Administration Of Staphylococcal Enterotoxin B Research Articles

Cellular immune response of Staphylococcus aureus enterotoxin B in Balb/c mice through intranasal infection.

Background and Aim:Staphylococcus aureus produces various superantigen exotoxins, including staphylococcal enterotoxin B (SEB). It causes fatal anaphylactic reactions and toxic shock. This study aimed to evaluate the reaction of leukocytes and histopathological changes in the respiratory organs of Balb/c mice after intranasal infection with enterotoxigenic S. aureus (SEB).Materials and Methods:The presence of the seb gene in S. aureus was established in this study using polymerase chain reaction-specific primer. Two groups of 8-week-old male Balb-c mice consist of six mice in each group. The treated group was infected with 50 μL and 100 μL of SEB intranasal on days 1 and 14, respectively. NaCl was administered in the second group and was considered as a control group. Blood samples were collected through the retro-orbital plexus on days 1, 4, 7, 14, and 22 after infections. Total cell counts were analyzed with an independent sample t-test and compared using the statistical package for the social sciences (SPSS) version 16.0 (IBM Corp., NY, USA). The infected tissues of the respiratory organ were observed descriptively and compared to the control group.Results:The seb gene with a molecular size of 478 bp, indicating the SEB strain, is present in S. aureus used in this study. Intranasal administration of SEB showed increased leukocytes, lymphocytes, monocytes, and eosinophils on day 22 post-infection. Significant leukocytosis was seen on days 6 and 14; lymphocytosis on days 1, 4, 6, and 16; and eosinophilia on days 6, 14, and 22 compared with the control group (p > 0.05). In contrast, the neutrophil decreased after an increase of immature band cells compared to the control group, indicating a severe acute infection with SEB. The lungs and trachea of the test group had an inflammatory cell accumulation in the respiratory organ.Conclusion:Intranasal route infection of S. aureus containing seb gene significantly induced the cellular immune response and caused pathological changes in the respiratory tissues of the Balb/c mice model. The hematological changes were aligned with marked pathological changes in the respiratory tract. Balb/c mice could be an excellent experimental model to study toxic and anaphylactic shock against SEB to define the future therapeutic agents.

Prenatal Exposure of Rats to Staphylococcal Enterotoxin B Alters the Expression of Th1 and Th2 Cytokines via Decreased Methylation in the Spleen of Adult but not Neonatal Offspring.

The methylation of IFN-γ and IL-4 genes is regarded as an epigenetic regulation that maintains the Th1 or Th2 phenotype. To explore the influence of prenatal administration of the staphylococcal enterotoxin B (SEB) in pregnant rats, on the IFN-γ or IL-4 expression in the offspring spleen. The SEB or PBS was administered intravenously to pregnant rats on the embryo-day 16. After normal delivery, the spleens from the fifth-day neonates and adult offspring were isolated under anesthesia. Quantitative PCR, western blot, ELISA and MeDIP-qPCR were applied to determine the levels of the splenic IFN-γ or IL-4 mRNAs, their protein levels, and methylation status, respectively. Prenatal administration of the SEB in pregnant rats decreased the levels of the splenic IFN-γ and IL-4 proteins in neonates, but increased their mRNA levels. However, prenatal administration of the SEB significantly augmented both mRNA and protein levels of the IFN-γ and IL-4 in the adult spleen. In addition, the prenatal SEB administration decreased the methylation of the splenic IFN-γ and IL-4 in adult but not neonatal offspring. The prenatal administration of SEB in pregnant rats can cause a mixed Th1 and Th2 cytokines response in the offspring spleen, and alter the cytokine expression of the Th1 and Th2 via decreasing the methylation in adult but, not neonatal offspring spleen.

Resveratrol-mediated attenuation of superantigen-driven acute respiratory distress syndrome is mediated by microbiota in the lungs and gut

Acute Respiratory Distress Syndrome (ARDS) is triggered by a variety of agents, including Staphylococcal Enterotoxin B (SEB). Interestingly, a significant proportion of patients with COVID-19, also develop ARDS. In the absence of effective treatments, ARDS results in almost 40% mortality. Previous studies from our laboratory demonstrated that resveratrol (RES), a stilbenoid, with potent anti-inflammatory properties can attenuate SEB-induced ARDS. In the current study, we investigated the role of RES-induced alterations in the gut and lung microbiota in the regulation of ARDS. Our studies revealed that SEB administration induced inflammatory cytokines, ARDS, and 100% mortality in C3H/HeJ mice. Additionally, SEB caused a significant increase in pathogenic Proteobacteria phylum and Propionibacterium acnes species in the lungs. In contrast, RES treatment attenuated SEB-mediated ARDS and mortality in mice, and significantly increased probiotic Actinobacteria phylum, Tenericutes phylum, and Lactobacillus reuteri species in both the colon and lungs. Colonic Microbiota Transplantation (CMT) from SEB-injected mice that were treated with RES as well as the transfer of L. reuteri into recipient mice inhibited the production of SEB-mediated induction of pro-inflammatory cytokines such as IFN-γ and IL-17 but increased that of anti-inflammatory IL-10. Additionally, such CMT and L. reuteri recipient mice exposed to SEB, showed a decrease in lung-infiltrating mononuclear cells, cytotoxic CD8+ T cells, NKT cells, Th1 cells, and Th17 cells, but an increase in the population of regulatory T cells (Tregs) and Th3 cells, and increase in the survival of mice from SEB-mediated ARDS. Together, the current study demonstrates that ARDS induced by SEB triggers dysbiosis in the lungs and gut and that attenuation of ARDS by RES may be mediated, at least in part, by alterations in microbiota in the lungs and the gut, especially through the induction of beneficial bacteria such as L. reuteri.

Discordant rearrangement of primary and anamnestic CD8+ T cell responses to influenza A viral epitopes upon exposure to bacterial superantigens: Implications for prophylactic vaccination, heterosubtypic immunity and superinfections.

Infection with (SAg)-producing bacteria may precede or follow infection with or vaccination against influenza A viruses (IAVs). However, how SAgs alter the breadth of IAV-specific CD8+ T cell (TCD8) responses is unknown. Moreover, whether recall responses mediating heterosubtypic immunity to IAVs are manipulated by SAgs remains unexplored. We employed wild-type (WT) and mutant bacterial SAgs, SAg-sufficient/deficient Staphylococcus aureus strains, and WT, mouse-adapted and reassortant IAV strains in multiple in vivo settings to address the above questions. Contrary to the popular view that SAgs delete or anergize T cells, systemic administration of staphylococcal enterotoxin B (SEB) or Mycoplasma arthritidis mitogen before intraperitoneal IAV immunization enlarged the clonal size of 'select' IAV-specific TCD8 and reshuffled the hierarchical pattern of primary TCD8 responses. This was mechanistically linked to the TCR Vβ makeup of the impacted clones rather than their immunodominance status. Importantly, SAg-expanded TCD8 retained their IFN-γ production and cognate cytolytic capacities. The enhancing effect of SEB on immunodominant TCD8 was also evident in primary responses to vaccination with heat-inactivated and live attenuated IAV strains administered intramuscularly and intranasally, respectively. Interestingly, in prime-boost immunization settings, the outcome of SEB administration depended strictly upon the time point at which this SAg was introduced. Accordingly, SEB injection before priming raised CD127highKLRG1low memory precursor frequencies and augmented the anamnestic responses of SEB-binding TCD8. By comparison, introducing SEB before boosting diminished recall responses to IAV-derived epitopes drastically and indiscriminately. This was accompanied by lower Ki67 and higher Fas, LAG-3 and PD-1 levels consistent with a pro-apoptotic and/or exhausted phenotype. Therefore, SAgs can have contrasting impacts on anti-IAV immunity depending on the naïve/memory status and the TCR composition of exposed TCD8. Finally, local administration of SEB or infection with SEB-producing S. aureus enhanced pulmonary TCD8 responses to IAV. Our findings have clear implications for superinfections and prophylactic vaccination.

Induction of specific T helper-9 cells to inhibit glioma cell growth.

The effects of Staphylococcal enterotoxin B (SEB) on regulation of immune response have been recognized; whether SEB can enhance the effects of immunotherapy on glioma remains to be investigated. This study tests a hypothesis that administration with SEB enhances the effects of specific immunotherapy on glioma growth in mice. In this study, a glioma-bearing mouse model was developed by adoptive transfer with GL261 cells (a mouse glioma cell line). The mice were treated with the GL261 cell extracts (used as an Ag) with or without administration of SEB. We observed that treating glioma-bearing mice with the glioma Ag and SEB induced glioma-specific Th9 cells in both glioma tissue and the spleen. Treating CD4+ CD25− T cells with SEB increased p300 phosphorylation, histone H3K4 acetylation at the interleukin (IL)-9 promoter locus, and increased the IL-9 transcriptional factor binding to the IL-9 promoter. Treating CD4+ CD25− T cells with both SEB and glioma Ag induced glioma-specific Th9 cells. The glioma-specific Th9 cells induced glioma cell apoptosis in the culture. Treating the glioma-bearing mice with SEB and glioma Ag significantly inhibited the glioma growth. In conclusion, SEB plus glioma Ag immunotherapy inhibits the experimental glioma growth, which may be a novel therapeutic remedy for the treatment of glioma.

Inhibition of squamous cancer growth in a mouse model by Staphylococcal enterotoxin B-triggered Th9 cell expansion.

Currently, therapy for squamous cancer (SqC) is unsatisfactory. Staphylococcal enterotoxin B (SEB) has strong immune regulatory activity. This study tests the hypothesis that SEB enforces the effect of immunotherapy on SqC growth in a mouse model. C3H/HeN mice and the SqC cell line squamous cell carcinoma VII were used to create an SqC mouse model. Immune cell assessment was performed by flow cytometry. Real-time RT-PCR and western blotting were used to evaluate target molecule expression. An apoptosis assay was used to assess the suppressive effect of T helper-9 (Th9) cells on the SqC cells. The results showed that immunotherapy consisting of SEB plus SqC antigen significantly inhibited SqC growth in the mice. The frequency of Th9 cells was markedly increased in the SqC tissue and mouse spleens after treatment. SEB markedly increased the levels of signal transducer and activator of transcription 5 phosphorylation and the expression of histone deacetylase-1 (HDAC1) and PU.1 (the transcription factor of the interleukin 9 (IL-9) gene) in CD4+ T cells. Exposure to SqC-specific Th9 cells markedly induced SqC cell apoptosis both in vitro and in vivo. In conclusion, the administration of SEB induces Th9 cells in SqC-bearing mice, and theseTh9 cells inhibit SqC growth.

Δ(9) Tetrahydrocannabinol attenuates Staphylococcal enterotoxin B-induced inflammatory lung injury and prevents mortality in mice by modulation of miR-17-92 cluster and induction of T-regulatory cells.

Staphylococcal enterotoxin B (SEB) is a potent activator of Vβ8+T-cells resulting in the clonal expansion of ∼30% of the T-cell pool. Consequently, this leads to the release of inflammatory cytokines, toxic shock, and eventually death. In the current study, we investigated if Δ(9) tetrahydrocannabinol (THC), a cannabinoid known for its anti-inflammatory properties, could prevent SEB-induced mortality and alleviate symptoms of toxic shock. We investigated the efficacy of THC against the dual administration (intranasal and i.p.) of SEB into C3H/HeJ mice based on the measurement of SEB-mediated clinical parameters, including cytokine production, cellular infiltration, vascular leak, and airway resistance. In addition, the molecular mechanism of action was elucidated in vitro by the activation of splenocytes with SEB. Exposure to SEB resulted in acute mortality, while THC treatment led to 100% survival of mice. SEB induced the miRNA-17-92 cluster, specifically miRNA-18a, which targeted Pten (phosphatase and tensin homologue), an inhibitor of the PI3K/Akt signalling pathway, thereby suppressing T-regulatory cells. In contrast, THC treatment inhibited the individual miRNAs in the cluster, reversing the effects of SEB. We report, for the first time a role for the miRNA 17-92 cluster in SEB-mediated inflammation. Furthermore, our results suggest that THC is a potent anti-inflammatory compound that may serve as a novel therapeutic to suppress SEB-induced pulmonary inflammation by modulating critical miRNA involved in SEB-induced toxicity and death.

Staphylococcal Enterotoxin B (SEB) Induces Memory CD4 T Cell Anergy in vivo and Impairs Recall Immunity to Unrelated Antigens.

IntroductionNaïve and memory T cells can utilize unique regulatory pathways to promote protection but prevent self-reactivity. A bacterial superantigen SEB exploits unique TCR proximal signaling processes in memory CD4 T cells to induce clonal anergy. The aim of this study was to determine if SEB could antagonize memory CD4 T cells in vivo and whether there would be consequences on recall immune responses. We evaluated Ab responses to a T-dependent antigen as a measurement of memory T cell helper function.MethodBALB/c mice were primed with TNP-RGG to elicit memory B cells and also immunized with an ovalbumin peptide to elicit memory helper T cells. Another group of TNP-RGG immunized mice were used as adoptive transfer recipients of exogenous DO11.10 memory T cells. Mice were challenged with TNP-OVA with or without prior administration of SEB. B cells secreting IgM or IgG TNP-specific Ab were enumerated by ELISPOT as indicators of primary versus secondary humoral immunity.ResultsComparing the SEB and non-SEB-treated groups, the SEB-treated group failed to produce TNP-specific IgG in response to challenge with TNP-OVA, even if they were previously immunized with OVA. All groups produced IgM, indicating that the primary Ab responses and naïve helper T cells were not impacted by SEB. SEB had no negative impact when DO11.10 × Fyn−/− memory T cells were used as donor cells.ConclusionThe present study indicated that SEB selectively targeted memory CD4 T cells in vivo and prevented helper function. Consequently, recall humoral immunity was lost. The data are most consistent with in vivo T cell anergy as opposed to indirect suppression as elimination of Fyn kinase restored helper function. These data suggest that bacterial superantigens can impair post-vaccination memory cell responses to unrelated antigens via their ability to target Vb families and antagonize memory cell activation.

Staphylococcal enterotoxin B administration during pregnancy imprints the increased CD4:CD8 T-cell ratio in the peripheral blood from neonatal to adult offspring rats.

Our previous study demonstrated that Staphylococcal enterotoxin B (SEB) administration during pregnancy could alter the percentage of T cells subpopulation in the thymus of the neonatal rats; however, little is known about the effect of maternal SEB administration during pregnancy on T cells subpopulation in the peripheral blood of the offspring rats. In the present study, pregnant rats at gestational day 16 were intravenously injected with 15 µg SEB. The present study found that prenatal exposure to SEB significantly decreased the percentages of CD8 T cells in the peripheral blood of both neonatal rats on the fifth day after delivery and the adult offspring rats. Furthermore, it significantly increased the percentage of CD4 T cells as well as the ratios of CD4 to CD8 T cells in both neonatal and adult offspring rats. Prenatal exposure to SEB significantly decreased the expression levels of IL-4 and IFN-γ in the plasma of neonatal and adult offspring rats. Furthermore, SEB restimulation significantly increased the percentage of CD8 T cells and significantly decreased the percentage of CD4 T cells. These data suggest the prenatal exposure to SEB can imprint the increased CD4:CD8 T cell ratio in the peripheral blood from the neonate to adulthood through the decreased CD8 T cells and the increased CD4 T cells, and altered the response characteristics of CD4 and CD8 T cells to secondary SEB administration in the peripheral blood of the adult offspring rats.

Decreased Vβ8.2 T-cells in neonatal rats exposed prenatally to Staphylococcal enterotoxin B are further deleted by restimulation in an in vitro cultured thymus.

Staphylococcal enterotoxin B (SEB) administration during adulthood can cause the anergy or deletion of variable portion of the ? chain (V?)?expressing T cells. However, the effect of maternal SEB administration during pregnancy on the thymocytes of neonatal rats remains to be elucidated. In the present study, pregnant rats at gestational day16 were intravenously injected with 15µg SEB. The present study revealed that prenatal exposure of SEB significantly increased the proportion of cluster of differentiation (CD)4?single positive (SP) Tcells and decreased the proportions of CD8?SP, CD4+ V?8.2+ and CD8+ V?8.2+ T cells in the thymus of neonatal rats between day 0 and 5 after delivery. In an invitro cultured thymus, SEB restimulation significantly increased the proportion of double positive cells and decreased the proportions of CD4?SP, CD8?SP, CD4+ V?8.2+ and CD8+ V?8.2+ Tcells. Furthermore, the decreased V?8.2+ T?cells in neonatal rats exposed prenatally to SEB were further deleted by SEB restimulation in an invitro cultured thymus. These data suggested the special response pattern of the remaining SEB?specific Tcells to SEB restimulation in neonatal rats exposed prenatally to SEB.

Amygdaloid Signature of Peripheral Immune Activation by Bacterial Lipopolysaccharide or Staphylococcal Enterotoxin B

Activated immune cells produce soluble mediators that not only coordinate local and systemic immune responses but also act on the brain to initiate behavioral, neuroendocrine and metabolic adaptations. Earlier studies have shown that the amygdala, a group of nuclei located in the medial temporal lobe, is engaged in the central processing of afferent signals from the peripheral immune system. Here, we compared amygdaloid responses to lipopolysaccharide (LPS) and staphylococcal enterotoxin B (SEB), two prototypic bacterial products that elicit distinct immune responses. Intraperitoneal administration of LPS (0.1mg/kg) or SEB (1mg/kg) in adult rats induced substantial increases in amygdaloid neuronal activity as measured by intracerebral electroencephalography and c-fos gene expression. Amygdaloid neuronal activation was accompanied by an increase in anxiety-related behavior in the elevated plus-maze test. However, only treatment with LPS, but not SEB, enhanced amygdaloid IL-1β and TNF-α mRNA expression. This supports the view of the immune system as a sensory organ that recognizes invading pathogens and rapidly relays this information to the brain, independent of the nature of the immune response induced. The observation that neuronal and behavioral responses to peripheral immune challenges are not necessarily accompanied by increased brain cytokine expression suggests that cytokines are not the only factors driving sickness-related responses in the CNS.

Acute Lung Injury Induced by Staphylococcal enterotoxin B: Disruption of Terminal Vessels as a Mechanism of Induction of Vascular Leak

The current hypothesis of alveolar capillary membrane dysfunction fails to completely explain the severe and persistent leak of protein-rich fluid into the pulmonary interstitium, seen in the exudative phase of acute lung injury (ALI). The presence of intact red blood cells in the pulmonary interstitium may suggest mechanical failure of pulmonary arterioles and venules. These studies involved the pathological and ultrastructural evaluation of the pulmonary vasculature in Staphylococcal enterotoxin B (SEB)-induced ALI. Administration of SEB resulted in a significant increase in the protein concentration of bronchoalveolar lavage fluid and vascular leak in SEB-exposed mice compared to vehicle-treated mice. In vivo imaging of mice demonstrated the pulmonary edema and leakage in the lungs of SEB-administered mice. The histopathological studies showed intense clustering of inflammatory cells around the alveolar capillaries with subtle changes in architecture. Electron microscopy studies further confirmed the diffuse damage and disruption in the muscularis layer of the terminal vessels. Cell death in the endothelial cells of the terminal vessels was confirmed with TUNEL staining. In this study, we demonstrated that in addition to failure of the alveolar capillary membrane, disruption of the pulmonary arterioles and venules may explain the persistent and severe interstitial and alveolar edema.

A protein superantigen, Staphylococcal Enterotoxin B (SEB), directly activates invariant Natural Killer T cells and leads to Acute Lung Injury and Acute Respiratory Distress Syndrome (ARDS) (89.37)

Abstract Invariant Natural Killer T (iNK T) express T and NK cell markers and secrete a variety of pro- and anti-inflammatory cytokines. Glycolipids such as alpha-galactosylceramide (α-GalCer) are potent activators of iNK T cells when presented by the antigen presenting molecule, CD1d. In this study, we investigated a novel protein antigen, SEB, for the activation of iNK T cells, using ARDS model in mice. In general, SEB activates T cells that posses T cell receptors (TCR) with Vβ8, -7 or -2 chains, and since iNK T cells also have this particular TCR, we hypothesized that SEB can directly activate iNK T cells. We demonstrate that after intranasal administration of SEB, vascular leak and infiltration of lymphocytes were significantly reduced in the lungs of Jα18-/- and CD1d-/- mice as compared to WT mice. The pore size in the endothelial cells of the capillaries of the lung was greater than 500nm for WT mice, while it was between 100 and 500nm for Jα18-/- and Cd1d-/- mice. We also show that cytokine storm induced by SEB was significantly reduced in the serum and bronchialveolar fluid of Jα18-/- and CD1d-/- mice. In vitro cultures that contained SEB-pulsed bone marrow derived dendritic cells (BMDCs) and FACS purified iNK T cells produced significant amounts of IFN-γ and IL-4, comparable to the α-GalCer-pulsed BMDC and iNK T cell cultures. We show for the first time that iNK T cells are activated by a protein antigen, and they are innate responders in Acute Respiratory Distress Syndrome.

Staphylococcal enterotoxin B can activate NKT cells independent of Cd1d

Staphylococcal enterotoxin B (SEB) is a superantigen that can activate 5 to 30% of the T cell population and unlike conventional antigens, it directly binds to the MHC Class II molecules and activates T cells with variable TCR β chain such as Vβ8. Such activation triggers massive cytokine release that leads to rash, fever, and can often lead to multi‐organ failure, coma and death. The precise role of NKT cells in the cytokine production and toxicity induced by SEB is not clear. In this study, we investigated the role of NKT cells following SEB immunization. We found that SEB administration in vivo triggered Vascular Leak Syndrome (VLS) in the lungs of wild‐type (WT) mice while it was significantly decreased in Cd1d−/−mice lacking NKT cells. This was reversed when WT spleenocytes were injected into the Cd1d−/−mice. Cd1d−/−mice also exhibited decreased cytokine production in response to SEB, particularly IL‐4. When bone marrow derived dendritic cells (DCs) from WT and Cd1d−/−mice were tested for their ability to activate NKT cells, we noted that DCs from Cd1d−/−mice were able to activate NKT cells. Together, our studies demonstrate that NKT cells can be directly activated independent of CD1d. Thus, the plethora of cytokines produced following SEB can also be attributed to initial NKT cell activation (Supported in part by NIH grants: AI053703, ES09098, AI058300, DA016545, HL058641 and P01AT003961).

Acute systemic immune activation following vaginal exposure to staphylococcal enterotoxin B—Implications for menstrual shock

Menstrual toxic shock syndrome (mTSS) is an acute systemic inflammatory disease associated with the superantigenic exotoxin, toxic shock syndrome toxin (TSST)-1, produced by Staphylococcus aureus and the use of high absorbency tampons. Even though S. aureus is capable of elaborating several other superantigenic exotoxins, only TSST-1 has been implicated in the pathogenesis of mTSS possibly because most other superantigenic exotoxins are known enterotoxins. Nonetheless, we have shown recently that one of the enterotoxigenic staphylococcal superantigens, staphylococcal enterotoxin B (SEB), can cause robust systemic immune activation following exposure through non-enteric mucosa, including nasal or conjunctival routes. In a similar manner, we show here that vaginal administration of SEB in HLA class II transgenic mice can cause robust systemic immune activation characterized by profound elevation of proinflammatory cytokines in the serum, activation and expansion of SEB-reactive CD4 + and CD8 + T cells in peripheral lymphoid organs and SEB-induced deletion of immature thymocytes. Vaginal administration of SEB also caused leukocytic infiltration in major organs, such as liver and lung, reminiscent of human toxic shock syndrome. Systemic immune activation following vaginal superantigen delivery was independent of the stage of the estrus cycle in the mouse. Using HLA class II transgenic mice, we have shown that exposure to SEB through the vaginal canal can cause robust systemic immune activation. SEB could thus play a role in the pathogenesis of mTSS.

INTRANASAL EXPOSURE TO STAPHYLOCOCCAL ENTEROTOXIN B ELICITS AN ACUTE SYSTEMIC INFLAMMATORY RESPONSE

Staphylococcus aureus produces a variety of superantigen exotoxins, including staphylococcal enterotoxin B (SEB). Little is known regarding the pathogenesis of SEB entering through the intranasal route. Intranasal exposure to SEB might occur because of nasal packing following surgical procedure, biologic warfare, or even S. aureus colonization. We evaluated the local and systemic effects of intranasally delivered SEB using a series of human leukocyte antigen (HLA) class II transgenic mice as conventional mice expressing endogenous class II molecules mount a poor immune response to SEB. Gene expression profiling using microarrays showed robust up-regulation of genes involved in several proinflammatory pathways as early as 3 h post-intranasal challenge with SEB in HLA class II transgenic mice. This was accompanied by a several hundred-fold increase in serum levels of pro-inflammatory cytokines such as IL-12, IL-6, TNF-alpha, IFN-gamma, as well as MCP-1 in HLA class II transgenic mice but not in C57BL/6 mice; CD4 or CD8 T-cells independently contributed to the systemic cytokine response. Defective IL-12 or IL-4 receptor signaling significantly decreased or increased serum IFN-gamma, respectively. Intranasal exposure to SEB resulted in neutrophil influx into bronchoalveolar lavage fluid and caused expansion of both CD4 and CD8 T-cells expressing TCR V beta 8 in the spleen. This was accompanied by mononuclear cell infiltration in the liver reminiscent of the systemic inflammatory response syndrome. Thus, we have shown, for the first time, that intranasal administration of SEB can cause systemic immune activation.

Role of CD44 and Its v7 Isoform in Staphylococcal Enterotoxin B-Induced Toxic Shock: CD44 Deficiency on Hepatic Mononuclear Cells Leads to Reduced Activation-Induced Apoptosis That Results in Increased Liver Damage

Exposure to bacterial superantigens such as staphylococcal enterotoxin B (SEB) leads to the induction of toxic shock syndrome which results in multiorgan failure, including liver damage. In the present study, we investigated the role of CD44 in SEB-induced liver injury. Injection of SEB into d-galactosamine-sensitized CD44 wild-type (WT) mice led to a significant increase in CD44 expression on liver T cells, NK cells, and NKT cells. Administration of SEB to CD44 knockout (KO) mice caused significantly enhanced liver damage which correlated with elevated numbers of T cells, NK cells, NKT cells, and macrophages in the liver and increased production of tumor necrosis factor alpha and gamma interferon compared to CD44 WT mice. Furthermore, liver mononuclear cells from CD44 KO mice were resistant to SEB-induced apoptosis, and cDNA microarray analysis revealed that SEB activation of such cells led to the induction of several antiapoptotic genes and repression of proapoptotic genes. Examination of CD44 isoforms revealed that SEB exposure altered CD44 variant 7 (v7) isoform expression. Interestingly, mice bearing a specific deletion of the CD44v7 exon exhibited increased susceptibility to SEB-induced hepatitis. Finally, treatment of CD44 WT mice with anti-CD44 monoclonal antibodies reduced expression of CD44 in liver mononuclear cells and caused increased susceptibility to SEB-induced liver injury. Together, these data demonstrate that the expression of CD44 and/or CD44v7 on SEB-activated liver mononuclear cells facilitates their rapid apoptosis, thereby preventing severe liver injury in wild-type mice, and suggest that CD44 plays an important role in the regulation and elimination of immune cells in the liver.

Behavioural endocrine immune-conditioned response is induced by taste and superantigen pairing

Administration of bacterial superantigen, such as staphylococcal enterotoxin B (SEB), induces in vivo stimulation of T cell proliferation and cytokine production such as interleukin-2 (IL-2). It has been previously reported that SEB administration induces fever, c-Fos expression in the brain, and hypothalamus–pituitary–adrenal axis activation, demonstrating that the brain is able to sense and respond to SEB. Previously it had been shown that immune functions can be behaviourally conditioned pairing a novel gustatory stimulus together with an immunomodulatory drug or an antigen. We designed an experimental protocol using Dark Agouti rats in which saccharin taste, as conditioned stimulus, was paired with an i.p. injection of SEB (2 mg/kg), as unconditioned stimulus. Six days later, when conditioned animals were re-exposed to the conditioned stimulus they displayed strong conditioned taste avoidance to the saccharin. More importantly, re-exposure to the conditioned stimulus significantly increased IL-2, interferon-γ and corticosterone plasma levels, in comparison with conditioned animals which had not been re-exposed to saccharin taste. These results demonstrate a behavioural-immune-endocrine conditioned response using a superantigen as unconditioned stimulus. In addition, they illustrate the brain abilities to mimic the unconditioned effects of a superantigen by yet unknown mechanisms.

Flare-up reaction on murine contact hypersensitivity: III. Effect of staphylococcal enterotoxin B

Staphylococcal enterotoxin B (SEB), a bacterial superantigen, is known as an immunomodulator because it activates an extremely large number of T-cells, and induces the production of large amounts of cytokines. In this study, we examined the effects of SEB on the contact hypersensitivity reaction (CHR). BALB/c mice were first sensitized through haptens applied to the back, and CHR was then induced through challenge to the left ear using the same haptens. SEB was administered intravenously 4 weeks later, causing a flare-up, peaking at 24 h post-administration, in the left ear that had previously exhibited CHR. This flare-up reaction was hapten non-specific, and was inhibited by anti-mouse tumor necrosis factor (TNF)-α antibodies. The flare-up was also suppressed by the oral administration of cyclosporin A prior to the administration of SEB. These results suggest that SEB induces a flare-up of CHR via the production of TNF-α.

In vivo cytokine responses in gut-associated lymphoid tissue (GALT) and spleen following oral administration of staphylococcal enterotoxin B

The mechanisms responsible for oral tolerance have been analyzed using various antigens. The induction of oral nonresponsiveness to low doses of Staphylococcal Enterotoxin B (SEB), in which SEB-reactive clonal anergy and deletion are involved, has also been confirmed. However, few reports have examined the cytokine milieu of the mucosal and peripheral lymphoid tissue during tolerance induction after the oral administration of SEB. In this study, to elucidate the mechanism responsible for the oral tolerance to low dose of SEB, the cytokine responses of the gut-associated lymphoid tissues (GALT) and the spleen were examined using the reverse transcription-polymerase chain reaction (RT-PCR) method. After the oral administration of a single low dose of SEB, the expressions of Th2 and TGF-β, and to a lesser extent Th1 mRNA were observed in the mucosa. In contrast, following repeated oral low doses of SEB, in a manner different from previous results using conventional antigens, no cytokine enhancements were demonstrated in the GALT or the spleen. In conclusion, the involvement of unknown inhibitory or regulatory cytokines, together with SEB-reactive clonal anergy and deletion, is suggested in the induction of oral tolerance to low dose SEB.