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Biomedical subjects

L A Boxer

Publications and source records attributed to L A Boxer.

At least 127 records · Page 7Linked to original sources
PubMed1983

Modulation of polymorphonuclear leukocyte function by cetiedil.

Cetiedil citrate monohydrate inhibits sickling of red cells and aggregation of platelets. We assessed its ability to attenuate polymorphonuclear leukocyte (PMN) function. PMN aggregation in response to 2 X 10(-7) M formyl-met-leu-phe (FMLP) was inhibited in a dose-dependent fashion by cetiedil concentrations ranging from 60 to 250 microM. Additionally, 125 microM cetiedil inhibited PMN aggregation in response to 2 X 10(-7) M FMLP, 20 ng/ml phorbol myristate acetate (PMA), and 1 X 10(-6) M A23187 by 69% +/- 18%, 72% +/- 20%, and 65% +/- 4%, respectively. Inhibition of FMLP-induced aggregation was provided by only 5 min of incubation of the drug with the cells and was partially reversible. Cell viability was unaffected by exposure of PMN to the drug. Correspondingly, 125 microM cetiedil prevented the translocation of calcium from the PMN membrane as assessed by chlorotetracycline fluorescence. Paralleling the effect of the drug on PMN aggregation, 125 microM cetiedil inhibited release of superoxide by 55% and decreased the number of available 3H-FMLP receptors. However, its effect on release of the primary granule constituent, myeloperoxidase, was minimal (4.5% inhibition), while the effect on release of the specific granule product, lactoferrin (27% inhibition), was modest. These studies indicate that cetiedil affects PMN aggregation and superoxide release to a much greater extent than PMN degranulation. Thus, cetiedil may have potential uses in modulating inflammatory response in vivo.

Azepines↗
PubMed1983

Comparative studies of functional characteristics of mononuclear cell subsets and granulocytes.

Two human peripheral blood monocyte subsets and lymphocytes were isolated by counterflow centrifugal elutriation (CCE). The cell volumes of 303 mu3 and 380 mu3 were measured for the smaller and larger monocyte populations, respectively. Superoxide release by large monocytes exposed to opsonized zymosan was five times more active than that of the small monocytes. The production of colony stimulating activity was two-fold greater and the myeloperoxidase activity was 1.4-fold greater by the larger monocytes. Enriched fractions of cytotoxic cells responsible for natural killer (NK) activity against neuroblastoma cells were also obtained by counterflow centrifugal elutriation. Natural killer cells were obtained in larger lymphocyte fractions and had a mean cell volume of 180 mu3. Compared with the NK activity against the neuroblastoma cells, both the small and large monocytes displayed greater antibody-dependent cellular cytotoxicity (ADCC) activity against human erythrocytes. The larger peripheral blood monocytes aggregated in response to FMLP (N-formyl-methionyl-leucyl-phenyl alanine peptide) and PAF (platelet activating factor). Unlike the granulocyte, monocyte aggregation in response to FMLP was not accompanied by degranulation nor was it potentiated by cytochalasin B. In addition, monocyte aggregation could be blocked by benoxaprofen, unlike the granulocyte. Thus, CCE provides a means of isolating subsets of monocytes and lymphocytes and obtaining large numbers of peripheral blood monocytes for functional studies.

Antibody-Dependent Cell Cytotoxicity↗
PubMed1983

The mechanism of action of the antiinflammatory agents dexamethasone and Auranofin in human polymorphonuclear leukocytes.

Human polymorphonuclear neutrophils (PMN) were treated with the antiinflammatory agents dexamethasone or Auranofin. PMN treated with dexamethasone in a dose range of 0.25-1 microM or Auranofin, 5-15 mM, were stimulated with 10(-7)M N-formyl-methionyl-leucyl-phenylalanine (FMLP). These agents were shown to inhibit the functional responses of degranulation and superoxide production in a dose-dependent manner. Similarly, the change in electrophoretic mobility, reflecting cell surface charge, was blocked. While both agents inhibited change in the fluorescence of the calcium chelate probe chlorotetracycline (CTC), the pattern of inhibition was significantly different. Dexamethasone appeared to inhibit the CTC response during its latter phases, while Auranofin inhibited all aspects of the CTC response. Auranofin was additionally shown to significantly decrease specific binding of FMLP, as well as the number of FMLP receptors. The two agents thus appear to act by different mechanisms. Dexamethasone is shown to have an effect on membrane-bound calcium release as measured by CTC, while Auranofin interferes with receptor binding.

Auranofin↗
PubMed1983

Disorders of stimulus-response coupling in neutrophils.

The initiation of both the normal and pathologic responses of human neutrophils to surface stimulation and the ensuing biochemical and physiologic events are elucidated. This knowledge has contributed to an understanding of the controlling mechanisms that may account for impaired phagocytic function in several clinical disorders associated with recurrent bacterial infections.

Animals↗
PubMed1983

Vitamin E--a selective inhibitor of the NADPH oxidoreductase enzyme system in human granulocytes.

The cellular sites of H2O2 formation in phagocytizing granulocytes have been identified with cerium chloride. A precipitate was visible in phagosomes and on plasma membranes from intact normal cells in the presence of either 0.71 mM NADH or NADPH. X-ray microanalysis permitted identification of cerium deposition within the phagosomes even in the absence of reduced pyridine nucleotides. Catalase ablated the formation of the reaction product. Intact granulocytes obtained from subjects receiving 1600 units of vitamin E daily for 2 weeks exhibited reaction product in the presence of NADH but not NADPH. Intact cells from subjects treated with vitamin E demonstrated diminished numbers of phagocytic vesicles containing reaction product. During phagocytosis the granulocytes treated with vitamin E consumed oxygen but exhibited significantly reduced rates of hydrogen-peroxide-dependent glucose-1-14C oxidation to 14CO2. Isolated phagocytic vesicles obtained from granulocytes after ingestion of opsonized lipopolysaccharide-paraffin oil droplets contained reaction product when exposed to 0.71 mM NADPH. No reaction product was evident at 0.71 mM NADH but was evident at 2.0 mM NADH. Isolated phagocytic vesicles from the granulocytes of subjects receiving vitamin E exhibited reaction product only in the presence of NADH. These observations suggest that vitamin E interferes with the electron transport chain apparently required for the oxidation of NADPH to form H2O2 in the phagocytizing granulocyte.

Cytochrome Reductases↗
PubMed1983

Synergistic effect of heparin and chemotactic factor on polymorphonuclear leukocyte aggregation and degranulation.

The in vitro effects of therapeutic amounts of polyanionic heparin on human polymorphonuclear leukocytes (PMN) aggregation and on the release of cationic lactoferrin from PMN-specific granules were investigated. Incubation of 1 X 10(7) human PMNs with 0.3 unit/ml of heparin followed by stimulation with the chemotactic peptide N-formyl-methionyl-leucyl-phenylalanine (FMLP) 2 X 10(-7) M significantly increased PMN aggregation, compared with controls. Cytochalasin B potentiated aggregation, which was further increased by incubation of the PMNs with heparin. Similarly, heparin also increased PMN degranulation and lactoferrin release following stimulation with FMLP with or without cytochalasin B, compared with controls. In addition, human lactoferrin complexed with heparin on a sucrose density gradient and caused a significant shift in the migration of 3H-heparin. Finally, rabbits pretreated with intravenous heparin resulting in prolongation of their activated partial thromboplastin time (APTT) to 1.5 to 2.5 times baseline had more profound reduction in PMN counts following a challenge with the secretagogue phorbol myristate acetate (PMA). These studies demonstrate that heparin can interact synergistically with chemotactic stimuli known to evoke lactoferrin release, which in turn leads to enhancement of PMN aggregation. Our data further suggest that heparin may be contraindicated in the treatment of syndromes with increased PMN aggregation such as endotoxin-induced Schwartzman-type reactions.

Animals↗
PubMed1983

Plasma lactoferrin reflects granulocyte activation in vivo.

N-formyl-met-leu-phe (FMLP) causes polymorphonuclear leukocytes (PMN) to secrete and become "sticky" in vitro. We related these events to in vivo FMLP-induced neutropenia. FMLP was intravenously administered to anesthetized rabbits in doses ranging from 0.01 microgram to 1.0 microgram. Controls received phosphate-buffered saline (PBS), the diluent for FMLP. Blood pressure, respiratory rate, and arterial Po2 were monitored. High and intermediate doses of FMLP caused a dramatic but transient decrease in blood pressure and an increase in respiratory rate. Prior to FMLP infusion, plasma lactoferrin level was 6.4 +/- 4.1 micrograms/ml, and the absolute granulocyte account (AGC) was 2008 +/- 1229 (mean +/- SD). There was a positive linear correlation between AGC and plasma lactoferrin level prior to injection of FMLP (R2 = 0.74, p less than 0.01). At 1 min after FMLP injection, the percent change in AGC decreased as an exponential function of dose to as low as 10% of baseline (R2 = 0.86, p = 0.002) and plasma concentration of lactoferrin increased as an exponential function of dose to as high as 30 micrograms/ml (R2 = 0.84, p = 0.006). Thus, FMLP-induced neutropenia is associated with increased levels of plasma lactoferrin, suggesting that PMN are induced to degranulate in vivo.

Animals↗
PubMed1983

The Chediak-Higashi gene in humans. III. Studies on the mechanisms of NK impairment.

Lymphocytes from six Chediak-Higashi (CH) patients were markedly depressed in their ability to lyse tumour cell targets in both 51Cr release and single cell cytotoxicity assays. The frequency of lymphocytes bearing the OKM1 marker and the frequency of T3+, T4+, T8+, Ia+, Mo1+, Mo2+ and B1+ cells was normal among sheep erythrocyte rosetting (E+) and non-rosetting (E-) peripheral blood leucocytes analysed by flow cytofluorography. Cells expressing the NK shared markers, OKM1, mac-1, FcR, and the characteristic large granular lymphocyte (LGL) morphology of NK cells were also present in normal numbers in the highly enriched NK fraction separated on Percoll density gradients. This fraction did not contain detectable numbers of cells expressing the Mo2 marker of human monocytes. Therefore most of the cells stained by monoclonal OKM1 and mac-1 in this fraction are likely NK cells, rather than monocytes, and we conclude that the size of the NK pool in CH patients is probably normal. The capacity of CH lymphocytes to recognize and bind to tumour cells was also normal as was the subsequent burst of oxygen intermediates produced by the NK cells in a chemiluminenscence assay. We have shown elsewhere that O2- generation is directly involved in activating subsequent steps in the NK cytolytic pathway. These results suggest that NK cells in CH patients are present in normal frequency but are blocked at some post-recognition, post-activation step in the cytolytic pathway subsequent to the burst of oxygen intermediates but preceding the lethal hit.

Adolescent↗
PubMed1982

The influence of vitamin E on human polymorphonuclear cell metabolism and function.

These studies on the effect of administration of 1,600 units of vitamin E to humans indicated the following responses to the PMNs (TABLE 6). Functional alterations occur with an increased ability to ingest particles but a mild decrease in bactericidal potency of the PMN. Although the respiratory burst is slightly enhanced as is superoxide anion release, H2O2 release from the PMN is markedly impaired. The hexose monophosphate shunt activity, which is dependent on intracellular H2O2 is decreased during phagocytosis. Membrane responses such as changes in order parameter during phagocytosis as reported by the stearic acid analogue probe 5DS are similar to those of normal PMNs. The release of arachidonic acid from membranes of vitamin E PMNs during phagocytosis of opsonized zymosan is slightly enhanced, indicating normal phospholipase A2 activation. NADH oxidase-derived H2O2 is not impaired within phagocytic generated by NADPH oxidase in phagocytic vesicles, accounting for impairment in HMPS activity and bactericidal activity in these cells.

Arachidonic Acid↗
PubMed1982

Purified human plasma kallikrein aggregates human blood neutrophils.

Exposure of human blood polymorphonuclear leukocytes (PMN) to purified active plasma kallikrein resulted in PMN aggregation when kallikrein was present at concentrations ranging from 0.4 to 0.6 U/ml (0.18-0.27 microM). Kallikrein-induced PMN aggregation was not mediated through C5-derived peptides, because identical responses were observed whether or not kallikrein had been preincubated with an antibody to C5. Moreover, kallikrein was specific for aggregating PMN, because no aggregation was observed with Factor XII active fragments (23 nM), Factor XIa (0.6 U/ml or 15nM), thrombin (1.6 microM), plasmin (2 microM), porcine pancreatic elastase (2 microM), bovine pancreatic chymotrypsin (2 microM), or bradykinin (1 microM). Bovine pancreatic trypsin (2 microM) aggregated PMN, but to a lesser extent than kallikrein (0.18 microM). Kallikrein was a potent aggregant agent for PMN because similar responses were observed with kallikrein (0.5 U/ml or 0.23 microM) and an optimal dose (0.2 microM) of N-formyl-methionyl-leucyl-phenylalanine. In addition, PMN incubation with kallikrein resulted in stimulation of their oxidative metabolism as assessed by an increased oxygen uptake. Neutropenia and leukostasis observed in diseases associated with activation of the contact phase system may be the result of PMN aggregation by plasma kallikrein.

Cell Aggregation↗
PubMed1982

Membrane-bound lactoferrin alters the surface properties of polymorphonuclear leukocytes.

Polymorphonuclear leukocytes (PMN) aggregate and avidly attach to endothelium in response to chemotactic agents. This response may be related in part to the release of the specific granule constituent lactoferrin (LF). We found by using immunohistology and biochemical and biophysical techniques that LF binds to the membrane and alters the surface properties of the PMN. Upon exposure of PMN treated with 5 micrograms/ml cytochalasin B to 2 x 10(-7) M formyl-methionine-leucine-phenylalanine for 5 min, the PMN mobilized LF to their surface as observed by immunoperoxidase staining for LF. At added LF levels ranging from 4 to 15 micrograms/10(7) PMN there was a dose-dependent reduction in PMN surface charge reaching 4 mV, when the partitioning into the membrane of a charged amphipathic nitroxide spin label was measured by electron spin resonance spectroscopy, whereas transferrin was without effect. When 125I-FeLF was added to human PMN in increasing amounts and the results corrected for the residual amount of free LF contaminating the cells, the PMN were saturated with LF at concentrations between 100 and 200 nM in the medium. Human PMN bound 1.35 x 10(6) molecules per cell and the calculated value for the association constant for these receptors was 5.2 x 10(6) M-1. Additionally, 6 micrograms/ml LF served as an opsonin for rabbit MN to promote PMN uptake by rabbit macrophages, when assessed by electron microscopy, but lysozyme did not. These studies indicate that LF can bind to the surface of the PMN and reduce its surface charge. This correlates with enhanced "stickiness" leading to a variety of cell-cell interactions.

Animals↗
PubMed1982

Further studies of natural killer cell function in Chediak-Higashi patients.

Spontaneous natural killer (NK) activity and antibody-dependent cellular cytotoxicity (ADCC) of blood lymphocytes against five human tumour cell lines (K562, Molt-4, HL-60, Chang, Daudi) and three mouse tumour lines (YAC, P815, RBL-5) were ten- to 100-fold lower than normal in six patients with Chediak-Higashi (CH) disease. NK and ADCC were defective at 4 hr, and less so at 18 hr. The NK activity in normals and CH patients was mediated in part by FcR+, E- effector cells. ADCC against human erythrocytes was normal in CH patients, as were lectin-dependent cytolysis and mixed lymphocyte proliferative responses. Phagocytosis of antibody-coated ox erythrocytes was normal in CH patients as well. These observations confirm that the CH syndrome is associated with a profound and selective defect in NK and ADCC activity against tumour cells, whereas other mononuclear cell-mediated functions are normal.

Adult↗
PubMed1982

Monocyte aggregation and superoxide anion release in response to formyl-methionyl-leucyl-phenylalanine (FMLP) and platelet-activating factor (PAF).

Like the PMN, human peripheral blood monocytes were capable of aggregating in response to FMLP and PAF. Monocyte aggregation was dependent on glycolysis and the presence of divalent cations. Unlike the PMN, monocyte aggregation in response to FMLP was not accompanied by degranulation, nor was it potentiated by cytochalasin B. Furthermore, cytochalasin B and its cogenitor, dihydrocytochalasin B, inhibited aggregation in response to PAF. PAF and FMLP appeared to react with the monocyte at separate receptors because sequential challenge of the monocyte with the same agents failed to elicit further aggregation, whereas rechallenge with the alternative agent induced further aggregation. Because the monocyte will aggregate to chemotactic agents in vitro, it is likely that the cell will be affected by these agents in vivo, which could lead to leukoembolization of circulating monocytes.

Calcium↗
PubMed1982

Relation of respiratory burst and arachidonate metabolism during phagocytosis by guinea pig alveolar macrophages.

We have examined the respiratory burst and arachidonic acid oxygenation that accompany phagocytosis in macrophages. Guinea pig alveolar macrophages were stimulated with opsonized zymosan in the presence of inhibitors of arachidonic acid metabolism: ASA, indomethacin, and ETYA, ASA, at concentrations as high as 60 micrograms/ml, had no effect on either oxygen consumption or superoxide ion formation. Indomethacin (4 x 10(-4) M) and ETYA (2 x 10(-5) M) did inhibit oxygen utilization and superoxide production. However, no indomethacin or ETYA inhibition of oxygen utilization was detected in the presence of 1 mM KCN, suggesting that the inhibitable portion of the respiratory burst observed with indomethacin or ETYA was dependent on mitochondrial respiration. Further study with ETYA showed that the inhibitor at 2 x 10(-5) M had little effect on uptake of 125I-labeled zymosan but did abolish the conversion of 14C-arachidonic acid to a compound that co-migrated with authentic 12-HETE on silica gel plates. Lower concentrations of ETYA (5 x 10(-6) M), which had no effect on the respiratory burst of phagocytosing alveolar macrophages, also inhibited arachidonic acid metabolism. We conclude therefore that the inhibition of oxygen consumption and superoxide production by ETYA at 2 x 10(-5) M is unrelated to inhibition of arachidonic acid metabolism. Furthermore, the oxygenation of arachidonic acid requires little of the oxygen consumed by phagocytosing alveolar macrophages.

5,8,11,14-Eicosatetraynoic Acid↗
PubMed1982

Metabolic, membrane, and functional responses of human polymorphonuclear leukocytes to platelet-activating factor.

The phospholipid mediator of anaphylaxis, platelet-activating factor (PAF) is chemotactic for polymorphonuclear leukocytes (PMN). We have examined this agent's effects on several other PMN functions. Human PMN were prepared from heparinized venous blood by Ficoll gradient. Metabolic burst was examined by measurement of O2 use and O2.- production in the presence or absence of PAF (10(-6)--10(-9) M). Unless cells were treated with cytochalasin-B (5 micrograms/ml), no significant respiratory burst was demonstrated. However, pretreatment with PAF (10(-7) M) enhanced approximately threefold the O2 utilization found when cells were subsequently stimulated with 10(-7) M FMLP. PAF also stimulated arachidonic acid metabolism in 14C-arachidonic acid-labeled PMN. Thin-layer chromatography analysis of chloroform-methanol extracts showed substances that comigrated with authentic 5-hydroxyeicosatetraenoic acid had a marked increase in radioactivity following PAF stimulation at 10(-7) M. PAF failed to stimulate release of granule enzymes, B-glucuronidase, lysozyme, or myeloperoxidase unless cytochalasin-B were added. PAF from 10(-6) M to 10(-10) M affected PMN surface responses. PMN labeled with the fluorescent dye, chlorotetracycline, showed decreased fluorescence upon addition of PAF, suggesting translocation of membrane-bound cations. Further, the rate of migration of PMN in an electric field was decreased following PAF exposure, a change consistent with reduced cell surface charge. PMN self-aggregation and adherence to endothelial cells were both influenced by PAF (10(-6) M--10(-9) M). Aggregation was markedly stimulated by the compound, and the percent PMN adhering to endothelial cell monolayers increased almost twofold in the presence of 10(-8) M PAF. Thus, PAF promotes a variety of PMN responses: enhances respiratory burst, stimulates arachidonic acid turnover, alters cell membrane cation content and surface charge, and promotes PMN self-aggregation as well as adherence to endothelial cells.

Cell Adhesion↗