Stephen M. Anderton

Do heat shock proteins control the balance of T-cell regulation in inlammatory diseases?

Abstract Heat shock proteins (Hsps) are remarkably immunogenic, despite their high degree of evolutionary conservation. Experimental and clinical observations on autoimmune diseases indicate that immune responses to Hsps arise spontaneously during the disease process. Based on current evidence, Willem van Eden and colleagues argue that such immunity to Hsps is part of a normal immunoregulatory T-cell response with disease controlling potential.

Hierarchy in the ability of T cell epitopes to induce peripheral tolerance to antigens from myelin

Nasal administration of peptide antigens has been shown to induce T cell tolerance. We have investigated the potential for peptide therapy of the autoimmune response to myelin antigens in experimental autoimmune encephalomyelitis (EAE). Three major encephalitogenic epitopes were studied for their ability to induce nasal tolerance to myelin antigens. These included epitopes Ac1 – 9 and 89 – 101 of myelin basic protein (MBP) and 139 – 151 from proteolipid protein (PLP). Peptide Ac1 – 9 from MBP effectively suppressed responses to both MBP epitopes, following immunization with whole myelin (linked suppression). The N‐terminal epitope failed, however, to modify the response to epitope 139 – 151 of PLP. The second MBP epitope (89 – 101) was poorly tolerogenic for the immune response to any naturally processed myelin epitope. By contrast, PLP[139 – 151] was able to induce bystander suppression of T cells responsive to both itself and the two epitopes from MBP. Furthermore, this epitope suppressed EAE induced with peptides derived from MBP and was capable of treating ongoing disease. The mechanism of bystander suppression, mediated by PLP[139 – 151], did not correlate with an overt switch from the T helper 1 to the T helper 2 phenotype. These results demonstrate how a complex autoimmune disease may be controlled by treatment with a single peptide epitope and reveal a hierarchy in the suppressive properties of different myelin antigens.

Mechanisms of central and peripehral T‐cell tolerance: lessons from experimental models of multiple sclerosis

Summary: Studies of experimental autoimmune encephalomyelitis in conventional and transgenic mouse models have clarified mechanisms of central and peripheral tolerance to myelin antigens. It is now clear that myelin antigens are expressed in the thymus and their expression can influence generation of the potential autoimmune T‐cell repertoire. How autoreactive T cells escape tolerance in the thymus is largely unclear. One mechanism has been revealed through the use of a transgenic mouse expressing a T‐cell receptor specific for a myelin antigen. T cells specific for the N‐terminal epitope of myelin basic protein escape tolerance through low avidity interaction. The affinity of antigen binding to MHC also proves to be important for induction of peripheral tolerance. Peptides may be administered in solution to adults in order to reinstate peripheral tolerance and suppress disease. Induction of antigen‐specific suppression with synthetic peptides can result in either linked or bystander suppression and this appears to involve the generation of T cells secreting suppressive cytokines. The use of altered peptide ligands fur induction of peripheral tolerance has been investigated. This can be achieved but the complexity of the approach argues against its use for treatment of human autoimmune diseases.

Therapeutic potential of TCR antagonists is determined by their ability to modulate a diverse repertoire of autoreactive T cells

The use of altered peptide ligands (APL) with TCR antagonist properties holds promise as an antigen‐specific therapy for autoimmune disorders. We are investigating the therapeutic potential of APL in experimental autoimmune encephalomyelitis (EAE) using the Ac1 – 9 peptide of myelin basic protein in H‐2u mice. Encephalitogenic T cells recognize Ac1 – 9 using residues 3Gln and 6Pro as the major TCR contact sites. Use of position 6 APL is compromised by the heterogeneous nature of the Ac1 – 9‐specific repertoire. Here we identify two position 3 APL that act as TCR antagonists on transgenic T cells expressing Ac1 – 9‐specific TCR and that inhibit EAE in H‐2u mice. However, the therapeutic capacity of these two APL correlated directly with the ability to maximally inhibit activation of a heterogeneous T cell pool. The implications of these findings for the requirements for EAE induction, the relative contribution of a given T cell subpopulation to pathology and the mechanism underlying EAE inhibition in this model are discussed.

Altered) Self Peptides and the Regulation of Self Reactivity in the Peripheral T cell Pool

Tolerance for self has appeared incomplete for many self antigens. We have obtained experimental evidence that both for self heat shock proteins and T cell receptor V-gene products, reactive T cells are part of the normal immune repertoire. Furthermore, it has become apparent that stimulation of T cell responsiveness to these antigens, by using peptide immunisation or by transfer of activated T cells, raises resistance to experimentally induced autoimmune arthritis. In addition, available evidence has suggested that these reactivities may be functional during natural processes of disease remission. The observations with regard to heat-shock proteins have indicated that mechanism leading to disease resistance are most efficiently triggered by exposing the immune system to non-self antigens such as bacterial hsps, which are similar to, but not identical to, self. Experimental evidence has been obtained, that conserved bacterial hsp peptides, may trigger self hsp reactive T cells, with disease suppressive regulatory potential. It is possible that such self hsp reactive T cells, being expanded by recognising bacterial peptides as full agonists, do, in fact, perceive the self epitopes as partial agonists, and therefore have the possibility of displaying downregulatory activity at the site of inflammation. Experiments with peptide analogs of self epitopes, being variants of disease critical T cell epitopes, have indeed suggested that also their activity in modulating disease may comply with the principles of dominant immunological tolerance.

Kinetics of Peptide Uptake and Tissue Distribution Following a Single Intranasal Dose of Peptide

Stimulation with the combination of PDB plus ionomycin induced LFA-l/ICAM-1-dependent homotypic adhesion of tonsil B cells. Adhesion of tonsil B cells in our system induced tyrosine phosphorylation of Pyk-2. Disruption of homotypic adhesion and concomitant inhibition of induction of protein tyrosine phosphorylation was achieved by physical separation of the cells and by treatment with methyl-2,5-dihydroxycinnamate (MDHC), an inhibitor of protein tyrosine phosphorylation. These results suggest that protein tyrosine phosphorylation that is associated with homotypic adhesion is mediated by LFA-l/ICAM-1 interactions.

Tolerance to an Arthritogenic T‐cell Epitope of HSP65 and the Regulation of Experimental Arthritis

Adjuvant arthritis (AA) is an extensively studied form of experimental arthritis resembling rheumatoid arthritis in a number of pathological aspects. It can be induced in susceptible (Lewis) rats by immunization with mycobacterial antigens. From several experiments it can be concluded that T-cell responses to hsp’s play an important immunomodulatory role in the induction AA. First it was shown that AA can be passivily transfered by T cells alone from diseased rats to syngeneic disease-free animals.’ Fuxtherniore passive transfer of a T-cell clone recognizing the nonconserved 180188 amino acid sequence in mycobacterial hsp65 was found to induce AA.z3 This T-cell clone also responded to cartilage proteoglycan but not to rat hsp60. The clone therefore showed tliat although self-cross-reactive or “mimicry” T cells are able to induce overt autoimmune disease, this was not related to the conserved nature of hsp’s. Several studies have shown that it is possible to induce antigen-specific T-cell tolerance in experimental autoimmune models! Most evidence so far has been collected in the model of experimental allergic encephalomyelitis (EAJZ). EAE is a demyelinating autoimmune disease caused by CD4’ T cells specific for myelin basic protein (MBP). It can be induced in susceptible animals by immunization with MBP in CFA. However, oral administration of MBP protects animals from developing E M . Protection can be established by adoptive transfer of CD8+ T cells, capable of producing TGF-P when stimulated with the relevant antigen^.^ Encephalogenic epitopes of MBP have been characterized, and nasal inhalation of the immunodominant epitope on MBP has also led to protection. In the present study we investigated whether tolerance could be induced similarly to the AA-related immunodominant epitope and whether this would lead to protection against AA, a disease induced by an antigen as complex as whole mycobacteria. Two 15-mer peptides containing the individual mycobacterial hsp65 sequences 176190 (M36) and 211-225 (M43) were used. M36 contains the epitope 180-188 recognized by the arthritogenic T-cell clone, A2b. Furthermore, it is the immunodominant T-cell epitope after induction of arthritis in AA.6 M43 is a codominant epitope both after induction

Antigen-Specific Tolerance Induction and the Immunotherapy of Experimental Autoimmune Disease

Antigen-specific tolerance induction is the ultimate goal for specific immunotherapy of autoimmune diseases. Here we will discuss recent experiments designed to induce tolerance following mucosal administration of antigens in a mouse model of experimental autoimmune encephalomyelitis (EAE). We were unable to induce oral tolerance either with whole myelin, myelin basic protein (MBP) or the immunodominant peptide antigen. Oral tolerance was possible, however, with an analogue of the immunodominant peptide modified to increase its affinity for the restricting major histocompatibility complex (MHC) antigen. By contrast, intranasal deposition of peptide antigen proved highly effective for both prevention and treatment of EAE. Prevention of disease was directly related to the antigenic property of the peptide which, in itself, was related to affinity for MHC. Notably, administration of a single peptide was shown to inhibit disease involving multiple epitopes. We investigated the resulting bystander regulation by studying the cellular basis of peripheral tolerance in a transgenic model. These studies indicate that bystander regulation may be the consequence of selective cytokine secretion.

Bacterial Heat-Shock Proteins and Autoimmune Disease

Of all the microorganisms, bacteria, especially, seem to exert continuous pressure on the immune system. The epithelial surfaces of the skin and the intestinal tract are colonized by a wide variety of bacterial species, forcing the immune system into a situation where it has to scan continuously an immense repertoire of foreign antigenic determinants. It may be self-evident that as much as the healthy immune system does not develop an aggressive response directed against self antigens, or autoantigens, the same immune system is not tuned to mount continuous and aggressive responses to antigens present in the resident bacterial flora. Nonetheless, the capacity to respond must be there. Within every healthy immune system, cells specific for autoantigens are present, and their potential for causing disease has been demonstrated in a variety of experimental autoimmune disease models. The mechanisms by which the immune