Browsing by Subject "Lipopolysaccharides"
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Item Open Access Anti-inflammatory effects of progesterone in lipopolysaccharide-stimulated BV-2 microglia.(PLoS One, 2014) Lei, Beilei; Mace, Brian; Dawson, Hana N; Warner, David S; Laskowitz, Daniel T; James, Michael LFemale sex is associated with improved outcome in experimental brain injury models, such as traumatic brain injury, ischemic stroke, and intracerebral hemorrhage. This implies female gonadal steroids may be neuroprotective. A mechanism for this may involve modulation of post-injury neuroinflammation. As the resident immunomodulatory cells in central nervous system, microglia are activated during acute brain injury and produce inflammatory mediators which contribute to secondary injury including proinflammatory cytokines, and nitric oxide (NO) and prostaglandin E2 (PGE2), mediated by inducible NO synthase (iNOS) and cyclooxygenase-2 (COX-2), respectively. We hypothesized that female gonadal steroids reduce microglia mediated neuroinflammation. In this study, the progesterone's effects on tumor necrosis factor alpha (TNF-α), iNOS, and COX-2 expression were investigated in lipopolysaccharide (LPS)-stimulated BV-2 microglia. Further, investigation included nuclear factor kappa B (NF-κB) and mitogen activated protein kinase (MAPK) pathways. LPS (30 ng/ml) upregulated TNF-α, iNOS, and COX-2 protein expression in BV-2 cells. Progesterone pretreatment attenuated LPS-stimulated TNF-α, iNOS, and COX-2 expression in a dose-dependent fashion. Progesterone suppressed LPS-induced NF-κB activation by decreasing inhibitory κBα and NF-κB p65 phosphorylation and p65 nuclear translocation. Progesterone decreased LPS-mediated phosphorylation of p38, c-Jun N-terminal kinase and extracellular regulated kinase MAPKs. These progesterone effects were inhibited by its antagonist mifepristone. In conclusion, progesterone exhibits pleiotropic anti-inflammatory effects in LPS-stimulated BV-2 microglia by down-regulating proinflammatory mediators corresponding to suppression of NF-κB and MAPK activation. This suggests progesterone may be used as a potential neurotherapeutic to treat inflammatory components of acute brain injury.Item Open Access Beta-arrestin-2 regulates the development of allergic asthma.(J Clin Invest, 2003-08) Walker, Julia KL; Fong, Alan M; Lawson, Barbara L; Savov, Jordan D; Patel, Dhavalkumar D; Schwartz, David A; Lefkowitz, Robert JAsthma is a chronic inflammatory disorder of the airways that is coordinated by Th2 cells in both human asthmatics and animal models of allergic asthma. Migration of Th2 cells to the lung is key to their inflammatory function and is regulated in large part by chemokine receptors, members of the seven-membrane-spanning receptor family. It has been reported recently that T cells lacking beta-arrestin-2, a G protein-coupled receptor regulatory protein, demonstrate impaired migration in vitro. Here we show that allergen-sensitized mice having a targeted deletion of the beta-arrestin-2 gene do not accumulate T lymphocytes in their airways, nor do they demonstrate other physiological and inflammatory features characteristic of asthma. In contrast, the airway inflammatory response to LPS, an event not coordinated by Th2 cells, is fully functional in mice lacking beta-arrestin-2. beta-arrestin-2-deficient mice demonstrate OVA-specific IgE responses, but have defective macrophage-derived chemokine-mediated CD4+ T cell migration to the lung. This report provides the first evidence that beta-arrestin-2 is required for the manifestation of allergic asthma. Because beta-arrestin-2 regulates the development of allergic inflammation at a proximal step in the inflammatory cascade, novel therapies focused on this protein may prove useful in the treatment of asthma.Item Open Access Brief Glutamine Pretreatment Increases Alveolar Macrophage CD163/Heme Oxygenase-1/p38-MAPK Dephosphorylation Pathway and Decreases Capillary Damage but Not Neutrophil Recruitment in IL-1/LPS-Insufflated Rats.(PLoS One, 2015) Fernandez-Bustamante, Ana; Agazio, Amanda; Wilson, Paul; Elkins, Nancy; Domaleski, Luke; He, Qianbin; Baer, Kaily A; Moss, Angela FD; Wischmeyer, Paul E; Repine, John EBACKGROUND: Glutamine (GLN) attenuates acute lung injury (ALI) but its effect on alveolar macrophages is unknown. We hypothesized that GLN pretreatment would induce the anti-inflammatory CD163/heme oxygenase (HO)-1/p38-MAPK dephosphorylation pathway in alveolar macrophages and reduce ALI in rats insufflated with interleukin-1 (IL-1) and lipopolysaccharide (LPS). METHODS: Male Sprague-Dawley rats were randomized to the following groups: GLN-IL-1/LPS-, GLN+IL-1/LPS-, GLN-IL-1/LPS+, and GLN+IL-1/LPS+. GLN pretreatment was given via gavage (1 g/kg L-alanyl-L-glutamine) daily for 2 days. ALI was subsequently induced by insufflating 50 ng IL-1 followed by 5mg/kg E.coli LPS. After 24h, bronchoalveolar lavage (BAL) protein, lactate dehydrogenase (LDH) and neutrophil concentrations were analyzed. BAL alveolar macrophage CD163+ expression, HO-1 and p38-MAPK concentrations were measured, as well as alveolar macrophage tumor necrosis factor (TNF)-α and interleukin (IL)-10 concentrations. Histology and immunofluorescence studies were also performed. RESULTS: Following IL-1/LPS insufflation, GLN pretreated rats had significantly decreased BAL protein and LDH concentrations, but not BAL neutrophil counts, compared to non-GLN pretreated rats. The number of alveolar macrophages and the number of CD163+ macrophages were significantly increased in GLN pretreated IL-1/LPS-insufflated rats compared to non-GLN pretreated, IL-1/LPS-insufflated rats. GLN pretreatment before IL-1/LPS also significantly increased HO-1 concentrations and dephosphorylated p38-MAPK levels but not cytokine levels in alveolar macrophages. Immunofluorescence localized CD163 and HO-1 in alveolar macrophages. CONCLUSION: Short-term GLN pretreatment activates the anti-inflammatory CD163/HO-1/p38-MAPK dephosphorylation pathway of alveolar macrophages and decreases capillary damage but not neutrophil recruitment in IL-1/LPS-insufflated rats.Item Open Access Co-regulation of nuclear respiratory factor-1 by NFkappaB and CREB links LPS-induced inflammation to mitochondrial biogenesis.(J Cell Sci, 2010-08-01) Suliman, Hagir B; Sweeney, Timothy E; Withers, Crystal M; Piantadosi, Claude AThe nuclear respiratory factor-1 (NRF1) gene is activated by lipopolysaccharide (LPS), which might reflect TLR4-mediated mitigation of cellular inflammatory damage via initiation of mitochondrial biogenesis. To test this hypothesis, we examined NRF1 promoter regulation by NFκB, and identified interspecies-conserved κB-responsive promoter and intronic elements in the NRF1 locus. In mice, activation of Nrf1 and its downstream target, Tfam, by Escherichia coli was contingent on NFκB, and in LPS-treated hepatocytes, NFκB served as an NRF1 enhancer element in conjunction with NFκB promoter binding. Unexpectedly, optimal NRF1 promoter activity after LPS also required binding by the energy-state-dependent transcription factor CREB. EMSA and ChIP assays confirmed p65 and CREB binding to the NRF1 promoter and p65 binding to intron 1. Functionality for both transcription factors was validated by gene-knockdown studies. LPS regulation of NRF1 led to mtDNA-encoded gene expression and expansion of mtDNA copy number. In cells expressing plasmid constructs containing the NRF-1 promoter and GFP, LPS-dependent reporter activity was abolished by cis-acting κB-element mutations, and nuclear accumulation of NFκB and CREB demonstrated dependence on mitochondrial H(2)O(2). These findings indicate that TLR4-dependent NFκB and CREB activation co-regulate the NRF1 promoter with NFκB intronic enhancement and redox-regulated nuclear translocation, leading to downstream target-gene expression, and identify NRF-1 as an early-phase component of the host antibacterial defenses.Item Open Access Differential inhibition of human immunodeficiency virus type 1 in peripheral blood mononuclear cells and TZM-bl cells by endotoxin-mediated chemokine and gamma interferon production.(AIDS Res Hum Retroviruses, 2010-03) Geonnotti, Anthony R; Bilska, Miroslawa; Yuan, Xing; Ochsenbauer, Christina; Edmonds, Tara G; Kappes, John C; Liao, Hua-Xin; Haynes, Barton F; Montefiori, David CBacterial lipopolysaccharide (endotoxin) is a frequent contaminant of biological specimens and is also known to be a potent inducer of beta-chemokines and other soluble factors that inhibit HIV-1 infection in vitro. Though lipopolysaccharide (LPS) has been shown to stimulate the production of soluble HIV-1 inhibitors in cultures of monocyte-derived macrophages, the ability of LPS to induce similar inhibitors in other cell types is poorly characterized. Here we show that LPS exhibits potent anti-HIV activity in phytohemagglutinin-stimulated peripheral blood mononuclear cells (PBMCs) but has no detectable anti-HIV-1 activity in TZM-bl cells. The anti-HIV-1 activity of LPS in PBMCs was strongly associated with the production of beta-chemokines from CD14-positive monocytes. Culture supernatants from LPS-stimulated PBMCs exhibited potent anti-HIV-1 activity when added to TZM-bl cells but, in this case, the antiviral activity appeared to be related to IFN-gamma rather than to beta-chemokines. These observations indicate that LPS stimulates PBMCs to produce a complex array of soluble HIV-1 inhibitors, including beta-chemokines and IFN-gamma, that differentially inhibit HIV-1 depending on the target cell type. The results also highlight the need to use endotoxin-free specimens to avoid artifacts when assessing HIV-1-specific neutralizing antibodies in PBMC-based assays.Item Open Access Differential Killing of Salmonella enterica Serovar Typhi by Antibodies Targeting Vi and Lipopolysaccharide O:9 Antigen.(PLoS One, 2016) Hart, Peter J; O'Shaughnessy, Colette M; Siggins, Matthew K; Bobat, Saeeda; Kingsley, Robert A; Goulding, David A; Crump, John A; Reyburn, Hugh; Micoli, Francesca; Dougan, Gordon; Cunningham, Adam F; MacLennan, Calman ASalmonella enterica serovar Typhi expresses a capsule of Vi polysaccharide, while most Salmonella serovars, including S. Enteritidis and S. Typhimurium, do not. Both S. Typhi and S. Enteritidis express the lipopolysaccharide O:9 antigen, yet there is little evidence of cross-protection from anti-O:9 antibodies. Vaccines based on Vi polysaccharide have efficacy against typhoid fever, indicating that antibodies against Vi confer protection. Here we investigate the role of Vi capsule and antibodies against Vi and O:9 in antibody-dependent complement- and phagocyte-mediated killing of Salmonella. Using isogenic Vi-expressing and non-Vi-expressing derivatives of S. Typhi and S. Typhimurium, we show that S. Typhi is inherently more sensitive to serum and blood than S. Typhimurium. Vi expression confers increased resistance to both complement- and phagocyte-mediated modalities of antibody-dependent killing in human blood. The Vi capsule is associated with reduced C3 and C5b-9 deposition, and decreased overall antibody binding to S. Typhi. However, purified human anti-Vi antibodies in the presence of complement are able to kill Vi-expressing Salmonella, while killing by anti-O:9 antibodies is inversely related to Vi expression. Human serum depleted of antibodies to antigens other than Vi retains the ability to kill Vi-expressing bacteria. Our findings support a protective role for Vi capsule in preventing complement and phagocyte killing of Salmonella that can be overcome by specific anti-Vi antibodies, but only to a limited extent by anti-O:9 antibodies.Item Open Access Effects of Lipopolysaccharide on Human First Trimester Villous Cytotrophoblast Cell Function In Vitro.(Biology of reproduction, 2016-02) Li, Liping; Tu, Jiaoqin; Jiang, Yao; Zhou, Jie; Yabe, Shinichiro; Schust, Danny JIt has been shown that adverse obstetrical outcomes such as pre-eclampsia and intrauterine growth retardation correlate with maternal infection. In this study, we investigated mechanisms involved in infection-associated abnormalities in cytotrophoblast function. Primary human first trimester cytotrophoblast cells were isolated and treated with lipopolysaccharide (LPS). Levels of the cytokines and chemokines were measured and cytotrophoblast invasion was investigated. In addition, first trimester decidual macrophages were isolated and treated with the conditioned medium from LPS-treated cytotrophoblast cells, and macrophage migration was assessed. Coculturing decidual macrophages with cytotrophoblast cells was conducted to investigate macrophage costimulatory molecule and receptor expression and intracellular cytokine production. We found that LPS exposure increased cytotrophoblast production of pro-inflammatory cytokines tumor necrosis factor (TNF)-alpha, interleukin (IL)-1beta and IL-6, and chemokines IL-8, macrophage inflammatory protein (MIP)-1alpha, and CXCL12 in a dose-dependent manner. In addition, LPS decreased cytotrophoblast invasion, and its effect was Toll-like receptor 4 (TLR4)-dependent and partly TNF-alpha-dependent. Conditioned medium from LPS-stimulated cytotrophoblast cells increased decidual macrophage migration and this effect was partly TLR4-dependent. Furthermore, coculturing decidual macrophages with LPS-exposed cytotrophoblast cells up-regulated macrophage CD80 and CD86 expression and intracellular TNF-alpha and IL-12p40 production, while down-regulating macrophage CD206 and CD209 expression and intracellular IL-10 secretion. LPS-stimulated macrophages also inhibited cytotrophoblast invasion. In conclusion, our results indicate that LPS increases the production of a subset of proinflammatory cytokines and chemokines by human first trimester cytotrophoblast cells, decreases cytotrophoblast invasion, and alters the cross talk between cytotrophoblast cells and decidual macrophages.Item Restricted Gene expression signatures of radiation response are specific, durable and accurate in mice and humans.(PLoS One, 2008-04-02) Meadows, Sarah K; Dressman, Holly K; Muramoto, Garrett G; Himburg, Heather; Salter, Alice; Wei, ZhengZheng; Ginsburg, Geoffrey S; Chao, Nelson J; Nevins, Joseph R; Chute, John PBACKGROUND: Previous work has demonstrated the potential for peripheral blood (PB) gene expression profiling for the detection of disease or environmental exposures. METHODS AND FINDINGS: We have sought to determine the impact of several variables on the PB gene expression profile of an environmental exposure, ionizing radiation, and to determine the specificity of the PB signature of radiation versus other genotoxic stresses. Neither genotype differences nor the time of PB sampling caused any lessening of the accuracy of PB signatures to predict radiation exposure, but sex difference did influence the accuracy of the prediction of radiation exposure at the lowest level (50 cGy). A PB signature of sepsis was also generated and both the PB signature of radiation and the PB signature of sepsis were found to be 100% specific at distinguishing irradiated from septic animals. We also identified human PB signatures of radiation exposure and chemotherapy treatment which distinguished irradiated patients and chemotherapy-treated individuals within a heterogeneous population with accuracies of 90% and 81%, respectively. CONCLUSIONS: We conclude that PB gene expression profiles can be identified in mice and humans that are accurate in predicting medical conditions, are specific to each condition and remain highly accurate over time.Item Open Access Genotypic analysis of B cell colonies by in situ hybridization. Stoichiometric expression of three VH families in adult C57BL/6 and BALB/c mice.(J Exp Med, 1987-07-01) Schulze, DH; Kelsoe, GThe filter paper disc method for cloning inducible lymphocytes was used to census the splenic B cell population of C57BL/6 and BALB/c mice for the expression of three VH gene-families, VH X-24, -Q52, and -J558. B cell colonies, arising from single founder lymphocytes, were identified by in situ hybridization with VH family- and C mu-specific cDNA probes. Some 6.7 X 10(4) C mu+ colonies were screened. Among C57BL/6- or BALB/c-derived colonies, approximately 3% were VH X-24+, approximately 19% were VH Q52+, and approximately 54% were VH J558+. These frequencies are consistent with a process of equiprobable expression for individual VH segments, and provide direct evidence that normal splenic B lymphocytes use a process of random genetic combinatorics to generate the antibody repertoire.Item Open Access Heat-labile enterotoxin: beyond G(m1) binding.(Toxins (Basel), 2010-06) Mudrak, Benjamin; Kuehn, Meta JEnterotoxigenic Escherichia coli (ETEC) is a significant source of morbidity and mortality worldwide. One major virulence factor released by ETEC is the heat-labile enterotoxin LT, which is structurally and functionally similar to cholera toxin. LT consists of five B subunits carrying a single catalytically active A subunit. LTB binds the monosialoganglioside G(M1), the toxin's host receptor, but interactions with A-type blood sugars and E. coli lipopolysaccharide have also been identified within the past decade. Here, we review the regulation, assembly, and binding properties of the LT B-subunit pentamer and discuss the possible roles of its numerous molecular interactions.Item Open Access Shared monocyte subset phenotypes in HIV-1 infection and in uninfected subjects with acute coronary syndrome.(Blood, 2012-11-29) Funderburg, Nicholas T; Zidar, David A; Shive, Carey; Lioi, Anthony; Mudd, Joseph; Musselwhite, Laura W; Simon, Daniel I; Costa, Marco A; Rodriguez, Benigno; Sieg, Scott F; Lederman, Michael MThe mechanisms responsible for increased cardiovascular risk associated with HIV-1 infection are incompletely defined. Using flow cytometry, in the present study, we examined activation phenotypes of monocyte subpopulations in patients with HIV-1 infection or acute coronary syndrome to find common cellular profiles. Nonclassic (CD14(+)CD16(++)) and intermediate (CD14(++)CD16(+)) monocytes are proportionally increased and express high levels of tissue factor and CD62P in HIV-1 infection. These proportions are related to viremia, T-cell activation, and plasma levels of IL-6. In vitro exposure of whole blood samples from uninfected control donors to lipopolysaccharide increased surface tissue factor expression on all monocyte subsets, but exposure to HIV-1 resulted in activation only of nonclassic monocytes. Remarkably, the profile of monocyte activation in uncontrolled HIV-1 disease mirrors that of acute coronary syndrome in uninfected persons. Therefore, drivers of immune activation and inflammation in HIV-1 disease may alter monocyte subpopulations and activation phenotype, contributing to a pro-atherothrombotic state that may drive cardiovascular risk in HIV-1 infection.Item Open Access The transcriptomic response of rat hepatic stellate cells to endotoxin: implications for hepatic inflammation and immune regulation.(PloS one, 2013-01) Harvey, Stephen AK; Dangi, Anil; Tandon, Ashish; Gandhi, Chandrashekhar RWith their location in the perisinusoidal space of Disse, hepatic stellate cells (HSCs) communicate with all of the liver cell types both by physical association (cell body as well as cytosolic processes penetrating into sinusoids through the endothelial fenestrations) and by producing several cytokines and chemokines. Bacterial lipopolysaccharide (LPS), circulating levels of which are elevated in liver diseases and transplantation, stimulates HSCs to produce increased amounts of cytokines and chemokines. Although recent research provides strong evidence for the role of HSCs in hepatic inflammation and immune regulation, the number of HSC-elaborated inflammatory and immune regulatory molecules may be much greater then known at the present time. Here we report time-dependent changes in the gene expression profile of inflammatory and immune-regulatory molecules in LPS-stimulated rat HSCs, and their validation by biochemical analyses. LPS strongly up-regulated LPS-response elements (TLR2 and TLR7) but did not affect TLR4 and down-regulated TLR9. LPS also up-regulated genes in the MAPK, NFκB, STAT, SOCS, IRAK and interferon signaling pathways, numerous CC and CXC chemokines and IL17F. Interestingly, LPS modulated genes related to TGFβ and HSC activation in a manner that would limit their activation and fibrogenic activity. The data indicate that LPS-stimulated HSCs become a major cell type in regulating hepatic inflammatory and immunological responses by altering expression of numerous relevant genes, and thus play a prominent role in hepatic pathophysiology including liver diseases and transplantation.