Biomedical subjects
Publications and source records attributed to L Adorini.
Experimental models of autoimmune diseases have demonstrated that such disease can be prevented or treated by selectively interfering with activation of any of these cell types: antigen-presenting cells, autoreactive T cells and regulatory T cells. Luciano Adorini and colleagues discuss these approaches to selective immunosuppression and examine how similar strategies may become applicable to the treatment of human autoimmune diseases.
Synthetic peptides spanning the entire sequence of both human and mouse beta 2-microglobulin (beta 2M) have been tested for their capacity to bind to three different mouse (I-Ad, I-Ed, and I-Ak) or human (DR1, DR2, and DR5) class II molecules. The results demonstrate that class II molecules do not discriminate between self and non-self peptides. When the immunogenicity of the human beta 2M peptides was measured by their ability to prime H-2d mice for in vitro T cell proliferation, it was found that peptides incapable of binding class II molecules in vitro were also non-immunogenic in vivo. Interestingly, however, several binders, including the human beta 2M peptide 1-16, the best binder in this series to Iad molecules, were found to be non-immunogenic. Since the corresponding mouse beta 2M peptide 1-16 was also capable of binding to Iad molecules, this suggested that lack of responsiveness to the non-self peptide could arise either from central or peripheral tolerance induced by the self homolog. Alternatively, lack of responsiveness could arise from other mechanisms, such as negative selection by other non-homolog sequences or lack of suitable T cell receptor genes. To discriminate between these possibilities, H-2d mice with disrupted beta 2M genes were immunized with the human beta 2M peptide 1-16. This peptide also failed to prime for T cell responsiveness in beta 2M-negative mice, suggesting that a hole in the T cell repertoire for this antigen was not mediated by negative selection or peripheral tolerance induced by self beta 2M peptides.
Multiple sclerosis (MS) is an autoimmune disease in which myelin proteins have been implicated as autoantigens recognized by pathogenic autoreactive T cells. To study the relationship between human myelin basic protein (hMBP) and HLA alleles associated to MS susceptibility, such as DRB1*1501, the binding of synthetic peptides spanning the entire hMBP sequence to 10 purified HLA-DR molecules was determined. All the hMBP peptides tested showed binding affinity for at least one of the DR molecules analyzed, but three hMBP peptides, included in sequences 13-32, 84-103, and 144-163 were found capable of binding to three or more DR molecules. The hMBP peptide 84-103 was the most degenerate in binding, in that it bound to 9 out of 10 DR molecules tested. Interestingly, it bound with highest affinity to DRB1*1501 molecules. To correlate the binding pattern of hMBP peptides to HLA class II molecules with their recognition by T cells, 61 hMBP-specific T cell lines (TCL) were established from the peripheral blood of 20 MS patients, who were homozygous, heterozygous, or negative for DRB1*1501. Analysis of hMBP epitopes recognized by these TCL and their HLA restriction demonstrated a very good correlation between binding data and T cell proliferation to hMBP peptides. Although virtually all hMBP peptides tested could be recognized by at least one TCL from MS patients, three immunodominant T cell epitopes were apparent among the TCL examined, corresponding exactly to the hMBP peptides capable of binding to several DR molecules. No major difference could be detected in the recognition of immunodominant hMBP peptides by TCL from DRB1*1501 positive or negative MS patients. These results have implications for the role of hMBP as relevant autoantigen, and of DRB1*1501 as susceptibility allele in MS.
Administration of major histocompatibility complex (MHC) class II-binding synthetic peptides can induce selective immunosuppression via different mechanisms of action. In this review four different ways to induce selective immunosuppression by peptide administration will be examined: MHC blockade, T cell receptor (TCR) antagonism, induction of peripheral tolerance, and activation of regulatory cells. These approaches to selective immunosuppression target three types of cells: antigen-presenting cells, antigen-specific T cells, and regulatory T cells. Understanding these forms of immunosuppression will provide new insights in basic immunology and may offer strategies for selective immunointervention in autoimmune diseases, allograft rejection, and allergy.
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Draining lymph node cells (LNC) from mice immunized with hen egg white lysozyme (HEL) display at their surface antigen-MHC complexes able to stimulate, in the absence of any further antigen addition, HEL peptide-specific, class II-restricted T cell hybridomas. Chloroquine addition to these LNC cultures fails to inhibit antigen presentation, indicating that antigenic complexes of class II molecules and HEL peptides are formed in vivo. MHC class II restriction of antigen presentation by LNC from HEL-primed mice was verified by the use of anti-class II monoclonal antibodies. Coinjection of HEL and the I-Ak-binding peptide HEL 112-129 in mice of H-2k haplotype inhibits the ability of LNC to stimulate I-Ak-restricted, HEL 46-61-specific T cell hybridomas. Similar results are obtained in mice coinjected with the HEL peptides 46-61 and 112-129. Inhibition of T hybridoma activation can also be observed using as antigen-presenting cells irradiated, T cell-depleted LNC from mice coinjected with HEL 46-61 and HEL 112-129, ruling out the possible role of either specific or nonspecific suppressor T cells. Inhibition of T cell proliferation is associated with MHC-specific inhibition of antigen presentation and with occupancy by the competitor of class II binding sites, as measured by activation of peptide-specific T cell hybridomas. These results demonstrate that administration of MHC class II binding peptide competitors selectively inhibits antigen presentation to class II-restricted T cells, indicating competitive blockade of class II molecules in vivo.
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Autoimmune diseases result from the activation of self-reactive T cells induced by autoantigens or by foreign antigens cross-reactive with an autoantigen. A striking characteristic of autoimmune diseases is the increased frequency of certain HLA alleles in affected individuals. Moreover, as demonstrated for example in rheumatoid arthritis and insulin-dependent diabetes mellitus, class II alleles positively associated with autoimmune diseases share amino acid residues in the hypervariable HLA regions involved in peptide binding. Therefore, it is likely that disease-associated HLA class II molecules have the capacity to bind the autoantigen and present it to T cells, thereby inducing and maintaining, under appropriate conditions, the autoimmune disease. The data reviewed here demonstrate MHC-selective inhibition of antigen-induced T cell responses in vivo by parenterally administered soluble, MHC-binding peptide competitors, under conditions in which the competitor is not immunogenic. This suggests the feasibility of a therapeutic approach based on blockade of MHC class II molecules in the treatment of HLA-linked autoimmune diseases.
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Class II MHC molecules on the surface of an APC present immunogenic peptides derived mainly from exogenous proteins to CD4+ T cells. During its transport to the cell surface, class II molecules intersect the endocytic pathway where they acquire peptides derived from endocytosed proteins. However, class II-restricted presentation of endogenously derived peptides can also occur. The current studies were undertaken to examine the ability of different types of APC to generate and present four different T cell determinants derived from an endogenous, nonsecreted, truncated form of hen-egg white lysozyme (HEL[1-80]-Kk). This was compared with the ability of these APC to generate the same determinants from exogenous HEL. All the peptides derived from endogenous HEL[1-80]-Kk tested, were presented by B cells to HEL-specific T cell hybridomas with an efficiency similar to presentation of the same determinants from exogenous HEL. In contrast, an I-Ak-bearing rat fibroblast was unable to generate the HEL peptide 25-43 from exogenous HEL, but could efficiently produce it from endogenous HEL[1-80]-Kk. The results indicate first, that peptides derived from an endogenous Ag can be presented by MHC class II molecules with an efficiency comparable to that of the presentation of the exogenous Ag. Second, that Ag-presenting B cells can generate the same repertoire of antigenic peptides from endogenous Ag as those generated from the exogenous protein. And third, that in contrast to B cells, certain "nonprofessional" APC can generate, from an endogenous protein, T cell determinants distinct from those generated after endocytosis of the exogenous protein. These results suggest that processing of exogenous and endogenous Ag by different APC take place in different intracellular compartments.
Antigen-presenting cells (APC) transfected with a construct encoding the hen egg-white lysozyme (HEL) amino acid sequence 1-80 constitutively present HEL peptides complexed to major histocompatibility complex (MHC) class II molecules to specific T cell hybridomas, indicating that endogenous cellular antigens can be efficiently presented to class II-restricted T cells. Here we show that exogenous peptide competitors added to HEL-transfected APC can inhibit the presentation of endogenous HEL peptides to class II-restricted T cells. The inhibition is specific for the class II molecule binding the competitor peptide, and it affects to the same extent presentation of exogenous or endogenous HEL peptides. These results, demonstrating that an exogenous competitor can inhibit class II-restricted T cell activation induced by endogenous as well as exogenous antigen, suggest lack of strict compartmentalization between endogenous and exogenous pathways of antigen presentation. Since autoreactive T cells may recognize endogenous, as well as exogenous antigens, the results have implications for the treatment of autoimmune diseases by MHC blockade.
We have analyzed the interaction of the hen egg-white lysozyme (HEL) peptide 107-116 with the MHC class II molecule I-Ek, using truncated and single residue substitution analogues to measure activation of I-Ek-restricted, 107-116-specific T cell hybridomas and competition for Ag presentation by I-Ek molecules. These results have been compared with previous findings on the interaction of the same peptide with the I-Ed molecule. Stimulation of T cell hybridomas by truncated peptides defines the sequence 108-116 as the minimum epitope necessary for activation of both I-Ek- and I-Ed-restricted T cell hybridomas. Substitution analysis pinpoints three residues (V109, A110, and K116) in the sequence 108-116 as being critical for binding to I-Ek molecules and demonstrates the involvement of most other residues in recognition by T cells. Results previously obtained for binding of HEL 107-116 to I-Ed molecules indicated that peptide residues R112, R114, and K116 were critical for interaction with I-Ed. Comparison of these results indicates a difference in the likely MHC contact residues between the HEL sequence 108-116 and I-Ed or I-Ek molecules, suggesting that the same HEL peptide assumes a different conformation in the binding site of these two MHC molecules. This in turn affects residues interacting with the specific T cell receptor. According to the hypothetical tridimensional structure predicted for class II molecules, the difference in MHC contact residues observed within the sequence 108-116 can be related to polymorphic amino acids in the binding site of I-Ek and I-Ed molecules. A search through published binding data for a common pattern in this and other I-Ek-binding peptides has permitted us to derive a possible motif for predicting peptide binding to I-Ek molecules. This putative motif was tested by determining binding to I-Ek of an unbiased panel of about 150 synthetic peptides. Binding data indeed demonstrate the presence of this motif in the majority of good binders to I-Ek molecules.
The influence of the class II-associated invariant chain (Ii) on the presentation of the protein antigens hen egg-white lysozyme (HEL) and ribonuclease A (RNase) was investigated. For this purpose the Ii- rat-2 fibroblasts were transfected with I-Ak genes with or without Ii. Transfectants expressing Ii were superior in the presentation of the complete HEL protein to a panel of I-Ak-restricted T hybridomas characterized by distinct specificities for different HEL peptides and by different sensitivities to antigen concentration. There appeared to be a correlation between the antigen-presenting capacity and the amount of Ii, in that transfectants expressing large amounts of Ii were the best antigen presentors. The presentation of synthetic HEL peptides was not influenced by Ii. In contrast to the findings with HEL, the presentation of RNase by the same set of transfectants was clearly independent of Ii. Both antigens, HEL and RNase, required processing in the chloroquine-sensitive compartment. However, only the presentation of HEL but not of RNase could be efficiently blocked by brefeldin A. These data confirm that presentation of HEL depends on de novo synthesized class II molecules, whereas the presentation of RNase seems to be predominantly mediated by a pool of pre-existing class II molecules whose interaction with endocytosed antigen does not depend on Ii. These results suggest different mechanisms for the presentation of HEL and RNase and they raise the possibility that different antigens intersect the class II pathway at distinct intracellular locations.
We have sought to determine how much amino acid diversity is tolerable at position 69 of the Ak alpha chain, a position previously implicated as a peptide contact site. Slot-machine mutagenesis was used to create a set of 11 mutant Ak alpha cDNAs, each specifying a different amino acid at position 69. These cDNAs were individually expressed in L cells together with a wild-type Ak beta cDNA to produce a panel of mutant antigen-presenting cell lines. The ability of each member of this panel to present a hen egg lysozyme and a bovine ribonuclease peptide to various T hybridomas was assessed. We found that a surprising degree of amino acid diversity is tolerable at Ak alpha position 69: even charged (Glu, Arg) or bulky (Trp, Tyr) residues can be accommodated without abrogating cell-surface expression of Ak, peptide binding to it, or T cell recognition of it. We discuss the implications of these findings for models of T cell recognition of the class II molecule/antigen duplex.
Autoimmune diseases result from the activation of self-reactive T cells induced by autoantigens or by foreign antigens cross-reactive with an autoantigen. A striking characteristic of autoimmune diseases is the increased frequency of certain HLA alleles in affected individuals. Moreover, as demonstrated for example in rheumatoid arthritis and insulin-dependent diabetes mellitus, class II alleles positively associated with autoimmune diseases share amino acid residues in the hypervariable HLA regions involved in peptide binding. Therefore, it is likely that disease-associated HLA class II molecules have the capacity to bind the autoantigen and present it to T cells, thereby inducing and maintaining, under appropriate conditions, the autoimmune disease. The data reviewed here demonstrate MHC-selective inhibition of antigen-induced T cell responses in vivo by parenterally administered soluble, MHC-binding peptide competitors, under conditions in which the competitor is not immunogenic. This suggests the feasibility of a therapeutic approach based on MHC blockade in the treatment of HLA-linked autoimmune diseases.
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A synthetic peptide corresponding to residues 53-61 of hen egg white lysozyme, as well as its N-succinyl C-amide, inhibit in vivo priming for T cell responses when administered in soluble form, in osmotic minipumps implanted s.c., 1 day before immunization. The inhibition exhibits MHC selectivity that corresponds to the binding specificity of these peptides for class II MHC molecules in vitro. Approximately fourfold higher doses of soluble competitor are required than of competitor in CFA-depot, in order to achieve comparable levels of inhibition. The inhibition of priming to a nonimmunodominant T cell determinant requires lower doses of soluble competitor than the suppression of priming to a dominant T cell determinant. The soluble competitors do not appear to induce class II MHC-restricted T cell response against themselves.
Peptides bound to class I or class II major histocompatibility complex (MHC)-encoded molecules are ligands for the antigen-specific T-cell receptor of T-cells carrying the CD8 and CD4 antigens, respectively. MHC class I-restricted T cells generally recognize peptides derived from processing of endogenously synthesized cellular antigens, whereas class II-restricted T cells usually recognize peptides derived from exogenous antigens entering antigen presenting cells. Accordingly, two separate pathways of antigen processing and presentation have been proposed. The fungal metabolite brefeldin A (BFA), an inhibitor of protein transport from the endoplasmic reticulum, inhibits presentation of endogenous antigens for MHC-restricted T-cell recognition. The selectivity of BFA activity has been inferred to reflect presentation of a given antigen processed through the cytosolic or the endocytic route. Here we show that BFA also greatly inhibits the presentation of exogenous protein antigens by MHC class II molecules to T cells, indicating a broader effect of this drug on antigen presentation and an additional similarity between the two processing pathways. As cycloheximide, a protein synthesis inhibitor, also inhibits presentation of protein antigens to class II-restricted T cells, the data indicate that peptides generated by processing of exogenous proteins binds to newly synthesized class II molecules for presentation to T cells.