Staphylococcal Enterotoxin C2 Mutant-Induced Antitumor Immune Response Is Controlled by CDC42/MLC2-Mediated Tumor Cell Stiffness.

As a biological macromolecule, the superantigen staphylococcal enterotoxin C2 (SEC2) is one of the most potent known T-cell activators, and it induces massive cytotoxic granule production. With this property, SEC2 and its mutants are widely regarded as immunomodulating agents for cancer therapy. In a previous study, we constructed an MHC-II-independent mutant of SEC2, named ST-4, which exhibits enhanced immunocyte stimulation and antitumor activity. However, tumor cells have different degrees of sensitivity to SEC2/ST-4. The mechanisms of immune resistance to SEs in cancer cells have not been investigated. Herein, we show that ST-4 could activate more powerful human lymphocyte granule-based cytotoxicity than SEC2. The results of RNA-seq and atomic force microscopy (AFM) analysis showed that, compared with SKOV3 cells, the softer ES-2 cells could escape from SEC2/ST-4-induced cytotoxic T-cell-mediated apoptosis by regulating cell softness through the CDC42/MLC2 pathway. Conversely, after enhancing the stiffness of cancer cells by a nonmuscle myosin-II-specific inhibitor, SEC2/ST-4 exhibited a significant antitumor effect against ES-2 cells by promoting perforin-dependent apoptosis and the S-phase arrest. Taken together, these data suggest that cell stiffness could be a key factor of resistance to SEs in ovarian cancer, and our findings may provide new insight for SE-based tumor immunotherapy.

Identification of a novel fully human anti-toxic shock syndrome toxin (TSST)-1 single-chain variable fragment antibody averting TSST-1-induced mitogenesis and cytokine secretion.

BACKGROUND: Staphylococcal superantigens are virulence factors that help the pathogen escape the immune system and develop an infection. Toxic shock syndrome toxin (TSST)-1 is one of the most studied superantigens whose role in toxic shock syndrome and some particular disorders have been demonstrated. Inhibiting TSST-1 production with antibiotics and targeting TSST-1 with monoclonal antibodies might be one of the best strategies to prevent TSST-1-induced cytokines storm followed by lethality. RESULTS: A novel single-chain variable fragment (scFv), MS473, against TSST-1 was identified by selecting an scFv phage library on the TSST-1 protein. The MS473 scFv showed high affinity and specificity for TSST-1. Moreover, MS473 could significantly prevent TSST-1-induced mitogenicity (the IC50 value: 1.5 µM) and cytokine production. CONCLUSION: Using traditional antibiotics with an anti-TSST-1 scFv as a safe and effective agent leads to deleting the infection source and preventing the detrimental effects of the toxin disseminated into the whole body.

Design of split superantigen fusion proteins for cancer immunotherapy.

Several antibody-targeting cancer immunotherapies have been developed based on T cell activation at the target cells. One of the most potent activators of T cells are bacterial superantigens, which bind to major histocompatibility complex class II on antigen-presenting cells and activate T cells through T cell receptor. Strong T cell activation is also one of the main weaknesses of this strategy as it may lead to systemic T cell activation. To overcome the limitation of conventional antibody-superantigen fusion proteins, we have split a superantigen into two fragments, individually inactive, until both fragments came into close proximity and reassembled into a biologically active form capable of activating T cell response. A screening method based on fusion between SEA and coiled-coil heterodimers was developed that enabled detection of functional split SEA designs. The split SEA design that demonstrated efficacy in fusion with coiled-coil dimer forming polypeptides was fused to a single chain antibody specific for tumor antigen CD20. This design selectively activated T cells by split SEA-scFv fusion binding to target cells.

Staphylococcal superantigen-like 12 induces the production of interleukin 4 in murine basophils.

S. aureus is associated with atopic dermatitis (AD). Several staphylococcal products including cell wall components, protease, and exotoxins, are thought to be involved in allergic inflammation of AD via activating immune cells such as T cells and mast cells. None of the staphylococcal exotoxins has been reported to activate a primary IL-4 inducer, basophils, that are known to produce large amounts of IL-4 in response to allergens as well as IgE-independent stimuli such as mites and helminth proteases. In this study, we investigated the ability of staphylococcal superantigen-like (SSL) family to activate basophils. SSL12, reported its activity to activate mast cells, induced the production of IL-4 in bone marrow derived basophils. SSL12 also evoked the release of IL-4 in freshly isolated murine basophils in bone marrow cells, as the depletion of basophils by basophils-specific antibodies against high-affinity IgE receptor and CD49b diminished the responsiveness of bone marrow cells for SSL12. These results propose the novel immune regulatory activity of SSL12 by inducing IL-4 in basophils, that contributes to the development of allergic inflammation disorders and the immune evasion of the cocci.

The Staphylococcus aureus superantigen SElX is a bifunctional toxin that inhibits neutrophil function.

Bacterial superantigens (SAgs) cause Vß-dependent T-cell proliferation leading to immune dysregulation associated with the pathogenesis of life-threatening infections such as toxic shock syndrome, and necrotizing pneumonia. Previously, we demonstrated that staphylococcal enterotoxin-like toxin X (SElX) from Staphylococcus aureus is a classical superantigen that exhibits T-cell activation in a Vß-specific manner, and contributes to the pathogenesis of necrotizing pneumonia. Here, we discovered that SElX can also bind to neutrophils from human and other mammalian species and disrupt IgG-mediated phagocytosis. Site-directed mutagenesis of the conserved sialic acid-binding motif of SElX abolished neutrophil binding and phagocytic killing, and revealed multiple glycosylated neutrophil receptors for SElX binding. Furthermore, the neutrophil binding-deficient mutant of SElX retained its capacity for T-cell activation demonstrating that SElX exhibits mechanistically independent activities on distinct cell populations associated with acquired and innate immunity, respectively. Finally, we demonstrated that the neutrophil-binding activity rather than superantigenicity is responsible for the SElX-dependent virulence observed in a necrotizing pneumonia rabbit model of infection. Taken together, we report the first example of a SAg, that can manipulate both the innate and adaptive arms of the human immune system during S. aureus pathogenesis.

IL-26 in allergic contact dermatitis: Resource in a state of readiness.

In this study, we investigated the role of IL-26 in allergic contact dermatitis (ACD), highlighting its' contribute in the cytotoxic mechanism responsible for the tissue injury. IL-26 is a signature Th17 cytokine, and immune cells are its predominant sources. Recently, it has shown that Th17 cell-derived-IL-26 functions like an antimicrobial peptide. Here, we hypothesized that IL-26 could be involved in cytotoxicity mechanism that underlies ACD. Indeed, we have attributed a role to IL-26 in this context, through PBMC cytotoxicity assays vs HaCat. To demonstrate that IL-26 was effectively involved in this activity, we performed the assay using transfected ACD PBMCs by siRNA for IL-26. Indeed, we demonstrated that these cells were less able to kill keratinocytes compared with ACD PBMCs (P < .01). In conclusion, our findings support the idea that this emergent cytokine, IL-26, is implicated in the killing mechanisms of KC observed during ACD.

Molecular basis determining species specificity for TLR2 inhibition by staphylococcal superantigen-like protein 3 (SSL3).

Staphylococcus aureus is a versatile opportunistic pathogen, causing disease in human and animal species. Its pathogenicity is linked to the ability of S. aureus to secrete immunomodulatory molecules. These evasion proteins bind to host receptors or their ligands, resulting in inhibitory effects through high affinity protein-protein interactions. Staphylococcal evasion molecules are often species-specific due to differences in host target proteins between species. We recently solved the crystal structure of murine TLR2 in complex with immunomodulatory molecule staphylococcal superantigen-like protein 3 (SSL3), which revealed the essential residues within SSL3 for TLR2 inhibition. In this study we aimed to investigate the molecular basis of the interaction on the TLR2 side. The SSL3 binding region on murine TLR2 was compared to that of other species through sequence alignment and homology modeling, which identified interspecies differences. To examine whether this resulted in altered SSL3 activity on the corresponding TLR2s, bovine, equine, human, and murine TLR2 were stably expressed in HEK293T cells and the ability of SSL3 to inhibit TLR2 was assessed. We found that SSL3 was unable to inhibit bovine TLR2. Subsequent loss and gain of function mutagenesis showed that the lack of inhibition is explained by the absence of two tyrosine residues in bovine TLR2 that play a prominent role in the SSL3-TLR2 interface. We found no evidence for the existence of allelic SSL3 variants that have adapted to the bovine host. Thus, within this paper we reveal the molecular determinants of the TLR2-SSL3 interaction which adds to our understanding of staphylococcal host specificity.

Superantigen staphylococcal enterotoxin C1 inhibits the growth of bladder cancer.

Superantigens can induce cell-mediated cytotoxicity preferentially against MHC II-positive target cells with large amounts of inflammatory cytokines releasing. In this study, superantigen staphylococcal enterotoxin C (SEC) 1 was investigated to evaluate its potential in bladder cancer immunotherapy in vitro and in vivo. Our results revealed that SEC1 could stimulate the proliferation of human peripheral blood mononuclear cells (PBMCs) in a dose-dependent manner, accompanied with the release of interleukin-2, interferon-γ, and tumor necrosis factor-α, and increased the population of CD4+ T cells and CD8+ T cells. PBMCs stimulated by SEC1 could initiate significant cytotoxicity towards human bladder cancer cells in vitro. The results of in vivo antitumor experiment indicated that SEC1 could decrease the rate of tumor formation and prolong the survival time of tumor-bearing mice. Our study demonstrated that SEC1 inhibited the growth of bladder cancer. And it is also suggested that SEC1 may become a candidate for bladder cancer immunotherapy.

Complement Inhibition: A Novel Form of Immunotherapy for Colon Cancer.

BACKGROUND: Complement is a central part of both the innate and adaptive immune response and its activation has traditionally been considered part of the immunosurveillance response against cancer. Its pro-inflammatory role and its contribution to the development of many illnesses associated with inflammatory states implicate complement in carcinogenesis. METHODS: We evaluated the role of three protein inhibitors of complement-cobra venom factor, humanized cobra venom factor, and recombinant staphylococcus aureus superantigen-like protein 7-in the setting of a transplantable murine colon cancer model. Outcomes were evaluated by monitoring tumor growth, and flow cytometry, ELISPOT, and quantitative real-time PCR were used to determine the impact of complement inhibition on the host immune response. RESULTS: Complement inhibitors were effective at depleting complement component C3 in tumor bearing mice and this was temporally correlated with a decreased rate of tumor growth during the establishment of tumors. Treatment with cobra venom factor resulted in increased CD8(+) T cells as a percentage of tumor-infiltrating cells as well as a reduced immunosuppressive environment evidenced by decreased myeloid derived suppressor cells in splenocytes of treated mice. Complement inhibition resulted in increased expression of the chemoattractive cytokines CCL5, CXCL10, and CXCL11. DISCUSSION: Complement depletion represents a promising mode of immunotherapy in cancer by its ability to impair tumor growth by increasing the host's effective immune response to tumor and diminishing the immunosuppressive effect created by the tumor microenvironment and ultimately could be utilized as a component of combination immunotherapy.

Naptumomab estafenatox: targeted immunotherapy with a novel immunotoxin.

Improvement of cancer therapy by introducing new concepts is still urgent even though there have been major advancements lately. Immunotherapy is well on the way to becoming an established tool in the cancer treatment armory. It seems that a combination of (1) activation of immune effector cells and selective targeting of them to tumors and (2) the inhibition of immune suppression often induced by the tumor itself are necessary to achieve the therapeutic goal. The immunotoxin naptumomab estafenatox was developed in an effort to activate and target the patient's own T cells to their tumor, by fusing a superantigen (SAg) variant that activates T lymphocytes to the Fab moiety of a tumor-reactive monoclonal antibody. Naptumomab estafenatox targets the 5T4 tumor antigen, a 72-kDa oncofetal trophoblast protein expressed on many carcinomas, including renal cell carcinoma. The therapeutic effect is associated with activation of SAg-binding T cells. The SAg-binding T lymphocytes expand, differentiate to effector cells, and infiltrate the tumor. The therapeutic efficacy is most likely related to the dual mechanism of tumor cell killing: (1) direct lysis by cytotoxic T lymphocytes of tumor cells expressing the antigen recognized by the antibody moiety of the fusion protein and (2) secretion of cytokines eliminating antigen-negative tumor cell variants. Naptumomab estafenatox has been clinically tested in a range of solid tumors with focus on renal cell carcinoma. This review looks at the clinical experience with the new immunotoxin and its potential.

Effective T-cell recall responses require the taurine transporter Taut.

T-cell activation and the subsequent transformation of activated T cells into T-cell blasts require profound changes in cell volume. However, the impact of cell volume regulation for T-cell immunology has not been characterized. Here we studied the role of the cell-volume regulating osmolyte transporter Taut for T-cell activation in Taut-deficient mice. T-cell mediated recall responses were severely impaired in taut(-/-) mice as shown with B16 melanoma rejection and hapten-induced contact hypersensitivity. CD4(+) and CD8(+) T cells were unequivocally located within peripheral lymph nodes of unprimed taut(-/-) mice but significantly decreased in taut(-/-) compared with taut(+/+) mice following in vivo activation. Further analysis revealed that Taut is critical for rescuing T cells from activation-induced cell death in vitro and in vivo as shown with TCR, superantigen, and antigen-specific activation. Consequently, reduction of CD4(+) and CD8(+) T cells in taut(-/-) mice upon antigen challenge resulted in impaired in vivo generation of T-cell memory. These findings disclose for the first time that volume regulation in T cells is an element in the regulation of adaptive immune responses and that the osmolyte transporter Taut is crucial for T-cell survival and T-cell mediated immune reactions.

Th17 cells expressing KIR3DL2+ and responsive to HLA-B27 homodimers are increased in ankylosing spondylitis.

CD4 Th cells producing the proinflammatory cytokine IL-17 (Th17) have been implicated in a number of inflammatory arthritides including the spondyloarthritides. Th17 development is promoted by IL-23. Ankylosing spondylitis, the most common spondyloarthritis (SpA), is genetically associated with both HLA-B27 (B27) and IL-23R polymorphisms; however, the link remains unexplained. We have previously shown that B27 can form H chain dimers (termed B27(2)), which, unlike classical HLA-B27, bind the killer-cell Ig-like receptor KIR3DL2. In this article, we show that B27(2)-expressing APCs stimulate the survival, proliferation, and IL-17 production of KIR3DL2(+) CD4 T cells. KIR3DL2(+) CD4 T cells are expanded and enriched for IL-17 production in the blood and synovial fluid of patients with SpA. Despite KIR3DL2(+) cells comprising a mean of just 15% of CD4 T in the peripheral blood of SpA patients, this subset accounted for 70% of the observed increase in Th17 numbers in SpA patients compared with control subjects. TCR-stimulated peripheral blood KIR3DL2(+) CD4 T cell lines from SpA patients secreted 4-fold more IL-17 than KIR3DL2(+) lines from controls or KIR3DL2(-) CD4 T cells. Strikingly, KIR3DL2(+) CD4 T cells account for the majority of peripheral blood CD4 T cell IL-23R expression and produce more IL-17 in the presence of IL-23. Our findings link HLA-B27 with IL-17 production and suggest new therapeutic strategies in ankylosing spondylitis/SpA.

Staphylococcus aureus α-toxin modulates skin host response to viral infection.

BACKGROUND: Patients with atopic dermatitis (AD) with a history of eczema herpeticum have increased staphylococcal colonization and infections. However, whether Staphylococcus aureus alters the outcome of skin viral infection has not been determined. OBJECTIVE: We investigated whether S aureus toxins modulated host response to herpes simplex virus (HSV) 1 and vaccinia virus (VV) infections in normal human keratinocytes (NHKs) and in murine infection models. METHODS: NHKs were treated with S aureus toxins before incubation of viruses. BALB/c mice were inoculated with S aureus 2 days before VV scarification. Viral loads of HSV-1 and VV were evaluated by using real-time PCR, a viral plaque-forming assay, and immunofluorescence staining. Small interfering RNA duplexes were used to knockdown the gene expression of the cellular receptor of α-toxin, a disintegrin and metalloprotease 10 (ADAM10). ADAM10 protein and α-toxin heptamers were detected by using Western blot assays. RESULTS: We demonstrate that sublytic staphylococcal α-toxin increases viral loads of HSV-1 and VV in NHKs. Furthermore, we demonstrate in vivo that the VV load is significantly greater (P < .05) in murine skin inoculated with an α-toxin-producing S aureus strain compared with murine skin inoculated with the isogenic α-toxin-deleted strain. The viral enhancing effect of α-toxin is mediated by ADAM10 and is associated with its pore-forming property. Moreover, we demonstrate that α-toxin promotes viral entry in NHKs. CONCLUSION: The current study introduces the novel concept that staphylococcal α-toxin promotes viral skin infection and provides a mechanism by which S aureus infection might predispose the host toward disseminated viral infections.

Peroxisome proliferator-activated receptor-γ agonists prevent in vivo remodeling of human artery induced by alloreactive T cells.

BACKGROUND: Ligands activating the transcription factor peroxisome proliferator-activated receptor-γ (PPARγ) have antiinflammatory effects. Vascular rejection induced by allogeneic T cells can be responsible for acute and chronic graft loss. Studies in rodents suggest that PPARγ agonists may inhibit graft vascular rejection, but human T-cell responses to allogeneic vascular cells differ from those in rodents, and the effects of PPARγ in human transplantation are unknown. METHODS AND RESULTS: We tested the effects of PPARγ agonists on human vascular graft rejection using a model in which human artery is interposed into the abdominal aorta of immunodeficient mice, followed by adoptive transfer of allogeneic (to the artery donor) human peripheral blood mononuclear cells. Interferon-γ-dependent rejection ensues within 4 weeks, characterized by intimal thickening, T-cell infiltrates, and vascular cell activation, a response resembling clinical intimal arteritis. The PPARγ agonists 15-deoxy-prostaglandin-J(2), ciglitazone, and pioglitazone reduced intimal expansion, intimal infiltration of CD45RO(+) memory T cells, and plasma levels of inflammatory cytokines. The PPARγ antagonist GW9662 reversed the protective effects of PPARγ agonists, confirming the involvement of PPARγ-mediated pathways. In vitro, pioglitazone inhibited both alloantigen-induced proliferation and superantigen-induced transendothelial migration of memory T cells, indicating the potential mechanisms of PPARγ effects. CONCLUSION: Our results suggest that PPARγ agonists inhibit allogeneic human memory T cell responses and may be useful for the treatment of vascular graft rejection.

Staphylococcal-derived superantigen enhances peanut induced Th2 responses in the skin.

BACKGROUND: The allergen-induced activation and expansion of IL-4 producing T helper type 2 (Th2) cells is a key event in the initiation and progression of allergic disease. Intriguingly, concomitant early childhood staphylococcal skin infections are being increasingly implicated in the allergen-induced switch of primary T cell responses towards the Th2 phenotype. OBJECTIVE: We sought to identify whether or not staphylococcal-derived superantigen can influence the primary T cell response in the skin to food allergens with a view to determining whether such exposures create the immune pathology that predisposes to the development of food allergy. METHODS: Using a novel Th2 reporter model, we determined the ability of the staphylococcal superantigen (SEB) to influence priming in the skin of IL-4 expressing Th2 cells by peanut extract (PE). Factors including the effect of SEB on the magnitude of the Th2 response in the skin draining lymph nodes, T cell receptor V region usage and the influence of endotoxin were evaluated. RESULTS: Primary exposure to PE and SEB lead to significantly enhanced PE specific Th2 responses when the mice were subsequently exposed to PE alone. The enhancement of the Th2 response was dependent on the Vß-binding properties of the SEB, but was not affected by endotoxin-mediated TLR-4 effects or strain differences in the mice. CONCLUSIONS AND CLINICAL RELEVANCE: These results identify that in the skin environment, the presence of SEB can significantly increase the numbers of allergen-induced Th2 cells which develop in response to subsequent allergen exposure. These data highlight the process by which individuals may become pathologically sensitized to food allergens in early life.

Activation of human γδ T cells and NK cells by Staphylococcal enterotoxins requires both monocytes and conventional T cells.

Staphylococcal enterotoxins (SE) pose a great threat to human health due to their ability to bypass antigen presentation and activate large amounts of conventional T cells resulting in a cytokine storm potentially leading to toxic shock syndrome. Unconventional T- and NK cells are also activated by SE but the mechanisms remain poorly understood. In this study, the authors aimed to explore the underlying mechanism behind SE-mediated activation of MAIT-, γδ T-, and NK cells in vitro. CBMC or PBMC were stimulated with the toxins SEA, SEH, and TSST-1, and cytokine and cytotoxic responses were analyzed with ELISA and flow cytometry. All toxins induced a broad range of cytokines, perforin and granzyme B, although SEH was not as potent as SEA and TSST-1. SE-induced IFN-γ expression in MAIT-, γδ T-, and NK cells was clearly reduced by neutralization of IL-12, while cytotoxic compounds were not affected at all. Kinetic assays showed that unconventional T cell and NK cell-responses are secondary to the response in conventional T cells. Furthermore, co-cultures of isolated cell populations revealed that the ability of SEA to activate γδ T- and NK cells was fully dependent on the presence of both monocytes and αß T cells. Lastly, it was found that SE provoked a reduced and delayed cytokine response in infants, particularly within the unconventional T and NK cell populations. This study provides novel insights regarding the activation of unconventional T- and NK cells by SE, which contribute to understanding the vulnerability of young children towards Staphylococcus aureus infections.

Superantigen antagonist protects against lethal shock and defines a new domain for T-cell activation.

Superantigens trigger an excessive cellular immune response, leading to toxic shock. We have designed a peptide antagonist that inhibits superantigen-induced expression of human genes for interleukin-2, gamma interferon and tumor necrosis factor-b, which are cytokines that mediate shock. The peptide shows homology to a b-strand-hinge-a-helix domain that is structurally conserved in superantigens, yet is remote from known binding sites for the major histocompatibility class II molecule and T-cell receptor. Superantigens depend on this domain for T-cell activation. The peptide protected mice against lethal challenge with staphylococcal and streptococcal superantigens. Moreover, it rescued mice undergoing toxic shock. Surviving mice rapidly developed protective antibodies against superantigen that rendered them resistant to further lethal challenges, even with different superantigens. Thus, the lethal effect of superantigens can be blocked with a peptide antagonist that inhibits their action at the beginning of the toxicity cascade, before activation of T cells takes place.

Superantigenic activity of toxic shock syndrome toxin-1 is resistant to heating and digestive enzymes.

AIMS: To elucidate the stability of superantigenic activity and pathogenesis of toxic shock syndrome toxin 1 (TSST-1) and staphylococcal enterotoxin A (SEA) against heating and digestive enzymes. METHODS AND RESULTS: Purified TSST-1 and SEA were treated with heating, pepsin and trypsin that are related to food cooking, stomach and intestine conditions. The integrity, superantigenic activity and toxicity of treated TSST-1 and SEA were analysed by Western blotting, spleen cell culture, cytokine assay and toxic shock models. Both TSST-1 and SEA showed strong resistance to heating, pepsin and trypsin digestion. Furthermore, the treated TSST-1 showed significant higher induction of interferon-γ and toxic shock compared with that of SEA. Pepsin- or trypsin-digested TSST-1 fragments still showed significant superantigenic and lethal shock toxicities. CONCLUSIONS: The superantigenic activity of TSST-1 was stable to heating and digestive enzymes. Pepsin- and trypsin-digested TSST-1 fragments still showed superantigenic and lethal shock activities, indicating that digested TSST-1 could cross epithelial cells and induce systemic toxicity. SIGNIFICANCE AND IMPACT OF THE STUDY: This study found, for the first time, that pepsin- or trypsin-digested smaller TSST-1 retained significant superantigenic and lethal shock activities. The different resistance of TSST-1 and SEA participates in the different pathogenic activities during food poisoning and toxic shock syndrome.

Staphylococcal TSST-1 Association with Eczema Herpeticum in Humans.

Atopic dermatitis (AD) is a condition affecting 30 million persons in the United States. AD patients are heavily infected with Staphylococcus aureus on the skin. A particularly severe form of AD is eczema herpeticum (ADEH), where the patients' AD is complicated by S. aureus and herpes simplex virus (HSV) infection. This study examined the S. aureus strains from 15 ADEH patients, provided blinded, and showed a high association of ADEH with strains that produce toxic shock syndrome toxin-1 (TSST-1; 73%) compared to 10% production by typical AD isolates from patients without EH and those from another unrelated condition, cystic fibrosis. The ADEH isolates produced the superantigens associated with TSS (TSST-1 and staphylococcal enterotoxins A, B, and C). This association may in part explain the potential severity of ADEH. We also examined the effect of TSST-1 and HSV-1 on human epithelial cells and keratinocytes. TSST-1 used CD40 as its receptor on epithelial cells, and HSV-1 either directly or indirectly interacted with CD40. The consequence of these interactions was chemokine production, which is capable of causing harmful inflammation, with epidermal/keratinocyte barrier disruption. Human epithelial cells treated first with TSST-1 and then HSV-1 resulted in enhanced chemokine production. Finally, we showed that TSST-1 modestly increased HSV-1 replication but did not increase viral plaque size. Our data suggest that ADEH is associated with production of the major TSS-associated superantigens, together with HSV reactivation. The superantigens plus HSV may damage the skin barrier by causing harmful inflammation, thereby leading to increased symptoms. IMPORTANCE Atopic dermatitis (eczema, AD) with concurrent herpes simplex virus infection (eczema herpeticum, ADEH) is a severe form of AD. We show that ADEH patients are colonized with Staphylococcus aureus that primarily produces the superantigen toxic shock syndrome toxin-1 (TSST-1); however, significantly but to a lesser extent the superantigens staphylococcal enterotoxins A, B, and C are also represented in ADEH. Our studies showed that TSST-1 uses the immune costimulatory molecule CD40 as its epithelial cell receptor. Herpes simplex virus (HSV) also interacted directly or indirectly with CD40 on epithelial cells. Treatment of epithelial cells with TSST-1 and then HSV-1 resulted in enhanced chemokine production. We propose that this combination of exposures (TSST-1 and then HSV) leads to opening of epithelial and skin barriers to facilitate potentially serious ADEH.

Proliferation is a prerequisite for bacterial superantigen-induced T cell apoptosis in vivo.

Staphylococcal enterotoxin B (SEB) is a bacterial superantigen that binds to major histocompatibility complex class II molecules and selectively interacts with T cells that bear certain T cell receptor (TCR) V beta domains. Administration of SEB in adult mice results in initial proliferation of V beta 8+ T cells followed by a state of unresponsiveness resulting from a combination of clonal deletion and clonal anergy in the SEB-reactive population. At this time, it is unclear what relationship exists between the T cells that have proliferated and those that have been deleted or have become anergic. Here we show that only a fraction of the potentially reactive V beta 8+ T cells proliferate in response to SEB in vivo, and that all the cells that have proliferated eventually undergo apoptosis. Virtually no apoptosis can be detected in the nonproliferating V beta 8+ T cells. These data demonstrate a causal relationship between proliferation and apoptosis in response to SEB in vivo, and they further indicate that T cells bearing the same TCR V beta segment can respond differently to the same superantigen. The implications of this differential responsiveness in terms of activation and tolerance are discussed.