3.4.24.83	diagnostics	the anthrax lethal toxin neutralization assay (TNA) measures the ability of antibodies to neutralize the cytotoxicity of anthrax lethal toxin rather than quantifying total antibody through a conjugated species-specific secondary antibody. TNA may provide a more relevant immunological measure, as it quantitates functional antibodies only rather than total protective antigen-binding antibodies. In this study, TNA data are generated in several different laboratories to measure the immune responses in rabbits, nonhuman primates, and humans. A collaborative study is conducted in which 108 samples from the three species are analyzed in seven independent laboratories. This study demonstrates that the TNA is a panspecies assay that can be performed in several different laboratories with a high degree of quantitative agreement and precision	697440	
3.4.24.83	diagnostics	the development, performance characteristics and validation using human serum of a robust and rugged format of the LTx neutralization activity (TNA) assay is reported and its application in evaluating immune serum from humans, Rhesus macaques and rabbits. This format uses standardized and characterized reagents in conjunction with customized interpretive software and a novel mathematical algorithm to calculate and extrapolate multiple reportable values, in addition to ED50, with high specificity, analytical sensitivity, accuracy and precision. This LTx neutralization activity (TNA) assay format is proposed as a unifying platform technology	696000	
3.4.24.83	medicine	administration of lethal factor proteases early in the infection inhibits dissemination of vegetative bacteria to the organs in the first 32 h following infection. Neutralizing antibodies against edema factor also inhibit bacterial dissemination with similar efficacy	733112	
3.4.24.83	medicine	anthrax lethal factor is a specific biomarker of active infection by Bacillus anthracis	718573	
3.4.24.83	medicine	anthrax toxin entry and activity differs among immune cells. Macrophages, dendritic cells, and B cells display higher activity of a of fusion protein of the anthrax toxin lethal factor N-terminal domain LFn, residues 1-254, with beta-lactamase, i.e. LFnBLA, than CD4+ and CD8+ T cells in both spleen cell suspension and the purified samples of individual cell types. Expression of anthrax toxin receptor CMG2 is higher in CD4+ and CD8+ T cells, which is not correlated to the intracellular LFnBLA activity	713352	
3.4.24.83	medicine	blood borne lethal toxin is a novel therapeutic target for combating anthrax	720419	
3.4.24.83	medicine	engineered lethal anthrax toxin prevents tumor growth by inhibiting angiogenesis (10 nmol/l engineered protective antigen + 5.5 nmol/l lethal factor)	700136	
3.4.24.83	medicine	human medical countermeasures for anthrax	669066	
3.4.24.83	medicine	in a murine model of intoxication, lethal factor causes the dose-dependent disruption of intestinal epithelial integrity, characterized by mucosal erosion, ulceration, and bleeding. The pathology correlates with a blockade of intestinal crypt cell proliferation, accompanied by marked apoptosis in the villus tips. Treated mice nearly uniformly develop systemic infections with commensal enteric organisms within 72 hours of administration. Intestinal pathology depends upon lethal factor proteolytic activity and is partially attenuated by co-administration of broad spectrum antibiotics	735046	
3.4.24.83	medicine	inducing strong mucosal and systemic immune responses against both anthrax toxins and bacilli after nasal immunization using a synthetic double-stranded RNA (dsRNA), polyriboinosinic-polyribocytidylic acid as adjuvant. The capsular poly-gamma-D-glutamic acid (PGA) from bacillus is immunogenic when conjugated to a carrier protein and dosed intranasally to mice. The nasal immunization with the poly-gamma-D-glutamic acid-carrier protein conjugate in combination with the anthrax protective antigen (PA) protein induces both anti-PGA and anti-PA immune responses in mouse sera and lung mucosal secretions. The anti-PA antibody response is shown to have anthrax lethal toxin neutralization activity. The anti-PGA Abs induced are able to activate complement and kill PGA-producing bacteria. It is feasible to develop a novel dual-action nasal anthrax vaccine	670987	
3.4.24.83	medicine	mice immunized with chloroplast-derived anthrax protective antigen survive anthrax lethal toxin challenge	668928	
3.4.24.83	medicine	neuronal nitric oxide synthase deficiency in mice causes strikingly increased sensitivity to anthrax lethal toxin, while deficiencies in NOS enzymes iNOS and eNOS have no effect on anthrax lethal toxin-mediated mortality. The increased sensitivity of nNOS2/2 mice is independent of macrophage sensitivity to toxin, or cytokine responses, and can be replicated in nNOS-sufficient wild-type mice through pharmacological inhibition of the enzyme with 7-nitroindazole. Anthrax lethal toxin induces architectural changes in heart morphology of nNOS2/2 mice, with rapid appearance of novel inter-fiber spaces but no associated apoptosis of cardiomyocytes. Anthrax lethal toxin-treated wild-type mice have no histopathology observed at the light microscopy level. Electron microscopic analyses reveal striking pathological changes in the hearts of both nNOS2/2 and wild-type mice, varying only in severity and timing. Endothelial andcapillary necrosis and degeneration, inter-myocyte edema, myofilament and mitochondrial degeneration, and altered sarcoplasmic reticulum cisternae are observed in both anthrax lethal toxin-treated wild-type and nNOS2/2 mice. Biomarkers of cardiac injury, myoglobin, cardiac troponin-I, and heart fatty acid binding protein, are elevated in anthrax lethal toxin-treated mice very rapidly and reach concentrations rarely reported in mice. The potent nitric oxide scavenger, 2-(4-carboxyphenyl)-4,5-dihydro-4,4,5,5-tetramethyl-1H-imidazolyl-1-oxy-3-oxide, i.e. carboxy-PTIO, shows some protective effect against anthrax lethal toxin	713382	
3.4.24.83	medicine	seven of eleven acute myeloid leukemia cell lines show cytotoxic responses to anthrax lethal toxin LeTx. Cytotoxicity is mimicked by the specific mitogen-activated protein/extracellular signal-regulated kinase kinase 1/2 inhibitor U0126, indicating involvement of the ERK1/2 branch of the MAPK pathway. The four LeTx-resistant cell lines are sensitive to the phosphatidylinositol 3-kinase inhibitor LY294002. with a lack of additive/synergistic effects when both pathways are inhibited. Phospho-ERK1/2 is only present in LeTx-sensitive cells. LeTx-induced cell death is caspase-independent and nonapoptotic	735320	
3.4.24.83	medicine	sublethal doses of Bacillus anthracis lethal toxin inhibit inflammation with lipopolysaccharide and Escherichia coli challenge but have opposite effects on survival	669743	
3.4.24.83	medicine	the antitumor toxin has potential for use in cancer therapy	680918	
3.4.24.83	medicine	the use of drugs capable of inhibiting Rho GTPase activity, such as statins, may provide a means to attenuate intoxication during Bacillus anthracis infection	698254	
3.4.24.83	molecular biology	anthrax lethal toxin treatment of neutrophils disrupts signaling to downstream MAPK targets in response to TLR stimulation. Following anthrax lethal toxin treatment, ERK family and p38 phosphorylation are nearly completely blocked, but signaling to JNK family members persists in vitro and ex vivo. In contrast to previous reports involving human neutrophils, anthrax lethal toxin treatment of murine neutrophils increases their production of superoxide in response to PMA or TLR stimulation in vitro or ex vivo. Although this enhanced superoxide production correlates with effects due to the lethal toxin-induced blockade of ERK signaling, it requires JNK signaling that remains largely intact despite the activity of anthrax lethal toxin	699322	
3.4.24.83	molecular biology	Bacillus anthracis represses the immune response, in part by altering chromatin accessibility of IL-8 promoter to NFkappaB in epithelial cells. This epigenetic reprogramming, in addition to previously reported effects of lethal toxin, represents an efficient strategy used by Bacillus anthracis for invading the host	700915	
3.4.24.83	molecular biology	celastrol is identified as an inhibitor of lethal toxin-mediated macrophage lysis and suggests an inhibitory mechanism involving inhibition of the proteasome pathway	700878	
3.4.24.83	molecular biology	it is shown that treatment of RAW 264.7 murine macrophage cells with anthrax lethaltoxin induces autophagy suggesting a protective role as autophagy inhibition using 3-methyladenine results in an accelerated cell death	696028	
3.4.24.83	molecular biology	lethal toxin triggers the formation of a membrane-associated inflammasome complex in murine macrophages consisting of caspase-1 and Nalp1b, resulting in cleavage of cytosolic caspase-1 substrates and cell death	698250	
3.4.24.83	molecular biology	microarray analysis is used to investigate the effects of Bacillus anthracis lethal toxin on human neutrophil-like NB-4 cells to identify markers of intoxication. Genes down-regulated after a 2 h lethal toxin exposure include those encoding chemokines and transcription factors. Significant decreases in the mRNA of interleukin-8, CCL20, CCL3 and CCL4 are observed using real-time PCR. The decreases are more pronounced at 4 and 8 h and are lethal toxin-specific. Decreases in chemokine protein levels are evident after 24 h and are sensitive to low concentrations of lethal toxin. Co-incubation with an anti-lethal factor mAb restores levels of interleukin-8 to 100% and 50%, respectively	696000	
3.4.24.83	molecular biology	primary keratinocytes are resistant to LeTx cytotoxicity, and MEK cleavage does not correlate with LeTx cytotoxicity. LeTx is considered as an anti-inflammatory agent, however it upregulates RANTES	698522	
3.4.24.83	molecular biology	proteasome inhibitors block anthrax lethal toxin-mediated caspase-1 activation and can protect against cell death, indicating that the degradation of at least one cellular protein is required for cell death. Proteins can be degraded by the proteasome via the N-end rule. Using amino acid derivatives that act as inhibitors of this pathway, it is shown that the N-end rule is required for anthrax lethal toxin-mediated caspase-1 activation and cell death. The Streptomyces olivoreti peptide bestatin, which inhibits leucine, alanine and arginine aminopeptidases, protects macrophages against anthrax lethal toxin. c-IAP1, a mammalian member of the inhibitor of apoptosis protein (IAP) family is identified, as a novel N-end rule substrate degraded in macrophages treated with anthrax lethal toxin	697197	
3.4.24.83	molecular biology	protein expression profile of murine macrophages RAW264.7 treated with LeTx is analyzed using two-dimensional polyacrylamide gel electrophoresis and MALDI-TOF MS. Among the differentially expressed spots, cleaved mitogen-activated protein kinase kinase 1 acting as a negative element in the signal transduction pathway, and glucose-6-phosphate dehydrogenase playing a role in the protection of cells from hyperproduction of active oxygen are up-regulated LeTx-treated macrophages	696413	
3.4.24.83	molecular biology	results suggest that this toxin delivery system is capable of stimulating protective immune responses where effective immunization requires stimulation of both classes of T cells	698243	
3.4.24.83	molecular biology	the cellular damage inflicted by anthrax lethal toxin depends not only on the innate responses but also on the maturation stage of the cell, which modulates the more general caspase-1-independent responses	697196	
3.4.24.83	molecular biology	the effects of lethal toxin on the transcriptional regulation of the VCAM1 gene, which contains binding sites in its promoter region for NF-kappaB, IFN regulatory factor-1 (IRF-1), Sp1, GATA-2, and AP-1, in primary human endothelial cells is examined. Lethal toxin enhances cytokine-induced activation of NF-kappaB and IRF-1 which are key factors in the lethal toxin-mediated enhancement of TNF-induced VCAM-1 expression. Altering the activity of key transcription factors involved in host response to infection may be a critical mechanism by which lethal toxin contributes to anthrax pathogenesis	699325	
3.4.24.83	molecular biology	the in vitro effects of thermal stress on the killing of murine macrophages by anthrax lethal toxin are investigated. Heat shock rapidly halts anthrax lethal toxin-induced cell death without any impact on toxin uptake or mitogen-activated protein kinases cleavage, by a mechanism independent of novel protein synthesis, p38 activation, HSP90 activity or proteasome inhibition. Rather, heat shock prevents the activation of procaspase-1 in anthrax lethal toxin -treated cells, apparently by the sequestration of pro-caspase-1 in a large, inhibitory complex. Heat-shocked cell lysates strongly inhibit the active caspase-1 heterotetramer in vitro, independent of a specific inflammasome platform. Results suggest the presence of a cellular, heat shock-inducible, caspase-1 inhibiting factor	697199	
3.4.24.83	molecular biology	toxin effects of lethal toxin and edema toxin of Bacillus anthracis in bone marrow dendritic cells stimulated with either LPS or Legionella pneumophila are analysed. Lethal toxin, not ET, is more toxic for cells from BALB/c mice than from C57BL/6 as measured by 7-AAD uptake. Results support the conclusion that lethal toxin and edema toxin are not uniformly suppressive of dendritic cell function but rather modulate function up or down depending on variables such as the function tested, the microbial stimulus used, and the genetic variation in innate immune response mechanisms in the host cell	697576	
3.4.24.83	synthesis	preparation of semisynthetic protective antigen-binding domain of anthrax lethal factor, LFN, by native chemical ligation of synthetic LFN residues 14-28 thioester with recombinant N29C-LFN residues 29-263 and comparison with two variants containing alterations in residues 14-28 of the N-terminal region. The properties of the variants in blocking ion conductance through the protective antigen pore and translocating across planar phospholipid bilayers in response to a pH gradient are consistent with current concepts of the mechanism of polypeptide translocation through the pore. The semisynthesis platform allows for investigation of the interaction of the pore with its substrates	710698