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Query: UNIPROT:P43026 (lipopolysaccharide)
 62,215 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Interaction of bacterial lipopolysaccharide (LPS) with monocytes stimulates production of a variety of mediators that are involved in the pathogenesis of septic shock and wound repair. We report here the mechanisms of LPS uptake and intracellular distribution of LPS in human monocytes. Ficoll-Hypaque-purified peripheral mononuclear cells (PBMC) were exposed to LPS from rough Escherichia coli (J5) or to biotin-conjugated LPS (biotin-LPS) from smooth E. coli (0111:B4), or to fluorescein isothiocyanate-conjugated LPS of E. coli (055:B5) at 37 degrees C for various times and processed for electron microscopy, immunocytochemistry, and flow cytometry. Monocytes were identified by the presence of numerous cytoplasmic peroxidase-positive granules or by monoclonal antibodies against monocyte. LPS micelles were identified by their specific bilayer structure, staining of horseradish peroxidase reaction product, or colloidal gold using biotin-LPS or a monoclonal antibody to LPS. Binding of LPS to cell surface was observed 5 min after incubation with LPS. Intracellular localization of LPS micelles was found 30 min following exposure to LPS. Prolonged incubation with LPS increased intracellular LPS. Intracellularly, LPS micelles were found in large membrane-bound vacuoles, in small vesicles, and in the cytoplasm and nucleus. They were also observed in association with the cytomembrane of various organelles. The overall results indicate that LPS may be taken up by monocytes by direct passive diffusion through ruptures of plasma membrane, pinocytosis, and phagocytosis, involving specific and/or nonspecific binding, and suggest that peripheral blood monocytes play an important role in clearance of LPS; that LPS may have broad effects on cell functions; and that the nonspecific binding to various cytomembranes may be destructive to cell organelles and cells in general.
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PMID:Ultrastructural and immunocytochemical study of the uptake and distribution of bacterial lipopolysaccharide in human monocytes. 211 60

The pharmacologic modulation of the effect of platelet-activating factor (PAF) on the interleukin-1 (IL-1) activity present in the supernatants from lipopolysaccharide (LPS)-stimulated macrophages was investigated. Rat spleen macrophages were isolated by centrifugation on a Ficoll-Hypaque gradient followed by adherence on plastic petri dishes for 60 min at 37 degrees C under a 5% CO2/95% air atmosphere. The IL-1 content in the cell-free supernatants was assessed using the mouse thymocyte proliferation assay. Preincubation of macrophages for 10 min with 10 fM PAF prior to stimulation with 20 micrograms/ml LPS for 24 h markedly increased the IL-1 activity present in the supernatants from macrophages whereas no direct effect of PAF was noted. Although they had no direct effect, addition of L-651,392 (10 microM), a lipoxygenase inhibitor, or the oxygen-derived free radical scavenger mannitol (10 microM) during the 10-min preincubation period with PAF reversed by 105.0% and 79.9%, respectively, the action of the autacoid on IL-1 activity. Pertussis toxin (PT, 1 microgram/ml) decreased by 30% the LPS-induced IL-1 activity. Association of PT with PAF suppressed the enhancing effect of 10 fM PAF on the IL-1 activity present in the supernatants from LPS-stimulated macrophages. Thus, the enhancing effect of PAF on IL-1 release appears to be due to the production of lipoxygenase metabolites, leading to superoxide production and alterations of cAMP levels.
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PMID:Modulation of the priming effects of platelet-activating factor on the release of interleukin-1 from lipopolysaccharide-stimulated rat spleen macrophages. 213 88

The production of interleukin (IL 1) by normal human peripheral blood monocytes purified by Ficoll-Hypaque density sedimentation, Percoll-gradient sedimentation, and plastic adherence can be detected as early as 30 min intracellularly, and extracellularly within 1 hr after stimulation with lipopolysaccharide (LPS). Production of mRNA coding for the isoelectric point 7.0 species of IL 1 was also detected as early as 1 hr after LPS stimulation and reached a maximum level at 6 hr. Cell-associated IL 1 activity could be extracted with CHAPS detergent from every cell fraction (i.e., membranes, cytosol, and particulates), but was present mainly (greater than 95%) in the cytosol of LPS-activated monocytes and the myelomonocytic cell line, THP-1. The apparent m.w. of IL 1 activity on high pressure liquid chromatography gel filtration in every cell fraction was approximately 23,000 daltons, with a minor peak at 31,000 daltons, whereas the IL 1 activity in the culture supernatants was 17,000 daltons. Western blotting analysis of LPS-stimulated monocyte extracts showed two forms of IL 1 corresponding to 31,000 daltons and 25,000 daltons. Exposure of viable cells to trypsin and plasmin released biologically active 23,000 dalton IL 1 only from IL 1-producing cells such as activated monocytes and IL 1-producing Ebstein-Barr virus B lymphocyte cell lines. Consequently, biologically active IL 1 is presumably exposed on the outer surface of cell membranes. Furthermore, IL 1 release by human monocytes in plasminogen-depleted fetal calf serum was considerably decreased. Conversely, supplementation of plasminogen-depleted serum with purified plasminogen restored the IL 1 production, suggesting that plasmin or plasmin-like factors may be involved in the regulation of the release of IL 1 from IL 1-producing cells. In conclusion, the results suggest that IL 1 is rapidly produced, is pooled in the cytosol, and in part is processed by enzymes, is transferred to the plasma membranes, and is then released from the cells. Tissue plasminogen activator and serum enzymes such as plasmin may therefore be involved in the release of IL 1 from IL 1-producing cells.
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PMID:Intracellular localization of human monocyte associated interleukin 1 (IL 1) activity and release of biologically active IL 1 from monocytes by trypsin and plasmin. 242 Aug 74

For much of the last decade, an increasing number of surgeons have been interested in objective assessment of cellular contributors to host defense function. In order to study many of these processes, it is apparently desirable that the cells be isolated to the extent feasible for the purpose of analyzing a more or less pure population of cellular elements. The purpose of this paper is to describe the physiologic activation of mononuclear cells that occurs as a result of the isolation process. Therefore, it follows logically that such cells are therein intrinsically less responsive to further physiologic manipulation in vitro. Analyses of such data without an awareness of this intrinsic aberration will undoubtedly lead to misinterpretation of the capacity of such cells for further modulation by immunostimulants or by the intrinsic processes related to injury, anesthesia, and operation. Furthermore, it may indicate that certain agents, e.g., cytokines, are unable to stimulate cellular function when, in fact, the defense function of the cell has been initially stimulated by the isolation procedure. Fractionation of human peripheral blood over Hypaque-Ficoll and subsequent purification of monocytes by adherence to plastic lead to an increase in the relative density of HLA-DR on monocytes. This increase occurred when carried out in endotoxin lipopolysaccharide (LPS)-contaminated or LPS-depleted reagents. LPS, added experimentally to whole blood, enhanced HLA-DR expression on monocytes without further manipulation. Monocyte HLA-DR expression measured in whole blood was reduced in patients with major sepsis (n = 19) compared to normal subjects (n = 10).(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Experimental and clinical significance of endotoxin-dependent HLA-DR expression on monocytes. 273 99

Human neutrophils were isolated from peripheral blood by four methods: 1) Ficoll-Hypaque gradients and erythrocyte lysis, 2) plasma-Percoll gradients, 3) a "lipopolysaccharide (LPS)-free" method yielding 85% neutrophils, and 4) by centrifugation of cells prepared by Method 3 through a plasma-Percoll gradient to produce pure neutrophils. The use of the Ficoll-Hypaque method resulted in spontaneous change of cell shape, enhanced formyl-methionyl-leucyl-phenylalanine (FMLP)-stimulated release of superoxide anion, increased release of lysosomal enzymes upon subsequent FMLP stimulation, and reduced chemotactic responsiveness, by comparison with the other methods. These effects were not due to erythrocyte lysis by NH4C1 but were reproduced by exposure of neutrophils prepared by the "LPS-free" method or the use of plasma-Percoll gradients to 10-100 ng/ml LPS. Neutrophil change of shape and stimulated O-2 production were particularly sensitive markers of these effects. The effects of trace concentrations of LPS in the modulation of neutrophil function may have relevance to the pathophysiology of endotoxemia and its resultant tissue injury.
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PMID:Modulation of multiple neutrophil functions by preparative methods or trace concentrations of bacterial lipopolysaccharide. 298 39

Bacterial endotoxin (lipopolysaccharide, LPS) and interleukin-2 (IL-2) are known to stimulate NK cell mediated cytotoxicity against tumor cells. In the present report we sought to correlate the stimulatory effect of LPS and IL-2 on NK cell activity with ultrastructural changes which occurred as a result of such stimulation. Peripheral blood mononuclear cells (PBMC) were purified from healthy donors by a Ficoll-Hypaque density gradient technique. Leu-11a+ NK cells were isolated by flow microfluorometry using a monoclonal FITC conjugated anti-Leu-11a antibody and a FACS II cell sorter. The PBMC were incubated, respectively, with E. coli LPS or recombinant IL-2 (IL-2) for various time periods. Sorted Leu-11a+ NK cells were incubated with LPS for 24 hours. The NK cytotoxicity in the PBMC and sorted Leu-11a+ cells was assessed by a 51Cr release technique using K562 tumor cells as targets. Leu-11a+ NK cells were identified by immunoelectron microscopy using anti-Leu-11a antibody and labeling with horseradish peroxidase or colloidal gold. Results showed that both LPS and IL-2 significantly enhanced the cytotoxic activity of PBMC. The cytotoxicity of sorted Leu-11a+ cells was augmented by LPS. Recombinant IL-2 induced a significant increase in the number of dense granules, hypertrophy of Golgi apparatus and rough endoplasmic reticulum, and mitosis of Leu-7+ cells and Leu-11a+ cells 4 or 7 days after stimulation. These data indicate that: (1) the effect of LPS on the enhancement of NK cytotoxicity in PBMC may be a direct and/or indirect process involving production of lymphokines; (2) LPS has a direct effect on sorted Leu-11a+ cells; (3) IL-2 stimulates mitosis of Leu-7+ cells and Leu-11a+ cells; and (4) the LPS or IL-2 induced ultrastructural changes in Leu-11a+ cells are consistent with the enhanced NK cytotoxicity.
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PMID:Ultrastructural and functional effects of lipopolysaccharide and interleukin-2 on human NK cells. 305 78

Interleukin 1 (IL 1) production by normal human B lymphocytes was investigated. Normal human peripheral blood B lymphocytes were purified by sequential separation with the use of Ficoll-Hypaque gradient centrifugation, sheep red blood cell rosette formation, Percoll gradients, and treatment with monoclonal antibodies (anti-Leu-M1, B73.1, and T101) and complement. Both purified large B lymphocytes (BL) and small B lymphocytes (BS) produced IL 1-like (thymocyte co-mitogenic and fibroblast mitogenic) activities in response to lipopolysaccharide. Maximal production of IL 1 activity by both BL and BS occurred at 48 hr. The m.w. of IL 1 activities from both BL and BS were about 20,000 with high pressure liquid chromatography, and the major isoelectric point of BL- and BS-derived IL 1 activity was 7.0. A rabbit anti-human monocyte IL 1 antiserum inhibited the activity of B cell-derived IL 1, suggesting antigenic similarities of monocyte- and B lymphocyte-derived IL 1 moieties. These data suggest that normal B lymphocyte-derived IL 1 activity is biochemically and immunologically similar to monocyte-derived IL 1.
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PMID:Production of interleukin 1 activity by normal human peripheral blood B lymphocytes. 387 31

Responses of B cells with or without receptors for C3 (CR) to polyclonal B cell activators (PBA) were studied. Mouse spleen cells were incubated with sheep red blood cells (SRBC) coated with antibody and complement to form rosettes, and they were separated by Ficoll-Hypaque density sedimentation into populations depleted of and enriched with lymphocytes bearing CR (CRL). These 2 populations were cultured with lipopolysaccharide (LPS), purified protein derivative of tuberculin (PPD), or dextran sulfate (DxS) and assayed for anti-TNP PFC. The CRL-depleted population responded well to LPS, poorly to PPD, and it showed practically no response to DxS, whereas the CRL-enriched population seemed to respond poorly to LPS but well to both PPD and DxS. The low responsiveness of the cRL-depleted population to PPD and DxS could not be explained by a shift of time-kinetics, by the dose-response profile of the responding cells, or by the depletion of adherent cells. Suppressor T cells did not take part in the reduced responses, since the treatment of the population with anti-Thy 1.2 plus complement could not restore the responses. These results indicate that B cells with CR [CR(+) B cells] respond well both to PPD and DxS, whereas the cells without CR [CR(-) B cells] respond poorly to PPD and DxS. It was difficult to evaluate the low responsiveness of CR(+) B cells to LPS because of the high background PFC of the cRL-enriched population.
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PMID:Responses of B cells with or without C3 receptors to polyclonal B cell activators. 616 36

A precise method for quantitation of polymorphonuclear leukocyte (PMNL) accumulation in skin in vivo, has been developed so that the proinflammatory effects of various agents can be compared. This method can also be used to evaluate the effect of therapeutic agents on PMNL accumulation in vivo. Rabbit PMNLs were purified from heparinized blood by dextran sedimentation, hypotonic lysis, and separation on Ficoll-Hypaque. The PMNLs were labeled with 3-5 microCi per 10(6) cells of 111In oxine and reinfused coincidentally with different concentrations of different chemotactic and proinflammatory materials injected intradermally into the back. In some experiments, varying concentrations of acetic acid were applied topically. Four to 18 hours later, the rabbits were sacrificed. Eight-millimeter punch biopsies were obtained from the injection sites and counted in a gamma counter. The number of PMNLs infiltrating the dermis was also quantitated in histologic sections. A significant correlation was found between the percent increase in radioactivity and the percent increase in PMNL accumulation morphologically. Dose-response curves were generated using such proinflammatory materials as formyl-methionyl-leucyl-phenylalanine, lipopolysaccharide, activated serum, trypsin, glycogen, and acetic acid. These curves were highly reproducible from animal to animal. Using this assay, we found that as little as 1 microgram of trypsin induced detectable PMNL accumulation. This is 2-3 logs more sensitive than injecting mice intraperitoneally with trypsin. Diisopropyl fluorophosphate-inactivation of trypsin inhibited PMNL accumulation. This sensitive and quantitative bioassay of PMNL accumulation permits evaluation of multiple agents in the same animal, which decreases animal to animal variation.
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PMID:Labeling of peripheral blood polymorphonuclear leukocytes with indium-111: a new method for the quantitation of in-vivo accumulation of PMNLs in rabbit skin. 642 Apr 76

Phagocytosis and cellular cytotoxicity by mononuclear phagocytes of blood and intestinal mucosa were studied in patients with Crohn's disease and large bowel neoplasms. Antibody coated sheep erythrocytes were used for phagocytic assays and cellular cytotoxicity in vitro was measured by 24 hour isotope release from 75Selenium methionine-labelled RPMI 4788 human cancer cell cultures in the presence of mononuclear phagocyte-enriched effector populations. The mean percent of mononuclear phagocytes in Ficoll-Hypaque purified mononuclear cell suspensions of blood of healthy controls was 25.9 compared with 44.6 in patients with Crohn's disease, 45.6 in patients with colon neoplasms and 11.6 in intestinal mucosa. Phagocytic indices were similar in all groups, but the phagocytic capacity of mucosal macrophages was twice that of blood monocytes. Mean cytotoxicity of monocytes of patients with Crohn's disease was 12.8% compared with 22.9% for monocytes from normal controls, and 29.4% for patients with colon tumours. Mean cytotoxicity by mucosal macrophages was 18.0% compared with 13.2% by mucosal lymphocyte populations. Exposure of monocytes of Crohn's disease patients to bacterial lipopolysaccharide modestly increased cytotoxicity, but exposure did not alter phagocytosis by monocytes of patients or controls. The results indicate that monocytes of patients with Crohn's disease exhibit subnormal in vitro cytotoxicity. Mucosal macrophages from patients with various diseases show enhanced phagocytosis compared with blood monocytes, and they can mediate cellular cytotoxicity in vitro.
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PMID:Comparative studies of mononuclear phagocyte function in patients with Crohn's disease and colon neoplasms. 662 13


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