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Staphylococcal Enterotoxin B (staphylococcal + enterotoxin_b)
Selected AbstractsImproved Methods for Prepurification and Detection of Staphylococcal Enterotoxin B from Cell-Free Culture FiltrateBIOTECHNOLOGY PROGRESS, Issue 4 2005Maria Dainiak An improved ELISA method for the detection of Staphylococcal Enterotoxin B (SEB) in protein A preparations is presented. Fab fragments were obtained by digestion with papain of anti-SEB IgG bound to SEB immobilized on Sepharose 4B. Anti-SEB and peroxidase-labeled Fab fragments from secondary antibodies were successfully used in a modified ELISA of SEB in protein A preparations. SEB-Sepharose was used repeatedly for the production of anti-SEB Fab fragments by papain digestion without loss of affinity. In addition, for the purification of SEB from crude culture filtrates, an initial step utilizing a combined heat and pH treatment for the removal of significant amounts of contaminating proteins without losses of toxin activity is presented. This pretreatment step yielded positive effects in further downstream processing considering both shortened time and an increase in total recovery. [source] Effect of 5-lipoxygenase inhibitor MK591 on early molecular and signaling events induced by staphylococcal enterotoxin B in human peripheral blood mononuclear cellsFEBS JOURNAL, Issue 12 2008Chanaka Mendis Staphylococcal enterotoxin B (SEB) has been the focus of a number of studies due to its ability to promote septic shock and a massive impact on the human immune system. Even though symptoms and pathology associated with SEB is well known, early molecular events that lead to lethality are still poorly understood. Our approach was to utilize SEB induced human peripheral blood mononuclear cells (PBMCs) as a prototype module to further investigate the complexity of signaling cascades that may ultimately lead to lethal shock. Our study revealed the activation of multiple divergent intracellular pathways within minutes of SEB induction including components that interconnect investigated pathways. A series of performed inhibitor studies identified a specific inhibitor of 5-LO (MK591), which has the ability to block JNK, MAPK, p38kinase and 5-LO signaling-cascades and drastically reducing the activity of pro-inflammatory cytokine TNF-,. Further evaluation of MK591 utilizing cell proliferation assays in PBMCs, human proximal tubule cells and in vivo studies (monkey) showed a decrease in cell proliferation. The inhibitory effect of MK591 was reconfirmed at a genetic level through the utilization of a set of SEB specific genes. Signaling activities, inhibitor studies, cellular analysis and gene expression analysis in unison illustrated the significance of pathway interconnectors such as 5-LO as well as inhibiting such inter-connectors (using MK591) in SEB induced human PBMCs. [source] T-cell tolerance induced by repeated antigen stimulation: Selective loss of Foxp3, conventional CD4 T cells and induction of CD4 T-cell anergyEUROPEAN JOURNAL OF IMMUNOLOGY, Issue 4 2009Lena Eroukhmanoff Abstract Repeated immunization of mice with bacterial superantigens induces extensive deletion and anergy of reactive CD4 T cells. Here we report that the in vitro proliferation anergy of CD4 T cells from TCR transgenic mice immunized three times with staphylococcal enterotoxin B (SEB) (3× SEB) is partially due to an increased frequency of Foxp3+ CD4 T cells. Importantly, reduced number of conventional CD25, Foxp3, cells, rather than conversion of such cells to Foxp3+ cells, was the cause of that increase and was also seen in mice repeatedly immunized with OVA (3× OVA) and OVA,peptide (OVAp) (3× OVAp). Cell-transfer experiments revealed profound but transient anergy of CD4 T cells isolated from 3× OVAp and 3× SEB mice. However, the in vivo anergy was CD4 T-cell autonomous and independent of Foxp3+ Treg. Finally, proliferation of transferred CD4 T cells was inhibited in repeatedly immunized mice but inhibition was lost when transfer was delayed, despite the maintenance of elevated frequency of Foxp3+ cells. These data provide important implications for Foxp3+ cell-mediated tolerance in situations of repeated antigen exposure such as human persistent infections. [source] Effect of 5-lipoxygenase inhibitor MK591 on early molecular and signaling events induced by staphylococcal enterotoxin B in human peripheral blood mononuclear cellsFEBS JOURNAL, Issue 12 2008Chanaka Mendis Staphylococcal enterotoxin B (SEB) has been the focus of a number of studies due to its ability to promote septic shock and a massive impact on the human immune system. Even though symptoms and pathology associated with SEB is well known, early molecular events that lead to lethality are still poorly understood. Our approach was to utilize SEB induced human peripheral blood mononuclear cells (PBMCs) as a prototype module to further investigate the complexity of signaling cascades that may ultimately lead to lethal shock. Our study revealed the activation of multiple divergent intracellular pathways within minutes of SEB induction including components that interconnect investigated pathways. A series of performed inhibitor studies identified a specific inhibitor of 5-LO (MK591), which has the ability to block JNK, MAPK, p38kinase and 5-LO signaling-cascades and drastically reducing the activity of pro-inflammatory cytokine TNF-,. Further evaluation of MK591 utilizing cell proliferation assays in PBMCs, human proximal tubule cells and in vivo studies (monkey) showed a decrease in cell proliferation. The inhibitory effect of MK591 was reconfirmed at a genetic level through the utilization of a set of SEB specific genes. Signaling activities, inhibitor studies, cellular analysis and gene expression analysis in unison illustrated the significance of pathway interconnectors such as 5-LO as well as inhibiting such inter-connectors (using MK591) in SEB induced human PBMCs. [source] Proteomic analysis of exoproteins expressed by enterotoxigenic Staphylococcus aureus strainsPROTEINS: STRUCTURE, FUNCTION AND BIOINFORMATICS, Issue 12 2008Gabriella Pocsfalvi Dr. Abstract Pathogenic bacteria excrete a variety of virulence factors into extracellular medium and to the cell surface which have essential roles in the colonization and insurrection of the host cells, and thus reflect the degree of bacterial pathogenicity. For the exploration of virulence factors expressed in the secreted proteome fraction, different Staphylococcus aureus strains were analyzed using gel-based bottom-up proteomic approach. A total of 119 distinct proteins were identified for the enterotoxin gene cluster (egc) negative and seb gene positive S. aureus American Type Culture Collection (ATCC) 14458 strain by the use of one- and 2-DE based proteomics. Detailed analysis of enterotoxin region of the 2-D map confirmed, beside the highly expressed staphylococcal enterotoxin B (SEB), the presence of enterotoxin-like proteins SElK and SElQ previously predicted by genotyping (Sergeev et al.., J. Clin. Microbiol. 2004, 42, 2134,2143). Exoprotein patterns at the late-exponential (7,h) and stationary (24,h) phases of cellular growth show a high-level similarity in this region. Comparative analysis of enterotoxin region of five S. aureus strains including two clinical isolates (RIMD 31092 and A900322), a food derived strain (AB-8802) with highly prevalent egc positive operon and a nonenterotoxigenic reference strain (ROS) revealed the presence of different known enterotoxins and other virulence factors along with a number of core exoproteins. In addition, production of SElL (RIMD 31092) and SElP (A900322) was demonstrated for the first time at the protein level. Under the experimental conditions applied none of the enterotoxins encoded by the genes of egc operon was identified. [source] Effects of two novel cationic staphylococcal proteins (NP-taseand p70) and enterotoxin B on IgE synthesis and interleukin-4 and interferon-, production in patients with atopic dermatitisBRITISH JOURNAL OF DERMATOLOGY, Issue 4 2000A. Jahreis We have characterized the cell-mediated and humoral immune response of patients with atopic dermatitis (AD) and healthy controls in response to two novel staphylococcal antigens (NP-tase, p70) and the superantigen staphylococcal enterotoxin B (SEB). The parameters studied were IgE, interleukin (IL)-4 and interferon (IFN)-, synthesis by peripheral blood mononuclear cells (PBMC) after stimulation with NP-tase, p70 and SEB in vitro. Both antigens, as well as SEB, induced IL-4 and IFN-, secretion in patients and controls. However, patients with AD showed a significantly diminished IFN-, production in response to NP-tase or SEB. Furthermore, we demonstrated a good correlation between antigen-stimulated IgE production and the IL-4/IFN-, ratio in vitro. A distinct subgroup of PBMC showed impaired IFN-, synthesis and enhanced IL-4 secretion after incubation with p70 or NP-tase. These data support evidence that a subgroup of patients with AD, synthesizing low levels of IFN-, after stimulation with staphylococcal antigens, may have impaired abilities to clear Staphylococcus aureus colonization. Persistent staphylococcal antigens could then be responsible for inflammatory and allergic skin reactions in patients with AD. We therefore conclude that, besides superantigens, staphylococcal antigens may also play a part in the pathogenesis of AD. [source] A hexamer peptide ligand that binds selectively to staphylococcal enterotoxin B: isolation from a solid phase combinatorial libraryCHEMICAL BIOLOGY & DRUG DESIGN, Issue 2 2004G. Wang Abstract:, By screening a solid-phase combinatorial peptide library, a short peptide ligand, YYWLHH, has been discovered that binds with high affinity and selectivity to staphylococcal enterotoxin B (SEB), but only weakly to other SEs that share sequence and structural homology with SEB. Using column affinity chromatography with an immobilized YYWLHH stationary phase, it was possible to separate SEB quantitatively from Staphylococcus aureus fermentation broth, a complex mixture of proteins, carbohydrates and other biomolecules. The immobilized peptide was also used to purify native SEB from a mixture containing denatured and hydrolyzed SEB, and showed little cross-reactivity with other SEs. To our knowledge this is the first report of a highly specific short peptide ligand for SEB. Such a ligand is a potential candidate to replace antibodies for detection, removal and purification strategies for SEB. [source] | ||||||||||||||||