Superantigens in skin diseases

ARTICLE

The term superantigen has been introduced for proteins or glycoproteins that are able to activate large numbers of T cells (up to 30% of all T cells) without prior antigen processing [1, 2]. Therefore they differ from regular (nominal) antigens, which need processing by antigen presenting cells and activate only specific (clonotypic) T cells (usually less than 0,1% of all T cells). Nominal antigens (respectively the peptides in the binding groove of the MHC molecule) are recognized by all variable elements of the T cell receptor (V alpha, J alpha, Vß, Dß, Jß) and not only (or mainly) by the variable proportion of the ß chain like superantigens (compare Figure 1). Superantigens also have to be distinguished from mitogens, which activate T cells rather unspecifically and not only distinct subsets of them [1, 3, 4]. In analogy to the activation of whole subsets of T cells, certain compounds that bind to distinct heavy chain variable regions of immunoglobulins and thereby activate whole subsets of B cells have been termed immunoglobulin-superantigens.

Biological properties and immunological effects of superantigens

Sources of superantigens

Exogenous and endogenous (glyco-) proteins have been described as having superantigenic properties. Endogenous superantigens in mice such as Mls (minor lymphocyte stimulating antigens) are encoded by mouse mammary tumor proviruses (Mtv) carried in the germline of these mice. Their expression leads to the deletion of the reactive T cell populations in the affected strains of mice [5]. Endogenous superantigens have not been identified in humans until now. Exogenous superantigens are usually produced by microbial organisms, which are listed in Table I. However, some microbial products, which had been shown to have superantigenic properties failed to show such properties when they were highly purified or produced as recombinant molecules. This may indicate that contaminating proteins may be responsible for the superantigenic properties of such microbial products. Among these doubtful superantigens are staphylococcal proteins like exfoliative/epidermolytic toxin A (ETA) or streptococcal proteins like protein M. Both staphylococcal and streptococcal superantigens are strong activators of the immune system. This has been known since long before they were identified and designated as superantigens [6].

Molecular characteristics and ligands of superantigens

The following review will focus on the well-investigated staphylococcal superantigens, staphylococcal enterotoxin A-E, which are exotoxins secreted by Staphylococcus aureus for instance. Their molecular weight ranges between 26 and 28 kD (228-239 amino acids). They contain a disulfide loop and their structure is a ß-sheet with small alpha-helical parts [7]. They bind to MHC class II molecules with varying affinity [8] and they also bind to T cell receptors with certain variable elements of their ß-chain [3, 9, 10]. The various regions of T cell receptors are grouped into families according to homology on the amino acid level greater than 80%. Superantigens stimulate alphaß-T cells that express distinct Vß-elements of their T cell receptor as listed in Table II [9, 10]. Superantigens bind to the MHC class II molecules outside the regular antigen binding groove, but certain peptides inside the antigen binding grooves have been demonstrated to interfere with the binding of staphylococcal enterotoxins to MCH class II molecules and the binding of a superantigen may also inhibit the contact between a peptide-loaded MHC molecule to the T cell receptor. Furthermore, it has been demonstrated that zinc is involved in the binding of some staphylococcal enterotoxins to MHC class II molecules.

Effects of superantigens on antigen presenting cells

The binding of superantigens to MCH class II molecules on antigen presenting cells leads to the induction of signalling events and activation of cytokine production in antigen presenting cells like phosphoinositol breakdown, phospholipase C and protein kinase C activation, calcium-fluxes, induction of NF-kappaB and cytokine production like IL-1, IL-12 and TNF-alpha [11, 12].

Effects of superantigens on T cells ­ activation and expansion

The effects of superantigens on T cells depend very much on the surrounding conditions. Initially a strong proliferative response is induced in the superantigen-reactive Vß T cell subset. For this process binding to MHC class II molecules on antigen presenting cells is usually required [2], although it has been described that soluble superantigens may also have a mitogenic effect, especially when exogeneous monokines are added. Besides the direct interaction of the MHC class II molecule, the superantigen and the T cell receptor, additional co-stimulatory and adhesion factors are required for full T cell activation, like ICAM-1 and LFA-1, LFA-3 and CD2 and B7 and CD28, but the dependence on these factors may vary with the type of APC and T cell studied.

Besides proliferation, cytokine production is also induced in T cells. However, expression on the mRNA level has to be discriminated from real secretion of cytokines. Among the cytokines produced are TNF-alpha, IL-2 (mainly CD4⁺ T cells) and IFN-gamma (mainly CD8⁺ T cells [13]). IL-4 and IL-10 can be induced by SEB in the presence of IL-4, also leading to enhanced IgE production of B cells in vitro [14].

Effects of superantigens on T cells ­ inhibition of function (anergy, deletion and suppression)

Stimulation with superantigens does not necessary lead to activation and expansion of the reactive Vß⁺ T cells in the long run. Intraperitoneal or intravenous injections of SEB in the mg range leads to a rapid loss of surface expression of L-selectin (accompanied by initial activation and expansion of the reactive Vß⁺ T cells), while later a deletion especially of the CD4⁺ T cells can be observed [15]. In new born mice deletion of both CD4⁺ and CD8⁺ superantigen-reactive T cells occurs already in the thymus [1, 2]. Initial expansion of T cells (leading to "exhaustion") is not required for deletion of T cells later on, as repetitive application of small doses leads to deletion without initial proliferation. Despite the potency to prevent the induction of anergy, IL-2 is not able to prevent the deletion of T cells by superantigens in mice.

Deletion however is not the only negative immunological effect of superantigen exposure. Anergy and active suppression are also induced. Interestingly, TSST-1 is not able to induce anergy in CD4⁺ superantigen reactive T cell subsets like staphylococcal enterotoxins A and B [16]. Application of cyclohexamide together with SEB is able to prevent the induction of anergy, while it is not able to prevent subsequent deletion (apoptosis), indicating that induction of anergy is not a prerequisitive for later deletion. Additionally, anergy is not always followed by deletion, as cyclosporin A is able to prevent deletion of SEB reactive Vß⁺ T cells without preventing the induction of anergy. Deletion can also be prevented by pertussis toxin, while hydrocortisone enhances the deletion of T cells caused by superantigens. Induction of deletion and anergy by superantigens can be prevented by proper activation of the respective Vß⁺ T cells with their nominal antigen.

The third negative immunological effect of superantigens is the induction of active suppression, which can be achieved in vitro and in vivo and is mainly mediated by CD8⁺ T cells [17]. The induction of active suppressor cells can be prevented by pre-treatment with IL-12 and the suppressive effect of already induced suppressor cells can also be overcome by the treatment of recipient mice with IL-12.

Immunological effects of superantigens with special relevance to the skin

Effects of superantigens on T cells

The superantigenic staphylococcal enterotoxins and exfoliative toxin, but not mitogens, are able to induce cutaneous lymphocyte-associated antigen on human T cells (CLA, 12). This way even non-cutaneous exposure to superantigens may cause inflammation of the skin, as CLA is the special homing receptor of T cells for the skin.

Epicutaneous application or intradermal injection of SEB in ng amounts leads to a strong inflammatory response in the skin of mice. This response is dependent on the availability of SEB reactive T cells expressing the correct Vß⁺ elements, as mice who lack lymphocytes, T cells or certain Vß⁺ T cell subsets do not mount an inflammatory response at all or a much weaker response to cutaneous exposure to superantigens [18]. Dendritic epidermal T cells (DETC), which are gamma delta T cells, cannot substitute for the presence of SEB reactive alphaß T cells, indicating the strict Vß restriction of the SEB-effect. Cutaneous inflammation is followed by over-expression of SEB-reactive T cells among the increasing numbers of T cells in the local draining lymph nodes after cutaneous exposure to SEB [18]. Induction of anergy, suppression or deletion of Vß⁺ T cells can not be observed after a single exposure to SEB in the ng range [18]. In humans, epicutaneous application of SEB also induces an inflammatory response at the site of exposure [19].

In mice, repeated intradermal injections of low amounts or a single injection of larger amounts in the µg range of SEB lead to a functional down-regulation of SEB reactive Vß⁺ T cells being observable after about five days. Deletion of SEB-reactive T cells does not exceed 30% of the initial percentage of SEB reactive Vß⁺ T cell subsets, but allergic reactions depending on the functionality of the down-regulated Vß⁺ T cell subsets are strongly inhibited, concerning allergic reactions of the immediate type (ovalbumin) as well as of the delayed type (dinitrofluorbenzene).

Effects of superantigens on Langerhans cells/dendritic cells

Langerhans' cells are able to present superantigens to T cells in vitro with less UV-sensitivity than induction of T cells with nominal antigens, probably because antigen processing is not required. Superantigen exposure of skin sections leads to depletion of Langerhans' cells [20], probably due to their activation (as has also been observed in vivo [18]). When dendritic cells are used to present antigens to T cells the co-stimulatory pathway B7/CD28 seems to be involved. Dendritic cells are the most potent presenters of superantigens and bind up to 200-times more superantigen than B cells or monocytes.

Effects of superantigens on keratinocytes

Interferon-gamma-treated MHC class II⁺ keratinocytes are also able to present superantigens to T cells [21]. This process seems to be dependent on ICAM-1 and LFA-1 interactions but not on co-stimulation via CD28. Keratinocytes however are not only able to present superantigens to T cells, but they also are able to produce pro-inflammatory cytokines like TNF-alpha in response to superantigens [21] and express the adhesion molecule ICAM-1 on their surface, which may facilitate epidermal infiltration by immunocytes.

Potential clinical significance of superantigens to the field of dermatology

Superantigens may have clinical importance mainly in the field of autoimmunity and for inflammatory skin diseases. Autoimmune diseases may be triggered by activation and expansion of T cells that are able to cross-react with autoantigens [10]. Such mechanisms have been implicated for the initiation of acute guttate psoriasis (see below), rheumatoid arthritis and other autoimmune disorders [10].

Atopic dermatitis

Among inflammatory skin diseases the highest amounts of evidence for the importance of superantigens have been accumulated for atopic dermatitis and psoriasis. As demonstrated in mice, epicutaneous application or intradermal injection of superantigens, especially staphylococcal enterotoxins, induces inflammation of the skin [18]. This has also been demonstrated for humans, normal persons as well as patients with atopic dermatitis [19]. Therefore staphylococcal superantigens may cause acute exacerbations of atopic eczema, especially as staphylococci that are able to secret superantigenic enterotoxins can be frequently isolated from the skin of patients with atopic dermatitis [22]. Antibiotic treatment of atopic dermatitis is also often beneficial. Besides local exposure to superantigens, systemic exposure to superantigens may also up-regulate CLA on T cells, the homing receptor for skin T cells, leading to cutaneous inflammation [12]. In children with atopic dermatitis a higher proliferative response to SEB has also been demonstrated as being accompanied by higher IL-4 production, but lower interferon-gamma production compared to normal children. Lack of IFN-gamma production may be involved in insufficient irradication of Staphylococcus aureus from the skin of patients with atopic dermatitis [23]. Atopic eczema may also be aggravated by enhanced infiltration of the epidermis due to increased expression of ICAM-1 and TNF-alpha by keratinocytes after stimulation with superantigens [21, 24]. Another mechanism that may cause flare-up reactions of atopic eczema is induction of mast cell degranulation after cutaneous superantigen exposure in patients who have produced IgE specific for staphylococcal enterotoxins, which can be detected in the sera [25].

It has to be considered however that repeated or chronic exposure to superantigens may not only up-regulate immune responses, but may also inhibit immune responses, as superantigens are able to induce anergy, deletion of reactive T cells and active suppression as described above. This process may also be involved in insufficient irradication of Staphylococcus aureus in patients with atopic dermatitis as well as other pathogens or decreased reactions to contact allergens. Chronic superantigen exposure may also be responsible for decreased proliferative responsiveness to SEB of PMBC from atopic donors [26]. Although PBMC from atopic donors produce less interferon-gamma and IL-12 compared to normal subjects and more IL-4 and IL-5, staphylococcal antigens rather suppress IgE production [26]. The inhibitory effect of TSST-1 on IL-4-induced IgE production can be blocked by antibodies against interferon-gamma in normal donors, but not in atopic donors and can be blocked by antibodies against interferon-gamma in atopic donors, but not in normal donors. This indicates that the inhibitory effect of staphylococcal superantigens on IgE production may be dependent on T cell derived interferon-gamma in normal donors and on interferon-gamma derived from monocytes in atopic donors [27]. In the presence of IL-4 however staphylococcal superantigens are able to enhance IL-4 production and are able to spike IgE production [14]. Additionally, presentation of superantigens by keratinocytes instead of antigen presenting cells may also stimulate preferential IL-4 production due to the lack of IL-12 expression by keratinocytes.

Psoriasis vulgaris

Superantigens have been implicated in the pathogenesis of psoriasis [28]. Bacterial infections can also trigger this disease [29]. Of special relevance seem to be streptococcal infections of the throat and colonization of the skin with Staphylococcus aureus [29]. Streptococcus pyogenes lysates can induce increased keratinocyte proliferation after cutaneous injection and psoriatic lesions. As superantigens can cause skin inflammation and production of interferon-gamma, psoriatic reactions may result, as interferon-gamma is able to initiate the formation of such reactions. In a humanized SCID-mouse model the reconstitution of the mouse with immunocytes from psoriasis patients which have been stimulated with superantigens is able to provoke skin lesions after injection of superantigens into the skin that exhibit characteristics of psoriasis like epidermal hyperproliferation, papillomatosis, focal ICAM-1 expression and a dermal infiltration with T lymphocytes expressing the cutaneous lymphocyte associated antigen [30]. In humans a significant overexpression of Vß T cells that react with streptococcal pyogenes exotoxin C has been found in skin biopsies from patients with acute guttate psoriasis [31, 32]. In chronic plaque stage psoriasis, however, no constant pattern of Vß overexpression could be detected, although 20% of Vß T cell subsets present in the blood of individual donors were not present in the skin. This was true, however, not only for Vß T cell subsets but also for Valpha T cell subsets. This is consistent with the observation that the Vß repertoire of skin T cells can differ from that of the peripheral blood in healthy donors.

Lexicon of specific vocabularies

Superantigen (Proteinaceous) compound (usually of microbal origin) that activates a distinct subset of lymphocytes without the need for prior antigen processing

Vß Variable region of the ß chain of the alphaß T cell receptor. Grouped into families according to sequence homology (greater than 80% on the amino acid level)

Staphylococcal enterotoxins Exotoxins with superantigeneic properties secreted by (e.g.) Staphylococcus aureus

Streptococcus pyogenes toxins Superantigens secreted by (e.g.) streptococcus pyogenes

Abbreviations

APC ­ antigen presenting cell

CLA ­ cutaneous lymphocyte-associated antigen

ICAM ­ intercellular adhesion molecule

IFN ­ interferon

IL ­ interleukin

LFA ­ lymphocyte function-associated antigen

MHC ­ major histocompatibility complex

NF ­ nuclear factor

PBMC ­ peripheral blood mononuclear cells

(m)RNA ­ (messenger) ribonucleic assid

SE ­ staphylococcal enterotoxin

SPE ­ streptococcal pyogenes toxin

TCR ­ T cell receptor

TSST ­ toxic shock syndrome toxin

TNF ­ tumor necrosis factor

Vß ­ variable part of the ß chain of the TCR.

CONCLUSION

REFERENCES

1. White J, Herman A, Pullen AM, Kubo R, Kappler JW, Marrack P. The V beta-specific superantigen staphylococcal enterotoxin B: stimulation of mature T cells and clonal deletion in neonatal mice. Cell 1989; 56: 27-35.

2. Fleischer B, Schrezenmeier H. T cell stimulation by staphylococcal enterotoxins. Clonally variable response and requirement for major histocompatibility complex class II molecules on accessory or target cells. J Exp Med 1988; 167: 1697-707.

3. Kappler J, Kotzin B, Herron L, et al. V beta-specific stimulation of human T cells by staphylococcal toxins. Science 1989; 244: 811-3.

4. Tomai MA, Schlievert PM, Kotb M. Distinct T cell receptor V beta gene usage by human T lymphocytes stimulated with the streptococcal pyrogenic exotoxins and pep M5 protein. Infect Immun 1992; 60: 701-5.

5. Marrack P, Winslow GM, Choi Y, et al. The bacterial and mouse mammary tumor virus superantigens; two different families of proteins with the same functions. Immunol Rev 1993; 131: 79-92.

6. Donnelly RP, Rogers TJ. Immunosuppression induced by staphylococcal enterotoxin B. Cell Immunol 1982; 72: 166-77.

7. Micusan VV, Thibodeau J. Superantigens of microbial origin. Semin Immunol 1993; 5: 3-11.

8. Mollick JA, Cook RG, Rich RR. Class II MHC molecules are specific receptors for staphylococcus enterotoxin A. Science 1989; 244: 817-20.

9. Marrack P, Kappler J. The staphylococcal enterotoxins and their relatives. Science 1990; 248: 705-11.

10. Kotzin BL, Leung DY, Kappler J, Marrack P. Superantigens and their potential role in human disease. Adv Immunol 1993; 54: 99-166.

11. Chatila T, Geha RS. Signal transduction by microbial superantigens via MHC class II molecules. Immunol Rev 1993; 131: 43-59.

12. Leung DYM, Gately M, Trumble A, Ferguson-Darnell B, Schlievert P, Picker LJ. Bacterial superantigens induce T cell expression of the skin-selective homing receptor, the cutaneous lymphocyte-associated antigen, via stimulation of interleukin 12 production. J Exp Med 1995; 181: 747-53.

13. Herrmann T, Mac Donald HR. The CD8 T cell response to staphylococcal enterotoxins. Semin Immunol 1993; 5: 33-9.

14. Armerding D, van Reijsen FC, Hren A, Mudde GC. Induction of IgE and IgG1 in human B cell cultures with staphylococcal superantigens: role of helper T cell interaction, resistance to interferon-gamma. Immunobiology 1993; 188: 259-73.

15. Kawabe Y, Ochi A. Programmed cell death and extrathymic reduction of Vbeta8⁺ CD4⁺ T cells in mice tolerant to Staphylococcus aureus enterotoxin B. Nature 1991; 349: 245-8.

16. Hewitt CR, Lamb JR, Hayball J, Hill M, Owen MJ, O'Hehir RE. Major histocompatibility complex independent clonal T cell anergy by direct interaction of Staphylococcus aureus enterotoxin B with the T cell antigen receptor. J Exp Med 1992; 175: 1493-9.

17. Taub DD, Lin YS, Rogers TJ. Characterization and genetic restriction of suppressor-effector cells induced by staphylococcal enterotoxin B. J Immunol 1990; 144: 456-62.

18. Saloga J, Leung DYM, Reardon C, Giorno RC, Born W, Gelfand EW. Cutaneous exposure to the superantigen staphylococcal enterotoxin B elicits a T cell-dependent inflammatory response. J Invest Dermatol 1996; 106: 982-8.

19. Strange P, Skov L, Lisby S, Nielsen PL, Baadsgaard O. Staphylococcal enterotoxin B applied on intact normal and intact atopic skin induces dermatitis. Arch Dermatol 1996; 132: 27-33.

20. Shankar G, Pickard ES, Burnham K. Superantigen-induced Langerhans cell depletion is mediated by epidermal cell-derived IL-1 alpha and TNF alpha. Cell Immunol 1996; 171: 240-5.

21. Tokura Y, Yagi J, O'Malley M, et al. Superantigenic staphylococcal exotoxins induce T cell proliferation in the presence of Langerhans cells or class II-bearing keratinocytes and stimulate keratinocytes to produce T cell-activating cytokines. J Invest Dermatol 1994; 102: 31-8.

22. McFadden JP, Noble WC, Camp RD. Superantigenic exotoxin-secreting potential of staphylococci isolated from atopic eczematous skin. Br J Dermatol 1993; 128: 631-2.

23. Campbell DE, Kemp AS. Proliferation and production of interferon-gamma and IL-4 in response to staphylococcus aureus and staphylococcal superantigen in childhood atopic dermatitis. Clin Exp Immunol 1997; 107: 392-7.

24. Wakita H, Tokura Y, Furokawa F, Takigawa M. Staphylococcal enterotoxin B upregulates expression of ICAM-1 molecules on IFN-gamma-treated keratinocytes and keratinocyte cell lines. J Invest Dermatol 1995; 105: 536-42.

25. Leung DY, Harbeck R, Bina P, et al. Presence of IgE antibodies to staphylococcal exotoxins on the skin of patients with atopic dermatitis. Evidence for a new group of allergens. J Clin Invest 1993; 92: 1374-80.

26. König B, Neuber K, König W. Responsiveness of peripheral blood mononuclear cells from normal and atopic donors to microbal superantigens. Int Arch Allergy Immunol 1995; 106: 124-33.

27. Lester MR, Hofer MF, Renz H, Trumble AE, Gelfand EW, Leung DYM. Modulatory effects of staphylococcal superantigen TSST-1 on IgE synthesis in atopic dermatitis. Clin Immunol Immunopathol 1995; 77: 332-8.

28. Leung DY, Walsh P, Giorno R, Norris DA. A potential role for superantigens in the pathogenesis of psoriasis. J Invest Dermatol 1993; 100: 225-8.

29. Whyte JH, Baughman RD. Acute guttate psoriasis and streptococcal infection. Arch Dermatol 1964; 89: 350-6.

30. Boehncke WH, Zollner TM, Dressel D, Kaufmann R. Induction of psoriasiform inflammation by a bacterial superantigen in the SCID-hu xenogeneic transplantation model. J Cutan Pathol 1997; 24: 1-7.

31. Leung DY, Travers JB, Giorno R, Norris DA, Skinner R, Aelion J, Kazemi LV, Kim MH, Trumble AE, Kotb M, et al. Evidence for a streptococcal superantigen-driven process in acute guttate psoriasis. J Clin Invest 1995; 96: 2106-12.

32. Lewis HM, Baker BS, Bokth S, et al. Restricted T cell receptor V beta gene usage in the skin of patients with guttate and chronic plaque psoriasis. Br J Dermatol 1993; 129: 514-20.