussion
 References
 
Expression of HLA-DR1 molecules and size of the peripheral CD4+ T cell pool in HLA-DR1+/+/IAß0/0 and HLA-DR1+/+/IAß+/+ Tg mice
The cell surface expression of HLA-DR1 and H-2-IAb molecules, and the percentage of peripheral CD4+ T cells, was evaluated by flow cytometry analysis on B cell (B220+)-enriched and T cell-enriched splenocytes. As illustrated in Fig. 1(A), a similar level of HLA-DR1 expression was observed in HLA-DR1+/+/IAß0/0 and HLA-DR1+/+/IAß+/+ Tg mice, but not in non-Tg B6-IAß0/0 mice. As expected, a high level of H-2-IAb expression is observed exclusively in HLA-DR1+/+/IAß+/+ transgenic mice. CD4+ T cells represent 0.9% of the splenic cell population in B6-IAß0/0 mice as reported by Cosgrove et al. (20), 17% in HLA-DR1+/+/IAß+/+ and 14% in HLA-DR1+/+/IAß0/0 Tg mice (Fig. 1B).



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Fig. 1. (A) Surface expression of transgenic DR1 and endogenous IAb molecules on purified B220+ B cells from B6, DR1+/+ IAß0/0 and DR1+/+ IAß+/+ mice. B cells were stained with L243 (anti DR)–FITC or AF61201 (anti-IAß)–PE antibody and analyzed by flow cytometry. (B) Restitution of the peripheral pool of CD4+ T lymphocytes in DR1+/+ IAß0/0 mice. Double staining was performed with PE labeled anti-CD4 (y-axis, log scale) and FITC-labeled anti-CD8.

 
Peripheral CD4+ T cells in HLA-DR1+/+/IAß0/0 and HLA-DR1+/+/IAß+/+ Tg mice display similar TCR BV chain usage and CDR3 diversity
Expression of the various BV families was studied on purified CD4+ T cells by flow cytometry analysis using each of the 14 BV specific mAb. The percentage of CD4+ T cells expressing a given TCR BV is shown in Fig. 2(A) for HLA-DR1+/+/IAß0/0, HLA-DR1+/+/IAß+/+ Tg mice and wild-type mice. Although some differences were observed, they were not significant. Thus, all BV families and subfamilies are represented similarly in the three strains of mice. The CDR3 length distribution for each BV chain was also analyzed, using the RT–PCR-based immunoscope assay, as described in the Methods. Again, no significant differences between the three strains of mice were detected (Fig. 2B).



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Fig. 2. BV family usage in the CD4+ T cell peripheral population in DR1 IAß0/0, DR1 IAß+/+ and B6 mice. (A) Immunofluorescence analysis. Double staining was performed on non-activated purified T splenocytes with FITC-labeled BV specific and PE-labeled CD4 mAb. (B–D) Immunoscopic analysis of CDR3ß domain size distribution among CD4+ T cells from DR1 IAß0/0 (B), DR1 IAß+/+ (C) and B6 mice (D). Mouse splenic CD4 T cells were positively purified on an AutoMACS and processed as described in the Methods by immunoscope analysis with a set of 24 primers, including BV 5.3, BV 17 and BV 19, which correspond to a pseudogene in the B6 mice.

 
While numerous HLA class I-restricted CTL epitopes from various antigens are known, only a limited number of TH epitopes restricted by HLA-DR1 have been reported to date. In order to compare the two types of HLA-DR1-expressing mice, we thus took advantage of the fact that HLA-DR1- and H-2-IA-restricted epitopes have been identified for the hepatitis virus capsid (HBc) and envelope (HBs) proteins, by following the T cell responses induced by immunization with these antigens in HLA-DR1+/+/IAß0/0 mice and comparing them with those elicited in HLA-DR1+/+/IAß+/+ mice.

Comparative analysis of HBc- and HBs-specific CD4+ T cell responses in HLA-DR1+/+/IAß0/0 Tg mice, compared with HLA-DR1+/+/IAß+/+ Tg mice
HLA-DR1+/+/IAß0/0 and HLA-DR1+/+/IAß+/+ Tg mice were immunized subcutaneously with HBc or HBs antigen, as described in the Methods. The splenocytes derived from the primed mice were then stimulated in vitro with HLA-DR1-restricted peptides (HBc111–125, HBs179–194 or Gag263–277) or with H-2-IA-restricted epitopes (HBc128–140 or HBs126–138). In the HLA-DR1+/+/IAß0/0 Tg mice, a stronger proliferative response directed against the HLA-DR1-restricted peptides (HBc111–125, Fig. 3A; or HBs179–194, Fig. 4A) was observed, while the H-2-IA-restricted peptides were inefficient (Figs 3A and 4A). In contrast, in the HLA-DR1+/+/IAß+/+ Tg mice, the H-2-IA-restricted anti-HBc or anti-HBs responses were predominant (Figs 3B and 4B), and an additional in vitro recall with the HBc111–125 (Fig. 3C) or HBs179–194 (data not shown) peptides was necessary to detect a weak HLA-DR1-restricted response. As expected, no responses were induced by the Gag263–277 peptide in either type of Tg mouse. Thus, expression of the HLA-DR1 Tg molecule in the context of endogenous IA molecules diminishes HLA-DR1-restricted Ag-specific T cell proliferative responses.



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Fig. 3. (A and B) T cell proliferation to HBc-derived peptides in HLA-DR1+/+ IAß0/0 and HLA-DR1+/+ IAß+/+ mice. Mice were immunized twice with 4 µg of HBc protein in alum at the base of the tail. Splenocytes from primed transgenic mice were rechallenged in vitro with 20 µg/ml (grey histogram), 6 µg/ml (dark histogram) or 2 µg/ml (white histogram) of each indicated peptide. (C) Splenocytes from immunized HLA-DR1+/+ IAß+/+ mice were subjected to an additional in vitro stimulation before the test. Control samples were cultured in medium alone or with an irrelevant peptide, HIV-1 Gag263–277. Proliferation was determined by [3H]thymidine incorporation and represented as the SI means of duplicate experiments.

 


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Fig. 4. (A and B) T cell proliferation to HBs antigenic peptides in HLA-DR1+/+ IAß0/0 and HLA-DR1+/+ IAß+/+ mice. Mice were immunized twice with 2 µg of HBs protein in alum at the base of the tail. Splenocytes from primed transgenic mice were rechallenged in vitro with 20 µg/ml (grey histogram), 6 µg/ml (dark histogram) or 2 µg/ml (white histogram) of each peptide. Control samples were cultured in medium alone or with an irrelevant peptide. Proliferation was determined by [3H]thymidine incorporation and represented as the SI means of duplicate cultures of two representative experiments.

 
Frequency of anti-HBc-specific HLA-DR1-restricted CD4+ T cells
The number of HLA-DR1-restricted HBc111–125-specific CD4+ T cells was measured in the Tg mice immunized with HBc antigen, using an IL-4 secretion assay from Miltenyi Biotec. Figure 5 shows a representative experiment from three experiments that were performed. Thus, 1.27% and 0.78% HBc111–125-specific HLA-DR1-restricted, IL4-secreting CD4+ T cells, were observed in HLA-DR1+/+/IAß0/0 and HLA-DR1+/+/IAß+/+ mice, respectively. The splenocytes used in these experiments were from the mice shown in Figs 3(A) and (B). The HLA-DR1+/+/IAß0/0 mice displayed strong HLA-DR1-restricted HBc-specific responses, while the HLA-DR1+/+/IAß+/+ mice responded to the same antigen only weakly. Simultaneously, an IFN{gamma} secretion assay was performed on these splenocytes, and no HBc111–125-HLA-DR1-restricted, IFN{gamma}-secreting CD4+ T cells were observed (data not shown). Thus, the CD4+ T cells that proliferated after stimulation with the HBc-derived peptide had the TH2 phenotype.



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Fig. 5. Cytokine production after peptide stimulation. Splenocytes from DR1+/+ IAß0/0 or DR1+/+ IAß+/+ HBc immunized mice were analyzed on day 12. Cells were stimulated for 4 h with 20 µg/ml of HBc111–125 or irrelevant peptide. IL4 secretion was measured by flow cytometry. The percentages represent the percent of total cells in each quadrant.

 

    Discussion
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 Abstract
 Introduction
 Methods
 Results
 Discussion
 References
 
We have previously described the improved capacity of HLA-A2-transgenic mice deficient for H-2 class I expression (HLA-A2+/+/ß2m0/0) to mount HLA-A2-restricted CTL responses, as compared with HLA-A2 transgenic mice that still express their endogenous murine H-2b class I molecules (HLA-A2+/+/ß2m+/+) (12,13,24). We have demonstrated that the HLA-A2+/+/ß2m0/0 mice represent the most versatile animal model currently available for CTL epitope mapping and preclinical characterization of HLA-A2-restricted CTL responses. Recent studies have revealed a major role for CD4+ TH lymphocytes in both the initiation and maintenance of CD8+ immune responses. However, only a limited number of helper TH cell epitopes restricted by specific human alleles have been identified to date (25,26).

HLA class II transgenic mice were first designed in order to study auto-immune disease (19,27,28). More recently, HLA class II Tg animals that express HLA class II molecules in the absence of murine class II molecules (Aß0/0) have been generated and were rapidly used for screening of epitopes for human vaccinology (29,30). In the present study, we evaluated the influence of endogenous H-2 class II restricted T cell responses on the HLA-DR1-restricted T cell responses in human HLA class II Tg mice. We therefore compared the HLA-DR1-restricted T cell responses in HLA-DR1 Tg mice deficient for H-2 class II expression (HLA-DR1+/+/IAß0/0) where all CD4+ T cells are restricted by the HLA molecules, and in HLA-DR1+/+/IAß+/+ Tg mice.

The expression level of HLA-DR1 molecules was equivalent in both lineages (0.8 log increase over the control) and the HLA-DR1 transgene was fully functional in thymic T cell education, since 14% of CD4+ peripheral T cells were detected in HLA-DR1+/+/IAß0/0 mice, compared to the residual 0.9% found in IAß0/0 mice. While we observed a minor difference in the number of mobilizable CD4+ peripheral T cells in HLA-DR1+/+/IAß0/0 and HLA-DR1+/+/IAß+/+ mice, the Vß gene segment usage was similar. The small increase in the overall number of CD4+ T cells in HLA-DR1+/+/IAß+/+ mice could be due to the higher cell surface expression of endogenous IAb molecules and thus enhanced T cell thymic education in these mice. In addition, no significant difference in the CDR3 length distribution for each BV chain usage was observed between the three strains of mice, demonstrating that the transgenic complex HLA-DR1 is fully functional in the selection process, and suggesting that BV usage is determined mainly by intracellular processes (e.g. recombination events, promoter strength). We then examined and compared the antigen-specific HLA-DR1-restricted responses induced by immunization with HBV capsid (HBc) or envelope (HBs) protein. A high stimulation index was induced by either HLA-DR1-restricted HBc or HBs peptides in nearly all immunized HLA-DR1+/+/IAß0/0 mice (Table 1). The same peptides elicited weak responses in half of the HLA-DR1+/+/IAß+/+ immunized mice; interestingly, all the mice exhibited strong H-2-restricted T cell responses (Table 1). These findings indicate that coexpression of endogenous H-2 class II molecules interferes with the capacity of mice to respond efficiently to HLA-DR1-restricted antigenic peptides. The proliferative index (Fig. 3) correlated with the number of antigen-specific HLA-DR1-restricted T cells present in the immunized mice (Fig. 5), suggesting that the recruitment of antigen-specific HLA-DR1-restricted peripheral T cells in HLA-DR1+/+/IAß+/+ mice was reduced compared to HLA-DR1+/+/IAß0/0 Tg mice. In the case of immunization with HBc Ag, we showed that the frequency of HBc111–125 specific HLA-DR1-restricted CD4+ T cells was higher in HLA-DR1+/+/IAß0/0 mice than in HLA-DR1+/+/IAß+/+ mice, and that the cells secrete IL4 but not IFN{gamma}. This is not surprising, as it was reported that immunization with an antigen having alum as adjuvant generally drives the response toward a TH2 phenotype (31). The fact that there was no difference between both transgenic mice in Vß gene segment or CDR3 length distribution suggests that thymic education proceeds normally in both backgrounds. Nevertheless, in HLA-DR1+/+/IAß+/+ Tg mice, competition between HLA-DR1 and IAb molecules likely begins at the thymic educational stage. The higher level of expression of endogenous IAb and the more efficient interaction of mouse CD4+ molecules with IAb than with HLA-DR1 may both favor IA- over DR1-restricted precursor T cells in HLA-DR1+/+/IAß+/+ mice during the lymphocyte life cycle: thymic education, peripheral maintenance and antigen-driven final mobilization (32,33). In addition, as shown in vivo, peripheral competition between T lymphocyte populations each expressing monoclonally distinct TCR regulates the final size of these populations in lymphoid tissues (34). This competition for immunodominance between HLA-DR1- and H-2 class II-educated lymphocytes could provide a plausible explanation for the reduced size of the mobilized HLA-DR1 population in HLA-DR1+/+/IAß+/+ mice compared to HLA-DR1+/+/IAß0/0 Tg mice. Given the abundance of peptide vaccine candidates, there is a growing need to evaluate their in vivo immunogenicity and their efficacy at inducing desired T-cell responses related to human diseases. For this purpose, the HLA-DR1+/+/IAß0/0 Tg mice provide a more sensitive and specific screening model than the HLA-DR1+/+/IAß+/+ mice. The ‘humanized’ class II Tg mice thus represent a time-saving model for the identification and development of more promising vaccination strategies with relevance for human disease.


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Table 1. Proliferative response of immunized transgenic mice to HLA-DR1- or H-2 IA-restricted peptides derived from the HBc or HBs proteins

 

    Acknowledgements
 
The authors are grateful to N. Sauzet for secretarial assistance. This work was funded by the Institut Pasteur, the INSERM, the Agence Nationale de Recherche contre le SIDA, the Ligue contre le Cancer comité de Paris and the Association pour la Recherche sur le Cancer. A. Pajot was the recipient of a fellowship from the Agence Nationale de Recherche contre le SIDA.


    Abbreviations
 
BC   ß constant
BJ   ß junction
BV   ß variable
CDR   complementarity determining region
CTL   cytotoxic T lymphocyte
TH   helper T lymphocyte
HBc   hepatitis B virus capsid protein
HBs   hepatitis B virus surface protein

    Notes
 
Transmitting editor: E. Vivier

Received 25 February 2004, accepted 17 June 2004.


    References
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 Abstract
 Introduction
 Methods
 Results
 Discussion
 References
 

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