The JI
HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
QUICK SEARCH:   [advanced]


     
 

This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Martinez-Soría, E.
Right arrow Articles by Izui, S.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Martinez-Soría, E.
Right arrow Articles by Izui, S.
The Journal of Immunology, 2004, 173: 2842-2848.
Copyright © 2004 by The American Association of Immunologists

Epitope-Dependent Inhibition of T Cell Activation by the Ea Transgene: An Explanation for Transgene-Mediated Protection from Murine Lupus1

Eduardo Martinez-Soría2,*, Nabila Ibnou-Zekri2,*, Masahiro Iwamoto*, Marie-Laure Santiago-Raber*, Shuichi Kikuchi*, Marie Kosco-Vilbois{dagger} and Shozo Izui3,*

* Department of Pathology and Immunology, University of Geneva, Geneva, Switzerland; and {dagger} NovImmune, Geneva, Switzerland


   Abstract
Top
 Abstract
Introduction
 Materials and Methods
 Results
 Discussion
 References
 
A high level expression of the Ead transgene encoding the I-E {alpha}-chain is highly effective in the suppression of lupus autoantibody production in mice. To explore the possible modulation of the Ag-presenting capacity of B cells as a result of the transgene expression, we assessed the ability of the transgenic B cells to activate Ag-specific T cells in vitro. By using four different model Ag-MHC class II combinations, this analysis revealed that a high transgene expression in B cells markedly inhibits the activation of T cells in an epitope-dependent manner, without modulation of the I-E expression. The transgene-mediated suppression of T cell responses is likely to be related to the relative affinity of peptides derived from transgenic I-E {alpha}-chains (E{alpha} peptides) vs antigenic peptides to individual class II molecules. Our results support a model of autoimmunity prevention based on competition for Ag presentation, in which the generation of large amounts of E{alpha} peptides with high affinity to I-A molecules decreases the use of I-A for presentation of pathogenic self-peptides by B cells, thereby preventing excessive activation of autoreactive T and B cells.


   Introduction
Top
 Abstract
 Introduction
Materials and Methods
 Results
 Discussion
 References
 
Systemic lupus erythematosus (SLE)4 is considered to be the prototypic systemic autoimmune disease. Unlike organ-specific autoimmune diseases such as type I diabetes mellitus or multiple sclerosis, SLE has the potential to directly involve multiple organ systems as a consequence of the production of multiple autoantibodies against a wide range of self-Ags. Although the precise mechanisms by which these autoantibodies mediate SLE pathology have not yet been totally elucidated, some autoantibodies form circulating immune complexes, leading to the development of glomerulonephritis and systemic vasculitis.

There is considerable evidence that the development of SLE has a strong genetic basis, with contributions from the MHC-linked and multiple non-MHC-linked disease-associated genes (1, 2). Because the development of SLE is dependent on CD4+ T cells (3) and is blocked by treatment with anti-I-A Abs (4), the MHC class II molecules are probably involved in the development of lupus-like autoimmune syndrome. A role for MHC molecules in the regulation of murine SLE has been further supported by several genetic studies in H2 congenic lupus-prone mice (5, 6, 7, 8, 9, 10). Although little is known about how certain MHC alleles regulate the development of SLE, we have previously shown that the MHC class II Ea gene encoding the I-E {alpha}-chain (E{alpha}-chain) may act as a lupus protective gene in mice, because the introduction of two copies of the Ead transgene (Ea-Tg) in the lupus-prone E{alpha}-chain-deficient BXSB strain (I-E–, H2b) is sufficient to prevent the development of SLE (10). This was further supported by the demonstration that a high level expression of Ea-Tg is highly effective in the protection from SLE in several other lupus-prone mice (11, 12).

The mechanism responsible for the Ea-Tg-mediated protection from SLE is still poorly understood. Studies of transgenic and nontransgenic mixed bone marrow chimeras revealed that these chimeric mice are able to develop a typical lupus-like autoimmune syndrome in which anti-DNA autoantibody and T cell-dependent Ab formation is preferentially induced by nontransgenic B cells independently of expression of the transgene at the level of thymic epithelial cells and bone marrow-derived APCs (13, 14). These results suggest that B cells are the major target of transgene-mediated suppression of autoimmune responses, and that transgene expression may interfere with an efficient interaction between autoreactive T and B cells, possibly by modulating the presentation of pathogenic self-peptides by MHC class II molecules. This could occur as a result of increased formation of peptides derived from the transgenic E{alpha}-chains, as an E{alpha}52–68 peptide has been identified as one of the major self-peptides presented by I-Ab and I-Ad molecules (15, 16). The hypothesis that the action of the transgene is mediated through the interaction of its products with the MHC class II molecules, thus competing with potentially pathogenic self-peptides, was further supported by a recent demonstration that the transgene effect is highly dependent on the host H2 haplotype (17).

To explore more directly the possible modulation of Ag-presenting capacity as a result of Ea-Tg expression in B cells, we assessed the ability of transgenic B cells to present antigenic peptides and activate Ag-specific T cells in vitro. We show in this study that the expression of the Ea-Tg resulted in a marked reduction of the Ag-presenting ability of B cells to activate CD4+ T cells in an epitope-dependent manner. This provides an explanation for the ability of the Ea-Tg to prevent the development of lupus-like autoimmune syndrome in mice.


   Materials and Methods
Top
 Abstract
 Introduction
 Materials and Methods
Results
 Discussion
 References
 
Mice

Two BXSB (H2b) Ea-Tg lines, BXSB-E7 and BXSB-E3, which contain, respectively, a single and ~100 copies of the transgene, and BXSB.H2d and BXSB.H2k congenic mice, created by backcross procedures at the 12th generation, were established as previously described (7, 10, 11, 13). Sp6 anti-DNP IgM transgenic (18) and D011.10 OVA323–339-I-Ad TCR transgenic BALB/c mice (19) were obtained from Dr. A. Rolink (University of Basel, Basel, Switzerland). The presence of Ea-Tg in F1 offspring was screened by Southern blot analysis, as described previously (11, 13). Expression of the Sp6 and D011.10 transgenes was detected by surface staining of peripheral blood B and T cells with biotinylated idiotypic anti-Sp6 (20.5) (20) and anti-D011.10 (KJ1-26) mAb (19), respectively. Three- to 4-mo-old female mice were used in this study.

Quantification of E{alpha}52–68-I-Ab complexes on B cells

To obtain a quantitative assessment of the portion of I-Ab molecules loaded with the E{alpha}52–68 peptide in B cells, Ab blocking experiments were performed according to the method described by Ignatowicz et al. (21). PBMC from BXSB female mice were first stained with FITC-conjugated anti-mouse µ-chain (LO-MM-9) mAb, then incubated with 100 µg of purified inhibitor mAb, either anti-E{alpha}52–68-I-Ab (Y-Ae) (22) or anti-I-Ab (Y-3P) (23), for 30 min at 4°C in the presence of saturating concentrations of 2.4G2 anti-Fc{gamma}RII/III mAb. After washing, the cells were stained with biotinylated Y-3P mAb, followed by PE-conjugated streptavidin. Cells were then washed, and the mean net fluorescence (MNF) was determined with a FACSCalibur flow cytometer (BD Biosciences, Mountain View, CA). Because the Y-3P and Y-Ae mAb compete for binding to I-Ab molecules loaded with the E{alpha}52–68 peptide (24), the percentage of occupancy of I-Ab with the E{alpha}52–68 peptide was estimated as: 100 x (Y-3P MNF – Y-3P MNF with excess Y-Ae)/(Y-3P MNF – Y-3P MNF with excess Y-3P).

Purification of splenic B and T cells

B and T cells were purified from the spleens of a pool of three female mice at 3–4 mo of age, as described previously (25). Briefly, B cells were purified from spleen cells by adherence of macrophages to plastic culture plates for 1 h at 37°C and subsequent treatment with anti-Thy-1.2 (AT-83) mAb in the presence of rabbit complement (Cedarlane Laboratories, Hornby, Canada). CD4+ T cells were purified by treatment with anti-CD8 (H35-17.2) and anti-B220 (RA3-3A1) mAb in the presence of rabbit complement. The purity of B and CD4+ T cells, as documented by cytofluorometric analysis, was >95%.

Ag presentation assays

Ag presentation assays were performed by incubating 2 x 105 splenic B cells, derived from (BALB/cxBXSB)F1 female mice expressing either both the Ea-Tg and Sp6 transgene or the Sp6 transgene alone or from BXSB female mice with or without the Ea-Tg with different concentrations of DNP-conjugated OVA or hen egg lysozyme (HEL) at 37°C for 24 h in a total volume of 200 µl of DMEM containing 5% FCS in 96-well microtiter plates. After washing the cells, 2 x 105 T cells from (BALB/cxBXSB)F1 females expressing the D011.10 OVA323–339-I-Ad TCR transgene or 5 x 104 BO4H9.1 HEL74–88-I-Ab (26), 2B5.1 HEL46–61-I-Ak, or 2G7.1 HEL1–18-I-Ek hybridoma T cells (27) were added to the culture. Supernatants were collected after 1 and 5 days of coculture and were tested for IL-2 and IgM contents, respectively. IL-2 concentrations were measured using IL-2-dependent CTLL-2 cells, with a standard curve obtained by rIL-2. Results are expressed as a stimulation index that represents the ratio of IL-2 concentrations in culture supernatants in the presence of Ags/IL-2 concentrations spontaneously released during the culture of B and T cells without Ags. IgM concentrations were determined by IgM-specific ELISA.

Flow cytometric analysis of HEL46–61-I-Ak complexes on B cells

To study the expression level of HEL46–61-I-Ak complexes on the surface of B cells, 5 x 105 spleen cells were incubated with different concentrations of DNP-HEL conjugates overnight at 37°C in a volume of 1 ml of DMEM containing 10% FCS. Then the cells were stained with FITC-labeled anti-mouse µ-chain (LO-MM-9) mAb and biotinylated anti-HEL46–61-I-Ak (C4H3) mAb (28) in the presence of saturating concentrations of 2.4G2 anti-Fc{gamma}RII/III mAb, followed by PE-conjugated streptavidin.

Statistical analysis

Statistical differences between groups from three or four independent experiments were analyzed by using a two-way repeated measures ANOVA. A value of p < 0.05 was considered significant.


   Results
Top
 Abstract
 Introduction
 Materials and Methods
 Results
Discussion
 References
 
Increased expression of E{alpha}52–68-I-Ab complexes in Ea-Tg B cells

A high level expression of Ea-Tg in BXSB-E3 transgenic mice carrying ~100 copies of the transgene led to an enhanced expression of E{alpha}52–68-I-Ab complexes, detectable with the Y-Ae mAb, on the surface of B cells (Fig. 1A). Their fluorescence intensity (mean of three mice ± SD, 209 ± 17) was ~4-fold higher than that seen in BXSB-E7 homozygous transgenics bearing two copies of the functional Ea genes (mean of three mice ± SD, 55 ± 7). To obtain a more quantitative assessment of the proportion of I-Ab molecules loaded with the E{alpha}52–68 peptide in transgenic B cells, we performed an Ab blocking analysis in which the percentage of cell surface I-Ab molecules occupied by the E{alpha}52–68 peptide was estimated from the ability of Y-Ae mAb to inhibit binding of Y-3P mAb (21). Y-Ae mAb inhibited binding of Y-3P mAb to BXSB-E3 transgenic B cells by 36% (Fig. 1B), suggesting that more than one-third of the I-Ab molecules were occupied by E{alpha}52–68 peptide in B cells expressing a high Ea Tg level. In contrast, ~10% of surface I-Ab molecules were estimated to be occupied by E{alpha}52–68 peptide on B cells from BXSB-E7 homozygous transgenics, in agreement with results obtained by quantitative binding analysis on B cells from B10.A(5R) mice expressing I-Ab and I-E (E{alpha}dE{beta}b) at homozygous levels (24). It is also worth noting that when murine B cell lines were incubated with exogenous HEL proteins, the percentage of I-Ak on the surface of B cells occupied by HEL46–61 peptides was 9% at maximum (29).



View larger version (12K):
[in this window]
[in a new window]
  		FIGURE 1. A, Expression levels of E{alpha}52–68-I-Ab complexes on B cells from BXSB-E3 transgenic (E3-Tg), BXSB-E7 homozygous transgenic (E7-Tg), and BXSB nontransgenic (N-Tg) female mice. Circulating B cells from different mice were stained with FITC-labeled anti-mouse µ-chain (LO-MM-9) mAb and biotinylated anti-E{alpha}52–68-I-Ab (Y-Ae) mAb in the presence of saturating concentrations of 2.4G2 anti-Fc{gamma}RII/III mAb, followed by PE-conjugated streptavidin. Histograms show representative results of Y-Ae staining of IgM+ B cells obtained from three mice. The specificity of the Y-Ae staining was controlled by using B cells from (BALB/cxBXSB)F1 mice (I-Ab- and I-E-positive) as a positive control and B cells from BXSB (I-Ab-positive, I-E-negative) and BALB/c (I-Ab-negative, I-E-positive) mice as negative controls (not shown). B, The proportion of I-Ab molecules occupied by E{alpha}52–68 peptide on B cells from BXSB-E3 transgenic (•), BXSB-E7 homozygous transgenic ({blacktriangleup}), and BXSB nontransgenic ({circ}) female mice. Results are shown as the percent inhibition of anti-I-Ab (Y-3P) binding by excess amounts of anti-E{alpha}52–68-I-Ab (Y-Ae) mAb in three individual mice.

 
Lower activation of OVA323–339-I-Ad-specific T cells by anti-DNP B cells expressing a high level of Ea-Tg in the presence of DNP-OVA conjugates

An increased occupation of cell surface I-Ab molecules by E{alpha}52–68 peptide in B cells expressing a high level of Ea-Tg suggested that the presentation of peptides derived from foreign Ags by I-A molecules may be inhibited in Ea-Tg B cells. If so, the extent of this inhibition should be dependent on the relative affinity of E{alpha}52–68 peptide vs antigenic peptides to I-A molecules. Because the affinity of {alpha}52–68 peptide to I-Ad is apparently >10 times higher than that of OVA323–339 peptide (16), we determined whether the expression of Ea-Tg is able to inhibit the presentation of OVA323–339 peptide to OVA323–339-I-Ad-specific T cells. For this purpose, BXSB-E3 mice expressing a high level of Ea-Tg were crossed with BALB/c mice carrying the Sp6 anti-DNP IgM transgene (BALB.Sp6), and Sp6/Ea double-transgenic or Sp6 single-transgenic B cells (H2d/b) were incubated with DNP-conjugated OVA for 24 h. Then we assessed their abilities to stimulate OVA-specific T cells from (BALB/cxBXSB)F1 mice bearing the D011.10 OVA323–339-I-Ad TCR transgene. T cell responses after coculture with anti-DNP B cells expressing Ea-Tg were markedly limited compared with those of cells stimulated with anti-DNP B cells lacking Ea-Tg (Fig. 2A). IgM secretion by anti-DNP Ea-Tg B cells, determined 5 days after coculture, was also strongly inhibited (Fig. 2B). The observed inhibition was dependent on the expression level of Ea-Tg, because the ability of anti-DNP B cells expressing a low Ea-Tg level, derived from (BALB.Sp6xBXSB-E7)F1 mice, to stimulate OVA-specific T cells was not significantly diminished (Fig. 2C).



View larger version (24K):
[in this window]
[in a new window]
  		FIGURE 2. Capacity of Ea-Tg and nontransgenic B cells bearing the H2d/b or H2d/d haplotype to activate OVA-specific T cells in vitro. A and B, T cells derived from D011.10 OVA323–339-I-Ad TCR transgenic (BALB/cxBXSB)F1 female mice (H2d/b) were stimulated with splenic anti-DNP B cells derived from H2d/b (BALB.Sp6xBXSB-E3)F1 Sp6 transgenic female mice expressing a high level of Ea-Tg (E3-Tg; •) or lacking Ea-Tg (N-Tg; {circ}) in the presence of different concentrations of DNP-conjugated OVA. After 24 h, IL-2 contents in supernatants were determined using IL-2-dependent CTLL-2 cells, and results are expressed as the stimulating index (A). After 5 days, IgM concentrations in supernatants were measured by ELISA, and results are expressed as micrograms per milliliter (B). Mean values (±1 SEM) from four independent experiments are shown, in which differences in IL-2 and IgM secretion in culture with N-Tg B cells vs Ea-Tg B cells were significant (p < 0.0005 and p < 0.001, respectively). C, OVA323–339-I-Ad TCR transgenic T cells were stimulated with H2d/b (BALB.Sp6xBXSB-E7)F1 Sp6 transgenic female mice expressing a low level of Ea-Tg (E7-Tg; {blacktriangleup}) or lacking Ea-Tg (N-Tg; {circ}) in the presence of different concentrations of DNP-conjugated OVA. After 24 h, IL-2 contents in supernatants were determined, and results are expressed as the stimulating index. Mean values (±1 SEM) from three independent experiments are shown, in which no significant differences in IL-2 secretion in culture with N-Tg B cells vs Ea-Tg B cells were observed. D, T cells derived from OVA323–339-I-Ad TCR transgenic (BALB/cxBXSB.H2d)F1 female mice (H2d/d) were stimulated with splenic anti-DNP B cells from H2d/d (BALB.Sp6xBXSB.H2d-E3)F1 Sp6 transgenic female mice expressing a high level of Ea-Tg (E3-Tg; •) or lacking Ea-Tg (N-Tg; {circ}) in the presence of different concentrations of DNP-conjugated OVA. After 24 h, IL-2 contents in supernatants were determined, and results are expressed as the stimulating index. Mean values (±1 SEM) from three independent experiments are shown, in which the difference in IL-2 secretion in culture with N-Tg B cells vs Ea-Tg B cells was significant (p < 0.05).

 
Because (BALB.Sp6xBXSB-E3)F1 mice are H2d/b heterozygous, the high level Ea-Tg expression led not only to increased generation of E{alpha}52–68 peptide, but also to enhanced expression of both conventional I-Ed (E{alpha}dE{beta}d) and mixed haplotype E{alpha}dE{beta}b heterodimers, as shown previously in H2d/b (NZBxBXSB-E3)F1 mice (12). Therefore, the observed inhibition of OVA-specific T cell responses by H2d/b anti-DNP Ea-Tg B cells could also result from the increased expression of I-E molecules, leading to competition between I-A and I-E molecules for peptide presentation, as reported in Ea-Tg NOD mice by Deng et al. (30, 31). To exclude the latter effect and to better define the contribution of the E{alpha} peptide-dependent inhibitory mechanism, we assessed OVA-specific T cell responses upon stimulation with anti-DNP Ea-Tg or control B cells from H2d/d homozygous (BALB.Sp6xBXSB.H2d-E3)F1 mice. In these F1 mice, both Ea-Tg and control B cells expressed only conventional I-Ed molecules, the expression level of which was unchanged even in the presence of the Ea-Tg (mean fluorescence intensities of three mice ± SD: Ea-Tg, 313 ± 17; nontransgenics, 309 ± 12). Thus, if the presentation of OVA323–339 peptides by I-Ad molecules is inhibited by high level expression of Ea-Tg, this cannot be due to competition between I-A and I-E molecules for OVA peptide presentation in this model. Indeed, when OVA-specific T cells were cocultured with H2d/d homozygous anti-DNP B cells expressing a high Ea-Tg level, significant suppression of T cell responses was again obtained (Fig. 2D). These data suggested that competition between E{alpha} peptide and OVA323–339 peptide for I-Ad binding played the major role in the Ea-Tg-mediated suppression of OVA-specific T cell activation.

Lower activation of HEL74–88-I-Ab-specific T cells by anti-DNP B cells expressing a high level of Ea-Tg in the presence of DNP-HEL conjugates

To further test the possibility that excessive generation of the E{alpha}52–68 peptide with a higher affinity to I-A can compete with antigenic peptides for I-A binding, we assessed the capacity of anti-DNP Ea-Tg B cells to activate HEL74–88-I-Ab-specific BO4H9.1 T cell hybridoma. The relative affinity for I-Ab molecules of the HEL74–88 peptide vs the E{alpha}52–68 peptide has not been defined. However, C57BL/6 mice expressing only I-Ab molecules are known to be genetically nonresponsive to HEL (32), indicating that the HEL74–88 peptide should display a very weak affinity to I-Ab or be poorly generated in B cells incubated with HEL proteins. Thus, it is expected that the E{alpha}52–68 peptide, the major I-Ab-binding peptide (15), could compete with presentation of the HEL74–88 peptide by I-Ab. As observed with the OVA323–339-I-Ad system, a high Ea-Tg expression in anti-DNP B cells, from H2d/b (BALB.Sp6xBXSB-E3)F1 mice, preincubated with DNP-HEL conjugates strongly inhibited the activation of HEL74–88-I-Ab-specific T cells (Fig. 3).



View larger version (20K):
[in this window]
[in a new window]
  		FIGURE 3. Capacity of H2d/b Ea-Tg and nontransgenic B cells to activate HEL-specific T cells in vitro. HEL74–88-I-Ab-specific BO4H9.1 T cells were stimulated with splenic anti-DNP B cells, derived from H2d/b (BALB.Sp6xBXSB-E3)F1 Sp6 transgenic female mice expressing a high level of Ea-Tg (•) or lacking Ea-Tg (N-Tg; {circ}) in the presence of different concentrations of DNP-conjugated HEL. After 24 h, IL-2 contents in supernatants were determined, and results are expressed as the stimulating index. Mean values (±1 SEM) from three independent experiments are shown, in which the difference in IL-2 secretion in culture with N-Tg B cells vs Ea-Tg B cells was significant (p < 0.01).

 
Efficient activation of HEL46–61-I-Ak-specific T cells by B cells expressing a high level of Ea-Tg in the presence of DNP-HEL conjugates

According to the E{alpha} peptide competition hypothesis, the expression of Ea-Tg should be unable to inhibit the Ag presentation and activation of Ag-specific T cells, if the affinity of antigenic peptides to the corresponding I-A molecules is higher than that of the E{alpha} peptide. In the system of HEL46–61 peptide and I-Ak, the affinity of the HEL46–61 peptide to the I-Ak molecule should be much higher than that of the E{alpha}52–68 peptide for the following reasons. First, peptide elution experiments on I-Ak-bearing splenic B cells did not identify peptides derived from the E{alpha}-chains as a major I-Ak-binding peptides (33), whereas the HEL46–61 peptide was readily detectable after incubation with HEL (34). Second, the direct binding analysis between the HEL46–61 peptide and I-Ak showed a high affinity interaction, ~40 times higher than that of the OVA323–336 peptide with I-Ad (35). By using B cells from H2d/k (BALB.Sp6xBXSB.H-2k-E3)F1 mice, we explored whether the high level expression of Ea-Tg suppresses the activation of HEL46–61-I-Ak-specific 2B5.1 T cell hybridoma. In marked contrast to the results obtained with the OVA323–339-I-Ad and HEL74–88-I-Ab systems, T cell responses were hardly inhibited after coculture with anti-DNP Ea-Tg B cells pulsed with DNP-HEL conjugates (Fig. 4A). Notably, the expression level of HEL46–61-I-Ak complexes on B cells after incubation with different concentrations of DNP-HEL proteins (as determined by staining with the C4H3 mAb specific for HEL46–61-I-Ak complexes) was indistinguishable between anti-DNP Ea-Tg and control B cells (Fig. 4B). These results indicated that the inhibition of T cell activation by B cells with high Ea-Tg expression was selective and dependent on T cell epitopes.



View larger version (16K):
[in this window]
[in a new window]
  		FIGURE 4. A, Capacity of H2d/k Ea-Tg and nontransgenic B cells to activate HEL-specific T cells in vitro. HEL46–61-I-Ak-specific 2B5.1 T cells were stimulated with splenic anti-DNP B cells derived from H2d/k (BALB.Sp6xBXSB.H2k-E3)F1 Sp6 transgenic female mice expressing a high level of Ea-Tg (•) or lacking Ea-Tg (N-Tg; {circ}) in the presence of different concentrations of DNP-conjugated HEL. After 24 h, IL-2 contents in supernatants were determined, and results are expressed as the stimulating index. Mean values (±1 SEM) from four independent experiments are shown, in which no significant differences in IL-2 secretion in culture with N-Tg B cells vs Ea-Tg B cells were observed. B, Surface expression of HEL46–61-I-Ak complexes on B cells from H2d/k Ea-Tg and nontransgenic mice. Spleen cells from H-2d/k (BALB.Sp6xBXSB.H2k-E3)F1 Sp6 transgenic female mice expressing a high level of Ea-Tg (•) or lacking Ea-Tg (N-Tg; {circ}) were incubated with different concentrations of DNP-conjugated HEL overnight at 37°C. The expression level of HEL46–61-I-Ak complexes on the surface of IgM+ splenic B cells was determined by staining with rat anti-HEL46–61-I-Ak (C4H3) mAb in the presence of saturating concentrations of 2.4G2 anti-Fc{gamma}RII/III mAb. Results are expressed as mean fluorescence intensities on IgM+ B cells obtained from three mice. SDs at different points in both groups of mice were <10% of each mean value. The specificity of C4H3 staining was controlled using B cells from H2k and H2b BXSB mice with or without incubation of HEL proteins (not shown).

 
Enhanced activation of HEL1–18-I-Ek-specific T cells by H2k/b heterozygous B cells expressing a high level of Ea-Tg in the presence of DNP-HEL conjugates

Results obtained with the OVA323–339-anti-I-Ab and HEL74–88-I-Ab systems were intriguing, because the extent of occupancy of I-Ab by the E{alpha}52–68 peptides in B cells expressing a high level of Ea-Tg was not >40% (see Fig. 1B). This suggested that relatively small differences in the availability of I-A molecules could have significant consequences for the activation of some Ag-specific T cells. In fact, we observed that Ea-Tg B cells, derived from H2k/b heterozygous BXSB-E3 transgenic mice and pulsed with DNP-HEL proteins, activated 2G7.1 HEL1–18-I-Ek-specific T cell hybridoma much more efficiently than nontransgenic H2k/b BXSB B cells (Fig. 5A). This enhancement is probably due to an increased expression of E{alpha}dE{beta}k, but not E{alpha}dE{beta}b, heterodimers in H2k/b Ea-Tg B cells for two reasons; first, a similarly increased activation of 2G7.1 T cell hybridoma was obtained with H2k homozygous BXSB B cells, and second, B cells expressing E{alpha}dE{beta}b from H2b BXSB-E3 transgenic mice were unable to activate 2G7.1 T cells (Fig. 5B). Notably, the peptide binding specificity of the E{alpha}dE{beta}k heterodimer should be identical with that of the conventional I-Ek (E{alpha}kE{beta}k) molecule, because the extracellular regions of the E{alpha}d- and E{alpha}k-chains differ only by a single amino acid localized at the junction between the {alpha}2 and transmembrane domains (36). The precise amounts of I-Ek and E{alpha}kE{beta}b expressed in H2k/b nontransgenic B cells cannot be estimated due to a cross-reactivity of available anti-E{beta}k mAb such as H9-14.8 and H40-242.3 (37, 38) to the E{beta}b-chains (data not shown). Considering the known restricted pairing of allelically matched {alpha}- and {beta}-chains of MHC class II heterodimers (39, 40), the E{alpha}k-chains could form I-E heterodimers with the E{beta}k-chains more efficiently than with the E{beta}b-chains in H2k/b nontransgenic B cells, whereas both E{beta}k- and E{beta}b-chains are fully expressed as a result of excessive synthesis of the transgenic E{alpha}d-chains in H2k/b Ea-Tg B cells. Accordingly, it can be expected that because of the limited amounts of available E{beta}k-chains, the total expression level of I-E molecules (I-Ek and E{alpha}dE{beta}k) involved in the 2G7.1 HEL1–18-I-Ek T cell response in H2k/b Ea-Tg B cells cannot be >2 times higher than that of I-Ek in nontransgenic B cells. Thus, these results indicated that relatively small changes in the availability of MHC class II molecules could lead to substantial differences in the activation of certain T cell responses.



View larger version (15K):
[in this window]
[in a new window]
  		FIGURE 5. Capacity of Ea-Tg and nontransgenic B cells bearing the H2k/b or H2k/k haplotype to activate HEL-specific T cells in vitro. HEL1–18-I-Ek-specific 2G7.1 T cells were stimulated with splenic B cells from H2k/b BXSB-E3 Ea-Tg ({circ}) or nontransgenic (N-Tg; •) female mice (A) or from BXSB nontransgenic female mice bearing the H2k/k ({circ}) or H2k/b (•) haplotype (B) in the presence of different concentrations of DNP-conjugated HEL. Notably, B cells derived from H2b BXSB-E3 Ea-Tg mice expressing E{alpha}kE{beta}b heterodimers were unable to activate 2G7.1 T cells ({blacktriangleup}; B). After 24 h, IL-2 contents in supernatants were determined, and results are expressed as the stimulating index. Mean values (±1 SEM) from three independent experiments are shown, in which differences in IL-2 secretion in culture with N-Tg B cells vs Ea-Tg B cells (A) and in culture with H2k/k B cells vs H2k/b B cells (B) were significant (p < 0.0005 and p < 0.0001, respectively).

 

   Discussion
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
References
 
In the present study by using four different combinations of antigenic epitopes and MHC class II molecules we have demonstrated that a high level of expression of Ea-Tg in B cells is able to inhibit the activation of Ag-specific CD4+ T cells in vitro in an epitope-dependent manner, and that this Ea-Tg-mediated suppression occurs independently of the modulation of I-E expression. These data suggest that the Ea-Tg-mediated inhibition is dependent on the generation of large amounts of the E{alpha}52–68 peptide derived from the transgenic E{alpha}-chains, displaying a high affinity for I-A molecules. Thus, the relative affinity of the E{alpha}52–68 peptide vs antigenic peptides for individual MHC class II molecules may play a key role in the Ea-Tg-mediated inhibition of T cell activation. Although our present demonstration is still indirect, the observed inhibition of T cell activation by a high level of expression of Ea-Tg probably reflects the Ea-Tg-mediated prevention of lupus-like autoimmune syndrome in mice. This idea is also supported by the finding that a low level expression of Ea-Tg, which conferred limited protection against the development of SLE (11, 12), failed to significantly inhibit T cell activation in vitro. Furthermore, in preliminary studies we have observed complete protection from SLE in BXSB.H2q-E3 transgenic mice without the induction of I-E expression, because of defects in both Ea and Eb genes in H2q mice (41, 42). Taken together, our results support the hypothesis that a high level expression of E{alpha}-chains and a subsequent increased formation of E{alpha} peptides in B cells may decrease the use of I-A molecules for the presentation of pathogenic self-peptides, thereby limiting the activation of autoreactive T and B cells.

It should be stressed that B cells expressing a high level of Ea-Tg are capable of presenting antigenic peptides and activating HEL46–61-I-Ak-specific T cells as efficiently as nontransgenic B cells. These results clearly rule out the possibility that the diminished T cell activation by the transgenic B cells in the OVA323–339-I-Ad and HEL74–88-I-Ab systems results from a nonspecific functional defect in B cells caused by the presence of a high copy number of the transgene, unlike those reported previously (43). In addition, our results argue against a role of E{alpha} peptides in the blocking of IL-2 signaling, unlike the effect observed with an HLA DQ65–79 peptide, which apparently antagonizes PI3K (44). Furthermore, we observed a remarkably enhanced activation of HEL1–18-I-Ek-specific T cells by Ea-Tg B cells as a consequence of increased expression (although <2-fold) of I-E molecules involved in this T cell response. This indicates that relatively small changes in the availability of MHC class II molecules can be sufficient to modulate some Ag-specific T cell responses to a large extent, consistent with limited activation by Ea-Tg B cells of certain T cell responses, despite the fact that the extent of the occupation of I-A molecules by the E{alpha}52–68 peptide in transgenic B cells was not >40%.

It is significant that the Ea-Tg-mediated suppressing effect on T cell activation in vitro was dependent on the types of Ags and MHC class II molecules. These results are in agreement with our recent demonstration that the level of protection conferred by Ea-Tg is highly dependent on the host H2 haplotype: strongest with the H2b haplotype, intermediate with the H2k haplotype, and weakest with the H2d haplotype (17). It should also be stressed that the autoimmune inhibitory effect mediated by the Ea-Tg is dependent on the type of autoimmune responses. For example, anti-DNA autoantibody production is more resistant to the transgene effect than anti-gp70 autoantibody production (12, 17). These differences are probably related to variabilities in the binding affinity of individual MHC class II molecules for the E{alpha} peptides vs pathogenic self-peptides. Thus, the overall disease-suppressing effect of Ea-Tg can be determined by the affinity of E{alpha} peptides for individual MHC class II molecules and the relative contributions of individual MHC class II molecules to the development of lupus-like autoimmune responses.

Despite the present demonstration that a high Ea-Tg expression is able to inhibit the activation of Ag-specific T cells in vitro, Ea-Tg expression does not affect immune responses against foreign Ags in vivo (11, 13, 14). This suggests that the autoimmune responses occurring in lupus-prone mice may be qualitatively different from immune responses against foreign Ags. The absence of T cell responses in Ea-Tg BXSB mice against the E{alpha}52–68 peptide, the dominant epitope of E{alpha}-chains in the context of I-Ab, indicates an efficient tolerance induction in T cells specific for dominant epitopes of self-Ags even in lupus-prone mice (11). Thus, autoreactive T cells implicated in SLE may be those specific to semidominant epitopes of self-Ags, which can evade tolerance induction. The avidity of these TCR for the self-peptide-MHC complexes is too low to induce their activation in the periphery (45), but such autoreactive T cells might be activated in lupus-prone mice, probably due to genetic defects predisposing them to autoimmune diseases. In this situation, the generation of large amounts of E{alpha}52–68 peptides in Ea-Tg mice could relatively easily down-modulate the presentation of semidominant epitopes of pathogenic autoantigens below the threshold levels, thereby limiting a low avidity interaction of autoreactive T and B cells. In contrast, this is not the case for immune responses against foreign Ags for two reasons. First, high affinity T cells capable of recognizing the dominant epitopes of foreign Ags are fully available. Second, immunization resulting in local accumulation of large amounts of peptides with a high affinity for MHC class II molecules would allow a sufficient presentation of foreign antigenic peptides, even in the presence of E{alpha} peptides, and lead to a normal immune response.

In addition to the Ea-Tg-mediated protection from SLE, the expression of different MHC class II transgenes has been reported to protect from other autoimmune diseases as well: the Ead or AakAbk transgene protects against type I diabetes mellitus in NOD mice (46, 47), the Ebd transgene protects against collagen-induced arthritis in B10.RQB3 mice (48), and the Eak transgene protects against myasthenia gravis (49). Notably, MHC class II A{beta}- and E{beta}-chains, in addition to E{alpha}-chains, are major sources of self-peptides displaying a high affinity for I-A molecules (33, 34, 50). Although the protective mechanisms in these transgenic mice have not been fully elucidated, the generation of MHC-derived peptides may lead to modification of the T cell repertoire, thereby modulating disease development, as proposed recently (51). Thus, the present results suggest that a peptide competition mechanism resulting from excessive generation of peptides derived from the transgene products might also be involved in the protection from autoimmune disease in these MHC class II transgenic mice. Further understanding of the protective mechanism(s) conferred by Ea and other MHC class II transgenes may elucidate the molecular and cellular bases central to the development of SLE and other autoimmune diseases and also have clinical implications in the design of future therapeutic strategies with self-peptides in autoimmune disorders.


   Acknowledgments
 
We thank G. Celetta and G. Brighouse for their excellent technical assistance, Dr. A. Y. Rudensky for the Y-Ae mAb, Dr. R. N. Germain for the C4H3 mAb, Dr. N. Shiastri for the BO4H9.1 T cell hybridoma, Dr. L. Adorini for the 2B5.1 and 2G7.1 T cell hybridomas, and Dr. P. Vassalli for critically reading the manuscript.


   Footnotes
 
The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

1 This work was supported by the Swiss National Foundation for Scientific Research. Back

2 E.M.-S. and N.I.-Z. contributed equally to this work. Back

3 Address correspondence and reprint requests to Dr. Shozo Izui, Department of Pathology, Centre Médical Universitaire, 1211 Geneva 4, Switzerland. E-mail: shozo.izui{at}medecine.unige.ch Back

4 Abbreviations used in this paper: SLE, systemic lupus erythematosus; BALB.Sp6, BALB/c mice expressing the Sp6 anti-DNP transgene; E{alpha}-chain, I-E {alpha}-chain; Ea-Tg, Ead transgene; HEL, hen egg lysozyme; MNF, mean net fluorescence. Back

Received for publication February 11, 2004. Accepted for publication June 1, 2004.


   References
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
 References
 

  1. Vyse, T. J., B. L. Kotzin. 1998. Genetic susceptibility to systemic lupus erythematosus. Annu. Rev. Immunol. 16:261.[Medline]
  2. Wandstrat, A., E. Wakeland. 2001. The genetics of complex autoimmune diseases: non-MHC susceptibility genes. Nat. Immunol. 2:802.[Medline]
  3. Wofsy, D., W. E. Seaman. 1985. Successful treatment of autoimmunity in NZB/NZW F1 mice with monoclonal antibody to L3T4. J. Exp. Med. 161:378.[Abstract/Free Full Text]
  4. Adelman, N. E., D. L. Watling, H. O. McDevitt. 1983. Treatment of (NZB x NZW)F1 disease with anti-I-A monoclonal antibodies. J. Exp. Med. 158:1350.[Abstract/Free Full Text]
  5. Hirose, S., R. Nagasawa, I. Sekikawa, M. Hamaoki, Y. Ishida, H. Sato, T. Shirai. 1983. Enhancing effect of H-2-linked NZW gene(s) on the autoimmune traits of (NZB x NZW)F1 mice. J. Exp. Med. 158:228.[Abstract/Free Full Text]
  6. Chiang, B.-L., E. Bearer, A. Ansari, K. Dorshkind, M. E. Gershwin. 1990. The bm12 mutation and autoantibodies to dsDNA in NZB.H-2bm12 mice. J. Immunol. 145:94.[Abstract]
  7. Merino, R., L. Fossati, M. Lacour, R. Lemoine, M. Higaki, S. Izui. 1992. H-2-linked control of the Yaa gene-induced acceleration of lupus-like autoimmune disease in BXSB mice. Eur. J. Immunol. 22:295.[Medline]
  8. Cohen, P. L., E. Creech, D. Nakul-Aquaronne, R. McDaniel, S. Ackler, R. G. Rapoport, E. S. Sobel, R. A. Eisenberg. 1993. Antigen nonspecific effect of major histocompatibility complex haplotype on autoantibody levels in systemic lupus erythematosus-prone lpr mice. J. Clin. Invest. 91:2761.
  9. Merino, R., M. Iwamoto, M.E. Gershwin, S. Izui. 1994. The Yaa gene abrogates the major histocompatibility complex association of murine lupus in (NZB x BXSB)F1 hybrid mice. J. Clin. Invest. 94:521.
  10. Ibnou-Zekri, N., M. Iwamoto, L. Fossati, P. J. McConahey, S. Izui. 1997. Role of the major histocompatibility complex class II Ea gene in lupus susceptibility in mice. Proc. Natl. Acad. Sci. USA 94:14654.[Abstract/Free Full Text]
  11. Iwamoto, M., N. Ibnou-Zekri, K. Araki, S. Izui. 1996. Prevention of murine lupus by an I-E {alpha} chain transgene: protective role of I-E {alpha} chain-derived peptides with a high affinity to I-Ab molecules. Eur. J. Immunol. 26:307.[Medline]
  12. Iwamoto, M., N. Ibnou-Zekri, T. Kobayakawa, S. Izui. 1998. Effect of genetic background on Ead transgene-mediated protection from murine lupus. J. Autoimmun. 11:241.[Medline]
  13. Merino, R., M. Iwamoto, L. Fossati, P. Muniesa, K. Araki, S. Takahashi, J. Huarte, K.-I. Yamamura, J.-D. Vassalli, S. Izui. 1993. Prevention of systemic lupus erythematosus in autoimmune BXSB mice by a transgene encoding I-E {alpha} chain. J. Exp. Med. 178:1189.[Abstract/Free Full Text]
  14. Creech, E. A., D. Nakul-Aquaronne, E. A. Reap, R. L. Cheek, P. A. Wolthusen, P. L. Cohen, R. A. Eisenberg. 1996. MHC genes modify systemic autoimmune disease: the role of the I-E locus. J. Immunol. 156:812.[Abstract]
  15. Rudensky, A. Y., P. Preston-Hurlburt, S. C. Hong, A. Barlow, C. A. Janeway, Jr. 1991. Sequence analysis of peptides bound to MHC class II molecules. Nature 353:622.[Medline]
  16. Hunt, D. F., H. Michel, T. A. Dickinson, J. Shabanowitz, A. L. Cox, K. Sakaguchi, E. Appella, H. M. Grey, A. Sette. 1992. Peptides presented to the immune system by the murine class II major histocompatibility complex molecule I-Ad. Science 256:1817.[Abstract/Free Full Text]
  17. Ibnou-Zekri, N., M. Iwamoto, M. E. Gershwin, S. Izui. 2000. Protection of murine lupus by the Ead transgene is MHC haplotype-dependent. J. Immunol. 164:505.[Abstract/Free Full Text]
  18. Rusconi, S., G. Köhler. 1985. Transmission and expression of a specific pair of rearranged immunoglobulin µ and k genes in a transgenic mouse line. Nature 314:330.[Medline]
  19. Murphy, K. M., A. B. Heimberger, D. Y. Loh. 1990. Induction by antigen of intrathymic apoptosis of CD4+CD8+TCRlo thymocytes in vivo. Science 250:1720.[Abstract/Free Full Text]
  20. Andersson, J., F. Melchers, A. Rolink. 1995. Stimulation by T cell independent antigens can relieve the arrest of differentiation of immature auto-reactive B cells in the bone marrow. Scand. J. Immunol. 42:21.[Medline]
  21. Ignatowicz, L., G. Winslow, J. Bill, J. Kappler, P. Marrack. 1995. Cell surface expression of class II MHC proteins bound by a single peptide. J. Immunol. 154:3852.[Abstract]
  22. Rudensky, A. Y., S. Rath, P. Preston-Hurlburt, D. B. Murphy, C. A. Janeway, Jr. 1991. On the complexity of self. Nature 353:660.[Medline]
  23. Janeway, C. A., Jr, P. J. Conrad, E. A. Lerner, J. Babich, P. Wettstein, D. B. Murphy. 1984. Monoclonal antibodies specific for Ia glycoproteins raised by immunization with activated T cells: possible role of T cellbound Ia antigens as targets of immunoregulatory T cells. J. Immunol. 132:662.[Abstract]
  24. Murphy, D. B., D. Lo, S. Rath, R. L. Brinster, R. A. Flavell, A. Slanetz, C. A. Janeway, Jr. 1989. A novel MHC class II epitope expressed in thymic medulla but not cortex. Nature 338:765.[Medline]
  25. Kosco-Vilbois, M. H., D. Gray, D. Scheidegger, M. Julius. 1993. Follicular dendritic cells help resting B cells to become effective antigen-presenting cells: induction of B7/BB1 and upregulation of major histocompatibility complex class II molecules. J. Exp. Med. 178:2055.[Abstract/Free Full Text]
  26. Kobori, J. A., L. Hood, N. Shastri. 1992. Structure-function relationship among T-cell receptors specific for lysozyme peptides bound to Ab or Abm12 molecules. Proc. Natl. Acad. Sci. USA 89:2940.[Abstract/Free Full Text]
  27. Adorini, L., J.-C. Guéry, S. Fuchs, V. Ortiz-Navarrete, G. J. Hämmerling, F. Momburg. 1993. Processing of endogenously synthesized hen egg-white lysozyme retained in the endoplasmic reticulum or in secretory form gives rise to a similar but not identical set of epitopes recognized by class II-restricted T cells. J. Immunol. 151:3576.[Abstract]
  28. Zhong, G., C. Reis e Sousa, R. N. Germain. 1997. Production, specificity, and functionality of monoclonal antibodies to specific peptide-major histocompatibility complex class II complexes formed by processing of exogenous protein. Proc. Natl. Acad. Sci. USA 94:13856.[Abstract/Free Full Text]
  29. Dadaglio, G., C. A. Nelson, M. B. Deck, S. J. Petzold, E. R. Unanue. 1997. Characterization and quantitation of peptide-MHC complexes produced from hen egg lysozyme using a monoclonal antibody. Immunity 6:727.[Medline]
  30. Deng, H., R. Apple, M. Clare-Salzler, S. Trembleau, D. Mathis, L. Adorini, E. Sercarz. 1993. Determinant capture as a possible mechanism of protection afforded by major histocompatibility complex class II molecules in autoimmune disease. J. Exp. Med. 178:1675.[Abstract/Free Full Text]
  31. Sercarz, E. E.. 2002. Processing creates the self. Nat. Immunol. 3:110.[Medline]
  32. Hill, S., E. E. Sercarz. 1975. Fine specificity of an immune response gene for the gallinaceous lysozymes. Eur. J. Immunol. 5:317.[Medline]
  33. Marrack, P., L. Ignatowicz, J. W. Kappler, J. Boymel, J. H. Freed. 1993. Comparison of peptides bound to spleen and thymus class II. J. Exp. Med. 178:2173.[Abstract/Free Full Text]
  34. Nelson, C. A., R. W. Roof, D. W. McCourt, E. R. Unanue. 1992. Identification of the naturally processed form of hen egg white lysozyme bound to the murine major histocompatibility complex class II molecule I-Ak. Proc. Natl. Acad. Sci. USA 89:7380.[Abstract/Free Full Text]
  35. Sette, A., S. Southwood, D. O’Sullivan, F. C. A. Gaeta, J. Sidney, H. Grey. 1992. Effect of pH on MHC class II-peptide interactions. J. Immunol. 148:844.[Abstract]
  36. Ayane, M., L. Mengle-Gaw, H.O. McDevitt, C. Benoist, D. Mathis. 1986. E{alpha}u and E{beta}u chain association: where lies the anomaly?. J. Immunol. 137:948.[Abstract]
  37. Pierres, M., F. M. Kourilsky, J. P. Rebouah, M. Dosseto, D. Caillol. 1980. Distinct epitopes on Ik gene products identified by monoclonal antibodies. Eur. J. Immunol. 10:950.[Medline]
  38. Pierres, M., C. Devaux, M. Doseeto, S. Marchetto. 1981. Clonal analysis of B- and T-cell responses to Ia antigens. I. Topology of epitope regions on I-Ak and I-Ek molecules analyzed with 35 monoclonal alloantibodies. Immunogenetics 14:481.[Medline]
  39. Germain, R. N., D. M. Bentley, H. Quill. 1985. Influence of allelic polymorphism on the assembly and surface expression of class II MHC (Ia) molecules. Cell 43:233.[Medline]
  40. Sant, A. J., N. S. Braunstein, R. N. Germain. 1987. Predominant role of amino-terminal sequences in dictating efficiency of class II major histocompatibility complex {alpha}{beta} dimer expression. Proc. Natl. Acad. Sci. USA 84:8065.[Abstract/Free Full Text]
  41. Vu, T. H., A. B. Begovich, F. M. Tacchini-Cottier, P. P. Jones. 1989. Molecular defects in the non-expressed H-2 Ea genes of the f and q haplotypes. J. Immunol. 142:2936.[Abstract]
  42. Tacchini-Cottier, F. M., P. P. Jones. 1988. Defective Eb expression in three mouse H-2 haplotypes results from aberrant RNA splicing. J. Immunol. 141:3647.[Abstract]
  43. Gilfillan, S., S. Aiso, S. A. Michie, H. O. McDevitt. 1990. Immune deficiency due to high copy number of an Abk transgene. Proc. Natl. Acad. Sci. USA 87:7319.[Abstract/Free Full Text]
  44. Boytim, M. L., P. Lilly, K. Drouvalakis, S. C. Lyu, R. Jung, A. M. Krensky, C. Clayberger. 2000. A human class II MHC-derived peptide antagonizes phosphatidylinositol 3-kinase to block IL-2 signaling. J. CIin. Invest. 105:1447.
  45. Kawai, K., P. S. Ohashi. 1995. Immunological function of a defined T-cell population tolerized to low-affinity self antigens. Nature 374:68.[Medline]
  46. Nishimoto, H., H. Kikutani, K. Yamamura, T. Kishimoto. 1987. Prevention of autoimmune insulitis by expression of I-E molecules in NOD mice. Nature 328:432.[Medline]
  47. Slattery, R. M., L. Kjer-Nielsen, J. Allison, B. Charlton, T. E. Mandel, J. F. A. P. Miller. 1990. Prevention of diabetes in non-obese diabetic I-Ak transgenic mice. Nature 345:724.[Medline]
  48. Gonzalez-Gay, M. A., G. H. Nabozny, M. J. Bull, E. Zanelli, J. Douhan, III, M. M. Griffiths, L. H. Glimcher, H. S. Luthra, C. S. David. 1994. Protective role of major histocompatibility complex class II Ebd transgene on collagen-induced arthritis. J. Exp. Med. 180:1559.[Abstract/Free Full Text]
  49. Christadoss, P., C. S. David, M. Shenoy, S. Keve. 1990. Ek {alpha} transgene in B10 mice suppresses the development of myasthenia gravis. Immunogenetics 31:241.[Medline]
  50. Rudensky, A. Y., P. Preston-Hurlburt, B. K. Al-Ramadi, J. Rothbard, C. A. Janeway, Jr. 1992. Truncation variants of peptides isolated from MHC class II molecules suggest sequence motifs. Nature 359:429.[Medline]
  51. Trembleau, S., G. Penna, E. Bosi, A. Mortara, M. K. Gately, L. Adorini. 1995. Interleukin 12 administration induces T helper type 1 cells and accelerates autoimmune diabetes in NOD mice. J. Exp. Med. 181:817.[Abstract/Free Full Text]



This article has been cited by other articles:


Home page
 J. Immunol.Home page
E. Martinez-Soria, M.-L. Santiago-Raber, L. Ho, T. Moll, and S. Izui
Protection of Murine Systemic Lupus by the Ea Transgene without Expression of I-E Heterodimers
J. Immunol., September 1, 2008; 181(5): 3651 - 3657.
[Abstract] [Full Text] [PDF]

Home page
 J. Immunol.Home page
N. K. Brown, D. J. McCormick, C. S. David, and Y.-c. M. Kong
H2E-Derived E{alpha}52-68 Peptide Presented by H2Ab Interferes with Clonal Deletion of Autoreactive T Cells in Autoimmune Thyroiditis
J. Immunol., May 15, 2008; 180(10): 7039 - 7046.
[Abstract] [Full Text] [PDF]

Home page
 BloodHome page
S. Kikuchi, H. Amano, E. Amano, L. Fossati-Jimack, M.-L. Santiago-Raber, T. Moll, A. Ida, B. L. Kotzin, and S. Izui
Identification of 2 major loci linked to autoimmune hemolytic anemia in NZB mice
Blood, August 15, 2005; 106(4): 1323 - 1329.
[Abstract] [Full Text] [PDF]

Home page
 J. Immunol.Home page
S. Kikuchi, L. Fossati-Jimack, T. Moll, H. Amano, E. Amano, A. Ida, N. Ibnou-Zekri, C. Laporte, M.-L. Santiago-Raber, S. J. Rozzo, et al.
Differential Role of Three Major New Zealand Black-Derived Loci Linked with Yaa-Induced Murine Lupus Nephritis
J. Immunol., January 15, 2005; 174(2): 1111 - 1117.
[Abstract] [Full Text] [PDF]

This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Martinez-Soría, E.
Right arrow Articles by Izui, S.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Martinez-Soría, E.
Right arrow Articles by Izui, S.

HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH