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Enzymatic Activities of Bovine Peripheral Blood Leukocytes and Milk Polymorphonuclear Neutrophils during Intramammary Inflammation Caused by Lipopolysaccharide  

Microsoft Academic Search

Leukocytes are recruited from peripheral blood into milk as part of the inflammatory response to mastitis. However, excessive accumulation of inflammatory cells alters the quality of milk and the proteases produced by polymorphonuclear neutrophils (PMNs) and macrophages may lead to mammary tissue damage. To investigate PMN recruitment and the kinetics of their intracytoplasmic enzymes in inflammation, we generated mastitis in

C. Prin-Mathieu; Y. Le Roux; G. C. Faure; F. Laurent; M. C. Bene; F. Moussaoui



Influence of 17?-Estradiol, Progesterone, and Dexamethasone on Diapedesis and Viability of Bovine Blood Polymorphonuclear Leukocytes  

Microsoft Academic Search

The aim of the current study was to investigate whether polymorphonuclear leukocyte (PMN) diapede- sis and viability are influenced by steroid hormones. Using an in vitro model with different types of cell layers (bovine mammary epithelial cells and fibro- blasts), we investigate whether steroid hormone treat- ments (17?-estradiol, progesterone, and dexametha- sone) have an influence on the diapedesis capacity and

I. Lamote; E. Meyer; L. Duchateau; C. Burvenich



Superoxide production by polymorphonuclear leukocytes  

Microsoft Academic Search

Phagocytosis by polymorphonuclear leukocytes triggers a burst of oxidative metabolism resulting in hydrogen peroxide and superoxide production, and these active oxygen species function in the killing of microorganisms. A new cytochemical technique, based on a manganese dependent diaminobenzidine oxidation, has been developed to detect superoxide in these cells. It has been shown that superoxide generation is associated with the plasma

R. T. Briggs; J. M. Robinson; M. L. Karnovsky; M. J. Karnovsky



Inhibitory effect of stobadine on FMLP-induced chemiluminescence in human whole blood and isolated polymorphonuclear leukocytes.  


The chemiluminescence (CL) technique with luminol and isoluminol was used to characterize the effect of stobadine on reactive oxygen metabolites (ROM) generation in human whole blood and in isolated polymorphonuclear leukocytes (PMNL) stimulated with N-formyl-methionyl-leucyl-phenyl-alanine (FMLP). In whole blood and in isolated PMNL, stobadine in the concentrations of 1, 10 and 100 micromol/L significantly inhibited the CL signal after FMLP, which activated predominantly extracellular generation of ROM. The same concentrations of stobadine were effective on CL in a cell-free system. On the other hand, myeloperoxidase (MPO) liberation was decreased by stobadine only in the concentration of 100 micromol/L. The results showed stobadine to act as a potent inhibitor/scavenger of extracellularly produced ROM in human PMNL and indicated interference of stobadine with ROM as well as with signalling events resulting in NADPH-oxidase activation and MPO liberation. PMID:16871535

Drábiková, Katarína; Jancinová, Viera; Nosál, Radomír; Pecivová, Jana; Macicková, Tatiana; Turcáni, Peter


Abnormal directed migration of blood polymorphonuclear leukocytes in rheumatoid arthritis. Potential role in increased susceptibility to bacterial infections.  

PubMed Central

Rheumatoid arthritis (RA) patients are at higher risks of bacterial infection than healthy subjects. Polymorphonuclear leukocytes (PMN) are the first line of nonspecific cellular defence against these infections. We tested the hypothesis that abnormal directed migration of PMN may be one reason for the increased infection rate of RA patients. PMN migration was investigated in 68 peripheral blood samples of 15 RA patients compared with 64 samples of healthy controls in a novel whole blood in vitro membrane filter assay. The migration of PMNs from RA patients and controls was stimulated using the bacterial chemoattractant N-formyl-methionyl-leucyl-phenylalanine (fMLP). Unstimulated PMN migration of RA patients was increased compared with healthy controls as measured by the following parameters: (a) absolute number of migrant PMNs (1954+/-87 vs. 1238 +/-58 PMN/mm2), (b) percentage of PMNs migrated into the filter (total migration index, TMI) (28.6+/-0.9 vs. 24.0+/-0.8%), (c) the distance half the migrating PMNs had covered (distribution characteristic, DC) (22.6+/-1.1 vs. 16.1+/-0.6 mm) and (d) the product of TMI and DC (neutrophil migratory activity, NMA) (669.0+/-45.0 vs. 389.0+/-18.9). fMLP stimulated PMNs of RA patients showed defective migration compared to unstimulated samples as shown by (a) a reduced number of migrant PMNs (1799+/-93 PMN/mm2), (b) lower TMI (26.1+/-0.9%), (c) unremarkable altered distribution characteristic (22.9+/-0.8 mm) and (d) significant reduced migratory activity (600.0+/-30.0). Our data suggest that the high incidence of infections in RA patients may partly be caused by defective migratory activity of PMNs to bacterial chemoattractants as demonstrated by fMLP.

Aglas, F; Hermann, J; Egger, G



Concanavalin A capping in polymorphonuclear leukocytes  

Microsoft Academic Search

Various polymorphonuclear leukocyte (PMN) functions are dependent on an intact intracellular cytoskeleton consisting of the microtubules and the microfilaments. To investigate the microtubule system in PMNs we observed the spontaneous, Colchicine and Diamide induced cap-formation by fluorescence microscopy in PMNs obtained from children with bacterial and viral infections, bronchial asthma, as well as during immunosuppressive therapy. Children with bacterial infections

M. Rister; F. Wegener; E. Gladtke



Simultaneous flow cytometric measurement of phagocytotic and oxidative burst activity of polymorphonuclear leukocytes in whole bovine blood  

Microsoft Academic Search

Polymorphonuclear neutrophils (PMN) are important in host defence against bacterial infection in the bovine mammary gland and techniques are needed to evaluate their functional activities. A rapid and sensitive two-color flow cytometric method for simultaneous measurement of phagocytosis arte and oxidative burst activity of bovine PMN in small blood samples is described. The method utilizes the oxidation of intracellular dihydrorhodamine

E. Smits; C. Burvenich; R. Heyneman



Superoxide anion release by peripheral polymorphonuclear leukocytes in welders  

Microsoft Academic Search

Objectives: To investigate peripheral blood neutrophil [polymorphonuclear leukocyte (PMN)] function in a group of 23 welders compared\\u000a with that in an age- and smoking habit-matched non-exposed control group. Methods: Stimulated release of superoxide anions from PMN isolated from peripheral blood of welders and of a matched group was carried\\u000a out. Results: The stimulated release of superoxide anions in PMNs from

Aviva Aloufy; Yehuda Lerman; Ada Tamir; Erica Hoffer



Functional states of polymorphonuclear leukocytes determined by chemiluminescent kinetic analysis.  


During the respiratory burst, upon stimulation with both soluble and particulate matter, polymorphonuclear leukocytes (PMN) generate reactive oxygen species (ROS) and emit chemiluminescence (CL) as a result of metabolic activation. The measurement of CL has been demonstrated to be a useful tool for in vitro assessment of the opsonophagocytic function of PMN. Using component analysis of CL kinetics, we characterized the functional state of PMN by three parameters of the respiratory burst: capacity, effectiveness and velocity (CEV space). The possibility of delimiting eight different functional states of PMN is discussed. The CL kinetics shown by blood PMN in different functional states was analysed, and revealed six out of eight functional states. We conclude that CEV-estimated functional states of PMN are relative, depending on both PMN readiness to generate ROS and conditions of the CL test. PMID:10862142

Magrisso, M Y; Alexandrova, M L; Markova, V I; Bechev, B G; Bochev, P G


Depression of monocyte and polymorphonuclear leukocyte oxidative metabolism and bactericidal capacity by influenza A virus.  

PubMed Central

Decreased host defense against bacterial disease associated with influenza infection may be related to virus-induced changes in phagocytic cell function. Influenza A virus initiates the respiratory burst in peripheral blood monocytes and polymorphonuclear leukocytes, with a peak chemiluminescent response approximately 3 min after virus is added to the cells in vitro. Electron micrographs of phagocytic cells incubated with influenza virus demonstrated virus attached to the cell membrane and within phagocytic vacuoles. After 20 min of incubation of the virus with phagocytic cells, the chemiluminescent response to opsonized zymosan or phorbol myristate acetate was decreased by 30 to 90%. Phagocytic activity of monocytes and polymorphonuclear leukocytes incubate with influenza virus was normal, but the bactericidal activity was significantly depressed. Influenza A virus therefore stimulates an oxidative burst in monocytes as well as polymorphonuclear leukocytes, leading to a subsequent depression of the oxidative metabolic response and bactericidal capacity of the phagocytic cells. Images

Abramson, J S; Mills, E L; Giebink, G S; Quie, P G



Labeling of peripheral blood polymorphonuclear leukocytes with indium-111: a new method for the quantitation of in-vivo accumulation of PMNLs in rabbit skin  

SciTech Connect

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 /sup 111/In 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.

Wahba, A.V.; Barnes, B.; Lazarus, G.S.



Arachidonic Acid Metabolism in Polymorphonuclear Leukocytes: Effects of Ionophore A23187  

Microsoft Academic Search

Addition of arachidonic acid and the divalent cation ionophore A23187 to a suspension of human peripheral blood polymorphonuclear leukocytes led to the formation of (5S)-hydroxy-6,8,11,14-icosatetraenoic acid, (15S)-hydroxy-5,8,11,13-icosatetraenoic acid, and (5S, 12R)-dihydroxy-6,8,10,14-icosatetraenoic acid. A method based on high-pressure liquid chromatography has been developed for assay of these metabolites. The addition of arachidonic acid to human polymorphonuclear leukocytes always resulted in formation

Pierre Borgeat; Bengt Samuelsson



Isolation, In-111 labeling, and abscess detection efficiency of rabbit polymorphonuclear leukocytes (PMN) from blood and peritoneal fluid  

SciTech Connect

In-111 labeled blood and peritoneal exudate PMN were compared for labeling efficiency and ability to migrate to sites of experimental abscesses using both direct sampling and visual imaging techniques. Blood PMN were prepared by combining heparinized blood with 6% Hetastarch for 1 hour and layering the plasma over a double density Ficoll-Hy-paque gradient (S.G. 1.076 over 1.141). The PMN layer (90-99% PMN) at the interface yielded 10/sup 6/-10/sup 7/ PMN from 80-120 ml of blood. Peritoneal PMN were obtained by infusion of 0.1% glycogen, followed by infusion of saline after 4 or 18 hours. The exudate yielded 10/sup 7/-10/sup 8/ PMN (80-99% PMN). PMN suspensions were labeled for 30 minutes by addition of 100 of In-111-oxine, then washed twice. Percent cell-associated radioactivity of the labeled blood, 4 hour, and 18 hour peritoneal PMN was 89%, 88%, and 86%. The labeled PMN were injected intravenously into rabbits which had two of three abdominal capsules (table tennis balls drilled with 250 1.5 mm holes) inoculated with Staphylococcus aureus 4 hours earlier. Peak venous recovery of circulating labeled PMN, for blood, 4 hour and 18 hour peritoneal PMN was 60%, 43%, and 19%. Gamma camera images 24 hours after infusion into infected rabbits were superior with 4 hour peritoneal PMN. The peritoneal PMN harvested 4 hours after glycogen stimulation are simple to prepare, are obtainable in greater numbers than blood PMN, and result in better abscess visualization.

Bettin, K.M.; Elson, M.K.; Gerding, D.N.; Bamberger, D.M.; Forstrom, L.A.; Shafer, R.B.



Polymorphonuclear leukocyte activation and hemostasis in patients with essential thrombocythemia and polycythemia vera  

Microsoft Academic Search

Thrombohemorrhagic complications are a major cause of morbidity and mortality in patients with essential thrombocythe- mia (ET) and polycythemia vera (PV). The pathogenesis of these complications is not completely clarified. Several studies have described abnormalities of red blood cells and platelets in these patients. How- ever, no studies are available on changes in the polymorphonuclear leukocytes (PMNs), which can play

Anna Falanga; Marina Marchetti; Virgilio Evangelista; Alfonso Vignoli; Marina Licini; Mara Balicco; Stefano Manarini; Guido Finazzi; Chiara Cerletti; Tiziano Barbui



Intracellular Penetration and Activity of Gemifloxacin in Human Polymorphonuclear Leukocytes  

PubMed Central

The intracellular penetration and activity of gemifloxacin in human polymorphonuclear leukocytes (PMN) were evaluated. Gemifloxacin reached intracellular concentrations eight times higher than extracellular concentrations. The uptake was rapid, reversible, and nonsaturable and was affected by environmental temperature, cell viability, and membrane stimuli. At therapeutic extracellular concentrations, gemifloxacin showed intracellular activity against Staphylococcus aureus.

Garcia, Isabel; Pascual, Alvaro; Ballesta, Sofia; Joyanes, Providencia; Perea, Evelio J.



Autooxidation as a Basis for Altered Function by Polymorphonuclear Leukocytes  

Microsoft Academic Search

To investigate the possibility that human polymorphonuclear leukocytes (PMN) elaborate sufficient amounts of hydrogen peroxide ( H202) and other radicals of re- duced oxygen to be autotoxic and retard directed cell movement and phagocytosis, the rate of ingestion of opsonized lipo- polysaccharide-paraffin oil particles and movement through Nuclepore filters were studied. Ingestion rates were increased under anaerobic conditions and in

RQbert L. Baehner; Laurence A. Boxer; John M. Allen; Jacqueline Davis



EDU pretreatment decreases polymorphonuclear leukocyte migration into rat lung airways.  


Pretreatment with the heterocyclic compound EDU (N-[2-(2-oxo-1-imidazolindinyl)ethyl]-N'-phenylurea) has previously been shown to reduce polymorphonuclear leukocyte (PMN) infiltration into the airways of ozone-exposed rats. The present study further examined the effects of 1 and 2 days EDU pretreatment on rat lung inflammatory responses by determining PMN infiltration in response to intratracheal instillation with the chemoattractant formyl-norleucine-leucine-phenylalanine (fNLP). Maximal recovery of PMNs by bronchoalveolar lavage was observed 4 hr after fNLP instillation with no alteration in the numbers of recoverable macrophages and lymphocytes. Although 1-day pretreatment with EDU did not affect PMN recovery from fNLP-instilled rat lungs, 2 days of EDU pretreatment prevented PMN infiltration as indicated by PMN recoveries that were similar to those obtained from saline-instilled lungs. Measurements of lung-marginated and interstitial pools of inflammatory cells using collagenase tissue digestion demonstrated no effect of 2 days EDU pretreatment. Although 2 days EDU pretreatment alone did not alter blood PMN content, lung permeability, and the lavage recoveries of inflammatory cells, blood PMN responses to chemotactic stimuli in vitro were impaired. In addition, EDU was shown to directly inhibit PMN chemotaxis and superoxide anion generation in vitro. These data demonstrated that EDU acts by interfering with PMN activation and migration rather than by decreasing PMN availability. EDU, by modulating the inflammatory response, represents a useful compound for preventing PMN-associated amplification of acute lung injuries. PMID:8048056

Bassett, D J; Elbon, C L; Ishii, Y; Yang, H; Otterbein, L; Boswell, G A; Kerr, J S



Effects of lead on the killing mechanisms of polymorphonuclear leukocytes  

SciTech Connect

The effects of lead on the killing mechanisms of rat polymorphonuclear leukocytes (PMN) were investigated, using male Long-Evans rats exposed to 1% lead acetate in the drinking water for varying periods of time to achieve blood lead levels ranging from 20-200 Studies of PMN bacterial and fungal killing activity, chemotaxis and phagocytosis demonstrated that: 1) bactericidal activity of PMN from rats exposed to lead was not altered; 2) chemotactic activity remained within normal limits; 3) the phagocytic ability of the PMN also remained unaltered. In addition to these normal findings, one major abnormality was demonstrated: a significant decrease in the ability of PMN from rats exposed to lead to kill Candida albicans. This defect was not related to age or to length of exposure. It could not be produced by addition of lead to the test system in vitro. Further investigation revealed significant decreases in PMN glucose-6-phosphate dehydrogenase, catalase, and myeloperoxidase activities. These data support two possible mechanisms for the abnormal fungicidal activity of PMN from lead-exposed rats: decrease in ability to reduce oxygen to active metabolites, or reduction in myeloperoxidase activity due to diminshed synthesis of the heme moiety required for its function.

Silberstein, C.F.



Metabolic, Membrane, and Functional Responses of Human Polymorphonuclear Leukocytes to Platelet-Activating Factor  

Microsoft Academic Search

The phospholipid mediator of anaphylaxis. platelet-activat- ing factor (PAF) is chemotactic for polymorphonuclear leukocytes (PMN). We have examined this agents effects on several other PMN functions. Human PMN were pre- pared from heparinized venous blood by Ficoll gradient. Metabolic burst was examined by measurement of #{176}2use and 0 2 production in the presence or absence of PAF (10-109M). Unless cells

Leah M. Ingraham; Thomas D. Coates; John M. Allen; Coleen P. Higgins; Robert L. Baehner; Laurence A. Boxer



Inhibition of human polymorphonuclear leukocyte chemotaxis by oxygenated sterol compounds  

SciTech Connect

When preincubated with certain oxygenated sterol compounds in lipoprotein-depleted serum (20% (vol/vol)), human polymorphonuclear leukocytes show inhibition of chemotaxis toward the synthetic dipeptide N-formylmethionylphenylalinine without alteration of random movement or loss of cell viability. These effects can occur at sterol concentrations as low as 6.25 and after as little as 5 min of preincubation, but they are increased at higher concentrations and longer preincubation times. The inhibition can be almost completely reversed by preincubation in lipoprotein-replete serum (human AB serum, 20% (vol/vol)) and may be partially corrected by addition of free cholesterol (0.125 mM) to the medium. These effects are unlikely to be due to inhibition of cellular sterol synthesis, competition for chemotaxin membrane binding sites, or deactivation of the leukocytes but they may be a consequence of insertion of the sterol molecule into the leukocyte plasma membranes.

Gordon, L.I.; Bass, J.; Yachnin, S.



Influenza A virus-induced polymorphonuclear leukocyte dysfunction in the pathogenesis of experimental pneumococcal otitis media.  

PubMed Central

The role of influenza A virus-induced polymorphonuclear leukocyte and eustachian tube dysfunction in the pathogenesis of acute purulent otitis media was studied in chinchillas. Polymorphonuclear leukocyte function, middle ear pressure, and the incidence of pneumococcal otitis media were observed after intranasal inoculation with influenza A virus, Streptococcus pneumoniae, or both. Results showed that depressed negative middle ear pressure and polymorphonuclear leukocyte chemiluminescence and chemotactic activity occurred after influenza inoculation, but not after inoculation with pneumococcus alone. The greatest incidence of pneumococcal otitis media occurred when the pneumococcus was inoculated just before the time of influenza-induced polymorphonuclear leukocyte dysfunction and negative middle ear pressure. Animals that had unilateral tympanostomy tubes placed before inoculation of influenza with pneumococcus showed no difference in the occurrence of pneumococcal otitis media in ventilated and nonventilated ears, suggesting that polymorphonuclear leukocyte dysfunction contributes more to the pathogenesis of pneumococcal otitis media than does negative middle ear pressure in this animal model.

Abramson, J S; Giebink, G S; Quie, P G



Uptake and intracellular activity of fluconazole in human polymorphonuclear leukocytes.  

PubMed Central

The penetration of fluconazole into human polymorphonuclear leukocytes (PMNs) and tissue culture epithelial cells (McCoy) was evaluated. At different extracellular concentrations (0.5 to 10 mg/liter), fluconazole reached cell-associated concentrations greater than the extracellular ones in either human PMNs (intracellular concentration to extracellular concentration ratio, > or = 2.2) or McCoy cells (intracellular concentration to extracellular concentration ratio, > or = 1.3). The uptake of fluconazole by PMNs was rapid and reversible but was not energy dependent. The intracellular penetration of fluconazole was not affected by environmental pH or temperature. Ingestion of opsonized zymosan and opsonized Candida albicans did not significantly increase the amount of PMN-associated fluconazole. At therapeutic extracellular concentrations, the intracellular activity of fluconazole against C. albicans in PMNs was significantly lower than that of amphotericin B. It was concluded that fluconazole reaches high intracellular concentrations within PMNs but shows moderate activity against intracellular C. albicans in vitro.

Pascual, A; Garcia, I; Conejo, C; Perea, E J



Fluorometric measurement of ofloxacin uptake by human polymorphonuclear leukocytes.  

PubMed Central

A fluorometric assay, based on the natural fluorescence of the quinolone nucleus, was used to determine the uptake of ofloxacin by human polymorphonuclear leukocytes. The ratio of cellular concentration to extracellular concentration (C/E) at 20 min and 37 degrees C was 7.2, using an extracellular concentration of 5 micrograms/ml. Uptake was rapid and was not affected by pH (5 to 9), but required elevated environmental temperature and cell viability. The metabolic inhibitors sodium fluoride and sodium cyanide significantly decreased the uptake of ofloxacin. The penetration of ofloxacin was not affected by the presence of glucose or adenosine, but was decreased by L-amino acids (lysine, leucine, and glycine). These results suggest that ofloxacin could be transported via an amino acid transport system and that the fluorometric assay is a useful method for determining the intracellular penetration of fluoroquinolones, avoiding the use of radiolabeled antimicrobial agents.

Pascual, A; Garcia, I; Perea, E J



Contrasting influence of peplomycin and azelastine hydrochloride (Azeptin) on reactive oxygen generation in polymorphonuclear leukocytes, cytokine generation in lymphocytes, and collagen synthesis in fibroblasts  

Microsoft Academic Search

The influence of peplomycin (PLM) and azelastine hydrochloride (Azeptin) on reactive oxygen (RO) and cytokine generation was examined in human peripheral blood mononuclear leukocytes, polymorphonuclear leukocytes (PMN), and rabbit alveolar macrophages (RAM). In addition, the infleunce of these drugs on DNA and collagen synthesis was investigated in human gingival and rabbit pulmonary fibroblasts. In vitro, PLM increased the FMLP-and PMA-induced

Eisaku Ueta; Tokio Osaki; Kazunori Yoneda; Tetsuya Yamamoto



Nitric oxide synthase in circulating vs. extravasated polymorphonuclear leukocytes.  


It is becoming increasingly apparent that certain forms of acute and chronic inflammation are associated with enhanced production of nitric oxide (NO). Although substantial information has been obtained describing the regulation of NO synthase (NOS) in macrophages, little information is available regarding the biochemistry and molecular biology of NOS in circulating vs. extravasated polymorphonuclear leukocytes (PMNs). The objective of this study was to characterize the molecular and biochemical properties of the inducible NO synthase (iNOS) in circulating vs. extravasated rat and human PMNs. Circulating rat and human PMNs were purified from peripheral blood and extravasated PMNs were elicited in rats by intraperitoneal injection of 1% oyster glycogen or in humans by peritoneal dialysis of patients with peritonitis. Inducible NOS mRNA from circulating and elicited PMNs was quantified using slot blot hybridization analysis with a cDNA probe specific for iNOS. iNOS protein was identified using Western immunoblot analysis, and NOS activity was quantified by measuring the NG-monomethyl-L-arginine (L-NMMA)-inhibitable conversion of 14C-labeled L-arginine to L-[14C]citrulline. In a separate series of experiments, circulating or extravasated PMNs were cultured for 4 h and the accumulation of L-NMMA-inhibitable nitrite (NO2-) in the supernatant was determined and used as a measure of NO production in vitro. We found that circulating PMNs (rat or human) contained no iNOS mRNA, protein, or enzymatic activity. Furthermore, circulating rat or human PMNs (2 x 10(6) cells/well) were unable to generate significant amounts of NO2- when cultured for 4 h in vitro. In contrast, iNOS mRNA levels in 4- and 6-h elicited rat PMNs increased 21- and 42-fold, respectively, when compared with circulating cells. Western blot analysis revealed the presence of iNOS protein in the elicited rat PMNs and iNOS enzymatic activity increased from normally undetectable levels in circulating rat PMNs to 81 and 285 pmol/min/mg for the 4- and 6-h elicited rat PMNs, respectively. Approximately 20-30% of the total iNOS activity was Ca(2+)-dependent. Nitrite formation by elicited rat PMNs in the absence of any exogenous stimuli increased from normally undetectable amounts for circulating PMNs to approximately 8 and 11 microM/10(6) cells for the 4- and 6-h elicited PMNs, respectively. Highly enriched preparations of extravasated human PMNs contained neither message, protein nor iNOS enzymatic activity. Taken together our data demonstrate that inflammation-induced extravasation of rat PMNs upregulates the transcription and translation of iNOS in a time-dependent fashion and that 20-30% of the total inducible NOS is Ca(2+)-dependent. In contrast, neither circulating nor extravasated human PMNs contained iNOS message, protein, or enzymatic activity. These data suggest that the human PMN iNOS gene is under very different regulation than is the rat gene. PMID:7595064

Miles, A M; Owens, M W; Milligan, S; Johnson, G G; Fields, J Z; Ing, T S; Kottapalli, V; Keshavarzian, A; Grisham, M B



Fc? R expression on polymorphonuclear leukocyte and superoxide generation in IgA nephropathy  

Microsoft Academic Search

Fc?R expression on polymorphonuclear leukocyte and superoxide generation in IgA nephropathy. Superoxide (O2?) production and Fc?R antigen expression of circulating polymorphonuclear leukocytes (PMNL) isolated from patients with IgA nephropathy (IgAN) and non-IgA mesangial proliferative glomerulonephritis (PGN) and healthy volunteers were investigated to establish their biological importance in the immunopathogenesis of mesangial proliferative glomerulonephritis. PMNL from both patient groups showed increased

Abul Kashem; Masayuki Endoh; Yasuo Nomoto; Hideto Sakai; Hiroe Nakazawa; Abul Kashem MBBS



Influenza A virus-induced polymorphonuclear leukocyte dysfunction.  

PubMed Central

Previous studies have shown that influenza A virus can activate the polymorphonuclear leukocyte (PMN) respiratory burst and that upon subsequent stimulation of the cell there is depressed metabolic function. We examined the mechanism by which influenza virus causes PMN dysfunction by measuring the effect upon the chemiluminescent activity of cells of varying the type of influenza virus used, the period of time that cells were exposed to virus, and the secondary stimulus that was used. The various types of intact influenza virus elicited different amounts of chemiluminescent activity, but when cells were subsequently stimulated with phorbol myristate acetate, each virus caused equivalent depression of the PMN response. Purified glycoproteins incorporated into a liposome structure similarly stimulated the PMN chemiluminescence, yet did not induce PMN dysfunction. Depressed PMN function was noted after as little as 5 min of incubation of cells with virus and occurred to both receptor-dependent (zymosan, N-formylmethionyl-leucyl-phenylalanine, and phorbol myristate acetate) and -independent (calcium ionophore A23187) stimuli. Images

Abramson, J S; Lyles, D S; Heller, K A; Bass, D A



Effects of irradiation on endothelial cell-polymorphonuclear leukocyte interactions  

SciTech Connect

Prominent early effects of irradiation include neutrophilic vasculitis and interstitial inflammation. To examine the role of the endothelium in these events, bovine aortic endothelial cells (EC) were irradiated (5 Gy) under ambient conditions followed by measurements of neutrophil chemotaxis toward conditioned media and adherence to EC. Neutrophil chemotactic activity increased at 4, 24, and 72 h in both the sham-treated (4.2 +/- 2.5, 15.2 +/- 4.8, and 20.0 +/- 2.7 microns, respectively) and irradiated EC-conditioned media (5.0 +/- 2.1, 18.7 +/- 4.5, and 24.1 +/- 3.4 microns, respectively), and the difference between them was significant at 72 h. The chemoattractant was trypsin sensitive, heat resistant, and chemotactic. It was not present in the EC sonicate. Adherence of neutrophils to EC that were irradiated 4 h earlier (19.3 +/- 4.2%) increased compared with controls (11.1 +/- 2.4%) and was similar to EC pretreated with zymosan-activated serum (22.0 +/- 4.0%), which is a potent inducer of adherence. Thus, following irradiation, bovine aortic EC have greater neutrophil chemotactic activity in their media and are more adherent to polymorphonuclear leukocytes.

Dunn, M.M.; Drab, E.A.; Rubin, D.B.



Interaction of chemotactic factors with human polymorphonuclear leukocytes: Studies using a membrane potential-sensitive cyanine dye  

Microsoft Academic Search

Summary Changes in the fluorescence intensity of the dye 3-3' dipentyloxacarbocyanine were measured in suspensions of purified human peripheral blood polymorphonuclear leukocytes (PMNs) during exposure to the chemotactic factors N-formyl-methionylleucyl-phenylalanine (f-met-leu-phe) and partially purified C5a. Incubation of PMNs with dye resulted in a stable fluorescence reflecting the resting membrane potential of the cell. Exposure of PMNs to dye did not

Bruce E. Seligmann; Elaine K. Gallin; David L. Martin; William Shain; John I. Gallin



Effect of Cyanide on NADPH Oxidation by Granules From Human Polymorphonuclear Leukocytes  

Microsoft Academic Search

man polymorphonuclear leukocytes. It also leukocyte. Based on studies of others, how- stimulates the oxidation of NADPH by a ever, it does not appear as though the particulate fraction derived from phagocy- enzyme is identical to myeloperoxidase. tizing cells. This stimulation of NADPH The mechanism of the CN stimulation oxidase is not observed in the presence of appears to involve

Lawrence R. DeChatelet; Linda C. McPhail; Pamela S. Shirley



Licofelone, a dual lipoxygenase-cyclooxygenase inhibitor, downregulates polymorphonuclear leukocyte and platelet function.  


Polymorphonuclear leukocytes are strongly implicated in the pathogenesis of inflammatory disease. Polymorphonuclear leukocyte recruitment at sites of inflammation, mainly sustained by the beta2-integrins, is followed by the synthesis and release of inflammatory mediators, such as leukotrienes, proteolytic enzymes and reactive oxygen species. Functional and metabolic interactions between polymorphonuclear leukocytes and platelets can contribute to and exacerbate the process. The effects of the dual 5-lipoxygenase and cyclooxygenase inhibitor licofelone ([2,2-dimethyl-6-(4-chlorophenyl)-7-phenyl-2,3-dihydro-1H-pyrrolizine-5-yl]-acetic acid) were studied on arachidonic acid transcellular metabolism occurring between polymorphonuclear leukocytes and platelets. The formation of leukotriene C(4), a leukotriene A(4)-derived metabolite, by mixed polymorphonuclear leukocyte/platelet suspensions stimulated with 10 microM A23187 was inhibited by licofelone with an IC(50) of 3.8 +/- 0.07 microM. The formation of 5,12-di-hydroxy-eicosatetraenoic acid (HETE) was abolished at concentrations > or = 10 microM. Licofelone also inhibited the generation of reactive oxygen species by polymorphonuclear leukocytes stimulated with 1 microM n-formyl-methionyl-leucyl-phenylalanine (fMLP), 10 nM complement fraction 5a (C5a) and 1 microM platelet activating factor (PAF) with IC(50)s of 24.4 +/- 0.6, 11.0 +/- 1.5 and 11.7 +/-1.2 microM; elastase release induced by the three agonists was inhibited with IC(50)s of 12.2 +/- 2.2, 23.5 +/- 8 and 2.6 +/- 1 microM, respectively. Homotypic polymorphonuclear leukocyte aggregation induced by fMLP, C5A and PAF was inhibited by licofelone with IC(50)s of 23.7 +/- 4.8, 15.6 +/- 3.4 and 15.4 +/- 4 microM, respectively. The present study extends the anti-lipoxygenase and anti-cyclooxygenase activities of licofelone to the production of arachidonic acid metabolites generated as a consequence of polymorphonuclear leukocyte-platelet transcellular metabolism and to polymorphonuclear leukocyte responses relevant to the pathogenesis of inflammation. The coexistence within the same molecule of a wide spectrum of anti-inflammatory properties is of interest. PMID:12393068

Rotondo, Serenella; Dell'Elba, Giuseppe; Krauze-Brzósko, Katarzyna; Manarini, Stefano; Martelli, Nicola; Pecce, Romina; Evangelista, Virgilio; Cerletti, Chiara



Changes of membrane fluidity in chemotactic peptide-stimulated polymorphonuclear leukocytes.  


Although the phenomenon of stimulus-response coupling in polymorphonuclear leukocytes involves a series of membrane events the influence of stimulation on membrane fluidity is to clarify. In our experiments we have used 1-(4-trimethylaminophenyl) 6-phenyl-1,3,5-hexatriene and 1,6-diphenyl-1,3,5-hexatriene fluorescence polarization technique to evaluate membrane fluidity in living polymorphonuclear leukocytes after stimulation with N-formyl-methyonil-leucyl-phenylalanine peptide which has a well defined membrane receptor on the plasma membrane. We report that polymorphonuclear leukocytes stimulation increases 1-(4-trimethylaminophenyl)-6-phenyl-1,3,5-hexatriene polarization, only when colcemid, a microtubule disrupting drug, is added to polymorphonuclear leukocytes. This can be viewed as an indirect evidence that microtubules are involved in the control of polymorphonuclear leukocytes membrane fluidity. On the contrary no changes have been observed with 1,6-diphenyl-1,3,5-hexatriene. This study indicates the potential use of 1-(4-trimethylaminophenyl)-6-phenyl-1,3,5-hexatriene to evaluate the involvement of plasma membrane physical state during intact cell activity. PMID:3814118

Valentino, M; Governa, M; Fiorini, R; Curatola, G



Impairment of chemotaxis of polymorphonuclear leukocytes from lead acid battery workers.  


Since lead impairs in vitro the functions of macrophagic cells, we have studied the chemotactic activity of polymorphonuclear leukocytes (PMNs) obtained from lead acid battery workers who were removed from exposure one month before, because they had an abnormal lead absorption. Controls were 18 age matched subjects without any history of occupational lead exposure. Both lead acid battery workers and controls had no alterations of the blood haematological and metabolic parameters. Chemotaxis was carried on in Boyden chambers using zymosan activated serum as chemotactic stimulus. The chemotactic indexes are 56.4 +/- 8.7 in acid battery workers and 75.6 +/- in controls. The difference, which is statistically significant, shows that lead workers have an impairment of PMNs chemotactic activity. PMID:3406721

Governa, M; Valentino, M; Visona, I; Scielso, R



Biosynthesis of leukotriene B4 in human polymorphonuclear leukocytes: regulation by cholesterol and other lipids.  


Leukotriene B4 (LTB4) is one of the most potent chemotactic compounds produced in macrophages and neutrophils. LTB4 is a product of arachidonic acid oxygenation by 5-lipoxygenase pathway. We present here the data on regulation of LT synthesis in human polymorphonuclear leukocytes by cholesterol, cholesterol sulfate and cholesterol phosphate. The addition of Pseudomonas aeruginosa lipopolysaccharides (LPS) with lipid vesicles containing phosphatidylcholine or phosphatidylcholine/cholesterol (70:30) showed that omitting cholesterol abolished the effect of LPS on LT synthesis. We show here the capacity of cholesterol sulfate, the most abundant sulfated sterol in human blood, to suppress LT production in human neutrophils and to neutralize the effect of P. aeruginosa LPS on LT synthesis. We suggest that sulfated lipids serve as specific endogenous regulators of LT synthesis in neutrophils, and anti-inflammatory therapy may be based on modification of cholesterol level and its conversion to anionic derivatives. PMID:19404868

Zagryagskaya, A N; Aleksandrov, D A; Pushkareva, M A; Galkina, S I; Grishina, Z V; Sud'ina, G F



Capsular antibodies induce type-specific phagocytosis of capsulated Staphylococcus aureus by human polymorphonuclear leukocytes.  

PubMed Central

Capsular types 5 and 8, which account for about 70% of Staphylococcus aureus strains isolated from the blood of patients, resisted in vitro phagocytosis by human polymorphonuclear leukocytes (PMN). Antisera and monoclonal antibody to type 5 and 8 capsular polysaccharides (CPS) induced type-specific in vitro phagocytosis of capsulated organisms by PMN. Antibodies directed against the O-acetyl moiety of the type 8 CPS were more effective in inducing phagocytosis of type 8 organisms by PMN. Either type-specific antiserum or monoclonal antibody reactive with the native O-acetylated type 8 CPS was most effective in inducing in vitro phagocytosis of type 8 organisms by PMN. These results provide further evidence that CPS of S. aureus are associated with host immunity to this organism. Images

Karakawa, W W; Sutton, A; Schneerson, R; Karpas, A; Vann, W F



Leukotriene B4 stimulates polymorphonuclear leukocyte adhesion to cultured vascular endothelial cells.  

PubMed Central

Adhesion of polymorphonuclear leukocytes (PMN) to the endothelial lining of blood vessels is an essential component of the inflammatory response. We have examined the effects of various lipoxygenase metabolites of arachidonic acid on PMN adhesion to cultured vascular endothelial cells, using a quantitative monolayer adhesion assay. Our results indicated that leukotriene B4 (LTB4) could effectively stimulate PMN adhesion to endothelial cell surfaces, in contrast to the sulfidopeptide leukotrienes C4, D4, and E4, and the monohydroxyacid lipoxygenase products of leukocytes and platelets, 5S-hydroxy-6-trans-8,11,14-cis-eicosatetraenoic acid and 12S-hydroxy-5,8-cis,10-trans,14-cis-eicosatetraenoic acid, respectively. LTB4-stimulation of PMN-endothelial adhesion did not appear to be dependent upon the generation of cyclooxygenase metabolites, nor was it inhibited by exogenous prostacyclin. Enhanced PMN adhesion was observed with endothelial cells that were cultured from different types of large vessels (arteries and veins) in several species. These findings suggest an important pathophysiologic role for LTB4 in regulating leukocyte-vessel wall interactions.

Gimbrone, M A; Brock, A F; Schafer, A I



Effect of human polymorphonuclear and mononuclear leukocytes on chromosomal and plasmid DNA of Escherichia coli. Role of acid DNase  

SciTech Connect

Phagocytosis and killing by polymorphonuclear and mononuclear leukocytes are important host resistance factors against invading microorganisms. Evidence showing that killing is rapidly followed by degradation of bacterial components is limited. Therefore, we studied the fate of Escherichia coli DNA following phagocytosis of E. coli by polymorphonuclear and mononuclear leukocytes. (/sup 3/H)Thymidine-labeled, unencapsulated E. coli PC2166 and E. coli 048K1 were incubated in serum, washed, and added to leukocytes. Uptake and killing of the bacteria and degradation of DNA were measured. Although phagocytosis and killing by mononuclear leukocytes was less efficient than that by polymorphonuclear leukocytes, only mononuclear leukocytes were able to degrade E. coli PC2166 DNA. Within 2 h, 60% of the radioactivity added to mononuclear leukocytes was released into the supernate, of which 40% was acid soluble. DNA of E. coli 048K1 was not degraded. To further analyze the capacity of mononuclear leukocytes to degrade E. coli DNA, chromosomal and plasmid DNA was isolated from ingested bacteria and subjected to agarose gel-electrophoresis. Only chromosomal DNA was degraded after phagocytosis. Plasmid DNA of E. coli carrying a gene coding for ampicillin resistance remained intact for a 2-h period after ingestion, and was still able to transform recipient E. coli cells after this period. Although we observed no DNA degradation during phagocytosis by polymorphonuclear leukocytes, lysates of both polymorphonuclear and mononuclear leukocytes contained acid-DNase activity with a pH optimum of 4.9. However, the DNase activity of mononuclear leukocytes was 20 times higher than that of polymorphonuclear leukocytes. No difference was observed between DNase activity from polymorphonuclear and mononuclear leukocytes from a chronic granulomatous disease patient with DNase activity from control polymorphonuclear and mononuclear leukocytes.

Rozenberg-Arska, M.; van Strijp, J.A.; Hoekstra, W.P.; Verhoef, J.



Defect of In Vitro Digestive Ability of Polymorphonuclear Leukocytes in Paracoccidioidomycosis  

PubMed Central

Selected functions of polymorphonuclear leukocytes were studied in patients with paracoccidioidomycosis (South American blastomycosis), in healthy control individuals, and in patients with diseases unrelated to paracoccidioidomycosis. Patients with paracoccidioidomycosis were also evaluated by standard immunological techniques. Phagocytosis and digestion of Paracoccidioides brasiliensis yeastlike cells in vitro was estimated by an original method. It was based on the appearance of phagocytosed P. brasiliensis in preparations stained by a modification of the Papanicolaou method and examined with phase-contrast optics. Interpretation of such findings was confirmed by electron microscopy. Two strains of P. brasiliensis were used. Strain 8506 was freshly isolated from a patient. Strain Pb9 was known to be nonpathogenic and to have a peculiar cell wall composition. Yeastlike cells of the Pb9 strain were digested significantly better than those of strain 8506. A higher number of leukocytes per fungus cells led to a higher proportion of digested P. brasiliensis. Leukocytes from patients with paracoccidioidomycosis phagocytosed the fungus in a normal way, but had a significant lower ability to digest it in vitro. When individual cases were analyzed, there was an excellent correlation between clinical evolution and digestive ability of polymorphonuclear leukocytes. There was good correlation between both of these and immunological parameters. Leukocytes from all groups behaved comparably in tests of general leukocyte function and in their abilities to kill and digest Candida albicans. Our results indicate that, as a group, polymorphonuclear leukocytes from patients with paracoccidioidomycosis had a significant, rather specific, defect in their in vitro digestive capacity against phagocytosed P. brasiliensis. There was also an inverse correlation between strain pathogenicity and its susceptibility to in vitro digestion by polymorphonuclear leukocytes. Our findings are concordant with and relevant to clinical reality. ImagesFig. 1

Goihman-Yahr, Mauricio; Essenfeld-Yahr, Ervin; De Albornoz, Maria C.; Yarzabal, Luis; De Gomez, MariA H.; Martin, Blanca San; Ocanto, Ana; Gil, Francisco; Convit, Jacinto



Intracellular Penetration and Activity of DX-619 in Human Polymorphonuclear Leukocytes  

PubMed Central

The intracellular penetration and activity of DX-619 in human polymorphonuclear leukocytes have been evaluated. DX-619 reached intracellular concentrations 10 times higher than the extracellular concentrations reached. Uptake was rapid, reversible, nonsaturable, and affected by environmental temperature, some metabolic inhibitors, and a soluble membrane activator. DX-619 showed intracellular activity against Staphylococcus aureus.

Garcia, Isabel; Ballesta, Sofia; Murillo, Concepcion; Perea, Evelio J.; Pascual, Alvaro



Pseudomonas aeruginosa tolerance to tobramycin, hydrogen peroxide and polymorphonuclear leukocytes is quorum-sensing dependent  

Microsoft Academic Search

The opportunistic human pathogen Pseudomonas aeruginosa is the predominant micro-organism of chronic lung infections in cystic fibrosis (CF) patients. P. aeruginosa colonizes the CF lungs by forming biofilm structures in the alveoli. In the biofilm mode of growth the bacteria are highly tolerant to otherwise lethal doses of antibiotics and are protected from bactericidal activity of polymorphonuclear leukocytes (PMNs). P.

Thomas Bjarnsholt; Peter Østrup Jensen; Mette Burmølle; Morten Hentzer; Janus A. J. Haagensen; Hans Petter Hougen; Henrik Calum; Kit G. Madsen; Claus Moser; Søren Molin; Niels Høiby; Michael Givskov



Altered Ca2+ homeostasis in polymorphonuclear leukocytes from chronic myeloid leukaemia patients  

Microsoft Academic Search

BACKGROUND: In polymorphonuclear leukocytes (PMNL), mobilization of calcium ions is one of the early events triggered by binding of chemoattractant to its receptors. Besides chemotaxis, a variety of other functional responses are dependent on calcium ion mobilization. PMNL from chronic myeloid leukaemia (CML) patients that were morphologically indistinguishable from normal PMNL were found to be defective in various functions stimulated

Chetana M Revankar; Suresh H Advani; Nishigandha R Naik



Synergistic effect of azithromycin on the phagocytic killing of Staphylococcus aureus by human polymorphonuclear leukocytes  

Microsoft Academic Search

Many macrolides have been shown to affect the interaction between bacteria and various immune defense mechanisms, such as chemotaxis, accumulation, and bioactivity within phagocytic cells. The interaction of azithromycin with human polymorphonuclear leukocytes (PMNs) was studied in vitro and compared with the interactions between other macrolides and PMNs. The opsonophagocytic killing ofStaphylococcus aureus was synergistically enhanced by azithromycin at concentrations

I. Herrera-Insúa; P. Pérez; C. Ramos; P. Martínez; M. L. Gómez-Lus; J. Prieto



Human polymorphonuclear leukocyte phagocytosis of crystalline cholesterol, bilirubin, and calcium hydroxyapatite in vitro  

Microsoft Academic Search

We tested the hypothesis that human polymorphonuclear leukocytes (PMNs) phagocytize crystalline cholesterol, bilirubin, or calcium hydroxyapatitein vitro and in the process release oxygen metabolites and enzymes involved in the inflammatory process. Chemiluminescence (CL), elicited by the respiratory burst (release and activation of oxygen metabolites and enzymes) of PMNs during phagocytosis of a target particle, was used to quantitate PMN phagocytosis

Jay B. Prystowsky; Jayashree S. Huprikar; Alfred W. Rademaker; Robert V. Rege




Microsoft Academic Search

Human neutrophil polymorphonuclear leukocytes (PMN) were studied to deter- mine the influence of cellular locomotion upon the redistribution and capping of concanavalin A (Con A) . Con A was detected by fluorescence (using Con A conjugated to fluorescein isothiocyanate (Con A-FITC)), or on shadow-cast replicas (using Busycon canaliculatum hemocyanin as a marker for Con A). After labeling with Con A




Participation of peripheral polymorphonuclear leukocytes in the oxidative stress and inflammation in patients with essential hypertension  

Microsoft Academic Search

Oxidative stress and inflammation have recently been linked to endothelial damage in essential hypertension (EH). Activated peripheral polymorphonuclear leukocytes (PMN) damage surrounding tissue by releasing reactive oxygen species (ROS) and proteolytic enzymes before self-necrosis. PMN necrosis further exacerbates inflammation and promotes chemotaxis and PMN recruitment. The number and properties of PMN from untreated EH patients is the focus of the

Batya Kristal; Revital Shurtz-Swirski; Judith Chezar; Joseph Manaster; Rivka Levy; Galina Shapiro; Irith Weissman; ShaulM Shasha; Shifra Sela



Modulation of polymorphonuclear leukocyte adherence by pulpotomy medicaments: effects of formocresol, glutaraldehyde, eugenol, and calcium hydroxide  

Microsoft Academic Search

Activated polymorphonuclear leukocytes (PMNs) release lysosomal enzymes and toxic oxygen-free radicals into their immediate environment. The persistent activation of PMNs by pulpotomy medicaments may contribute to the chronic inflammatory changes and root resorption seen in histologic sections. The authors examined the effects of pulpotomy medicaments commonly used in pediatric dentistry on PMN adherence, the earliest observable change in PMN behavior

W. Kim Seow; Y. H. Thong



Cigarette Smoking Causes Sequestration of Polymorphonuclear Leukocytes Released from the Bone Marrow in Lung Microvessels  

Microsoft Academic Search

Studies from our laboratory have shown that chronic cigarette smoke exposure causes a neutrophilia asso- ciated with a shortening of the mean transit time of polymorphonuclear leukocytes (PMN) though the post- mitotic pool of the marrow. The present study was designed to test the hypothesis that PMN newly re- leased from bone marrow by smoke exposure preferentially sequestered in pulmonary

Takeshi Terashima; Maria E. Klut; Dean English; Jennifer Hards; James C. Hogg; Stephan F. van Eeden


Human Polymorphonuclear Leukocytes Inhibit Aspergillus fumigatus Conidial Growth by Lactoferrin-Mediated Iron Depletion1  

Microsoft Academic Search

Aspergillus fumigatus, a common mold, rarely infects humans, except during prolonged neutropenia or in cases of chronic gran- ulomatous disease (CGD), a primary immunodeficiency caused by mutations in the NADPH oxidase that normally produces fungicidal reactive oxygen species. Filamentous hyphae of Aspergillus are killed by normal, but not CGD polymorphonuclear leukocytes (PMN); however, the few studies on PMN-mediated host defenses

Kol A. Zarember; Janyce A. Sugui; Yun C. Chang; Kyung J. Kwon-Chung; John I. Gallin


Increased hydroxyl radical generation by normal polymorphonuclear leukocytes incubated in sera from patients with leukocytoclastic vasculitis  

Microsoft Academic Search

The effect of sera from patients with untreated leukocytoclastic vasculitis was investigated on the generation of oxygen intermediates by normal polymorphonuclear leukocytes. Sera from untreated patients induced increased hydroxyl radical generation, which is one of the most potent oxidants capable of causing tissue damage. It is suggested that vascular injury may be mediated in part by enhanced production of hydroxyl

Yoshiki Miyachi; Keiko Yanase; Sadao Imamura; Yukie Niwa



Dynamic component chemiluminescent sensor for assessing circulating polymorphonuclear leukocyte activity of peritoneal dialysis patients.  


Recurrent bacterial peritonitis is a major complication in peritoneal dialysis (PD) patients, which is associated with polymorphonuclear leukocyte (PMN) functional changes and can be assessed by a chemiluminescent (CL) reaction. We applied a new approach of a dynamic component chemiluminescence sensor for the assessment of functional states of PMNs in a luminol-amplified whole-blood system. This method is based on the evaluation of CL kinetic patterns of stimulated PMNs, while the parallel measurements of intracellular and extracellular production of reactive oxygen species (ROS) from the same sample can be conducted. Blood was drawn from diabetic and nondiabetic patients during follow-up, and during peritonitis. Healthy medical personnel served as the control group. Chemiluminescence curves were recorded and presented as a sum of three biological components. CL kinetic parameters were calculated, and functional states of PMNs were assessed. Data mining algorithms were used to build decision tree models that can distinguish between different clinical groups. The induced classification models were used afterward for differentiating and classifying new blind cases and demonstrated good correlation with medical diagnosis (84.6% predictive accuracy). In conclusion, this novel method shows a high predictive diagnostic value and may assist in detection of PD-associated clinical states. PMID:18510343

Prilutsky, Daria; Rogachev, Boris; Vorobiov, Marina; Zlotnik, Moshe; Last, Mark; Lobel, Leslie; Marks, Robert S



Effects of statins on oxidative stress and primed polymorphonuclear leukocytes in hyperlipidemic patients.  


Inflammation and oxidative stress are among the factors that have been implicated in the pathogenesis of hyperlipidemia. In metabolic syndrome and hyperlipidemic patients, peripheral polymorphonuclear leukocytes (PMNL) are primed and they release uncontrolled superoxide that contributes to oxidative stress and inflammation. Recent studies have demonstrated that the anti-hyperlipidemic drug, Atrovastatin effects improvement in endothelial function, exhibits anti-oxidative characteristics and reduces lipid markers of oxidation. To evaluate possible nontraditional effects of treatment with Atrovastatin on PMNL priming, oxidative stress and inflammation in hyperlipidemia, 50 non-smoking hyperlipidemic patients were treated for 6 months with Atrovastatin and compared to age and gender-matched healthy controls. PMNL priming was assessed by the rate of superoxide release from separated, phorbol ester-stimulated PMNL and by PMNL-CD11b levels. Inflammation was reflected by blood inflammatory markers including albumin, transferrin, C-reactive protein (CRP) and fibrinogen levels, white blood cells (WBC), PMNL counts and PMNL apoptosis. Atrovastatin treatment showed a reduction in PMNL priming, PMNL apoptosis, fibrinogen and CRP levels concomitant with decreased lipid levels. Atrovastatin may be preferred for hyperlipidemic patients owing to its combined anti-PMNL priming and anti-inflammatory effects in addition to its anti-atherogenic effects. PMID:22989353

Farah, R; Jubran, F; Khamisy-Farah, R



Experimental bacterial pneumonia in rabbits: polymorphonuclear leukocyte margination and sequestration in rabbit lungs and quantitation and kinetics of /sup 51/Cr-labeled polymorphonuclear leukocytes in E. coli-induced lung lesions  

SciTech Connect

A relationship between the circulating and marginal polymorphonuclear leukocyte (PMN) pools was documented using /sup 51/Cr-labeled leukocytes as a marker. /sup 51/Cr-leukocytes marginating in the lungs were found to decrease following a first-order exponential decline, while /sup 51/Cr radioactivity accumulated in the liver and the spleen. Intravenously administered endotoxin caused a rapid selective disappearance of PMNs from the circulation. The percentage of infused /sup 51/Cr cells disappearing was equal to the percentage of disappearance of host cells. The PMNs were found to sequester in the lungs, with peak sequestration of labeled cells occurring 5 min after an endotoxin challenge. Over the next 25 min the /sup 51/Cr radioactivity in the lungs declined. Large numbers of PMNs, probably newly derived from the bone marrow, were observed histologically to be sequestered in the lung vasculature 90 min after an endotoxin dose, while the early sequestration of circulating leukocytes could not be assessed histologically. Pulmonary inflammatory lesions were induced selectively with Escherichia coli in the left lower lobes of rabbits, leaving the right lower lobes as intrinsic controls. PMN-accumulation into the lesions was quantitated using /sup 51/Cr-labeled blood leukocytes. With the aid of /sup 125/I-labeled E. coli, a logarithmic dose-response relationship was found between the number of E. coli and of PMNs. Over a 6-hr period circulating PMNs were found to accumulate in a lesion in the left lower lobe, whereas in the control right lower lobe, leukocyte radioactivity declined. These findings were confirmed with the aid of lavages of the right and left lungs. Two peaks of PMN-accumulation were found by studying leukocyte kinetics: a larger peak between 0 and 6 hr and a smaller peak 18-24 hr after instillation of the microorganisms. Histologic studies confirmed the accumulation of leukocytes, and by 3 weeks showed a complete resolution of the lesions.

Cybulsky, M.I.; Movat, H.Z.



Mode of activation of the metabolic burst in polymorphonuclear leukocytes by calcium oxalate crystals.  


Microcrystals of calcium oxalate cause an activation of the metabolic burst in polymorphonuclear leukocytes, measured as NBT reduction. Crystal-induced NBT reduction, which was mainly due to superoxide release, was accompanied by enzyme release. Modulation of calcium oxalate-induced activation by several agents resembles the effect of these agents on activation by opsonized zymosan and by phorbol myristate acetate. Both the activation of the metabolic burst and concomitant enzyme release could be counteracted by certain anions, such as oxalate, pyruvate and citrate, indicating that positive charges on the crystals play an important role in crystal-cell interaction. Removal of the negatively charged sialic acid from the cell surface by neuraminidase did not affect the the action of the crystals. The mechanism, by which calcium oxalate activates polymorphonuclear leukocytes, is discussed. PMID:3445822

Elferink, J G



Effect of human polymorphonuclear and mononuclear leukocytes on chromosomal and plasmid DNA of Escherichia coli. Role of acid DNase  

Microsoft Academic Search

Phagocytosis and killing by polymorphonuclear and mononuclear leukocytes are important host resistance factors against invading microorganisms. Evidence showing that killing is rapidly followed by degradation of bacterial components is limited. Therefore, we studied the fate of Escherichia coli DNA following phagocytosis of E. coli by polymorphonuclear and mononuclear leukocytes. (³H)Thymidine-labeled, unencapsulated E. coli PC2166 and E. coli 048K1 were incubated

M. Rozenberg-Arska; J. A. van Strijp; W. P. Hoekstra; J. Verhoef



The activation of gold complexes by cyanide produced by polymorphonuclear leukocytes--I. The effects of aurocyanide on the oxidative burst of polymorphonuclear leukocytes.  


It has been suggested that the antiarthritic gold complex, aurothiomalate (Autm), is activated by its conversion to aurocyanide by polymorphonuclear leukocytes (PMN) which generate cyanide from thiocyanate. In an examination of this hypothesis, a study has been conducted on the effects of aurocyanide on the oxidative burst of polymorphonuclear leukocytes (PMN) and monocytes activated by phorbol myristate acetate (PMA). Aurocyanide produced delayed inhibition of the oxidative burst as shown by its effect on both lucigenin and luminol-dependent chemiluminescence and on the production of superoxide. It was a more potent inhibitor of luminol-dependent chemiluminescence than free thiomalate and other by-products of the reaction between Autm and cyanide. Aurocyanide had a biphasic effect on the PMA-stimulated hexose monophosphate shunt of PMN, with enhancement at 0.1 microM and inhibition at 10 and 100 microM. The activity of aurocyanide was also compared with that of auranofin, an orally active gold complex, which inhibits a variety of functions of PMN and monocytes. At low concentrations, auranofin produced delayed inhibition of chemiluminescence in a similar fashion to aurocyanide but at high concentrations was an immediate inhibitor of the oxidative burst. PMID:2160817

Rudkowski, R; Graham, G G; Champion, G D; Ziegler, J B



Enhancement of Hexose Uptake in Human Polymorphonuclear Leukocytes by Activated Complement Component C5a  

Microsoft Academic Search

The polymorphonuclear leukocyte (PMNL) depends on glucose as a source of energy for motility, chemotaxis, phagocytosis, and bactericidal activity. Activated complement (C5a) at low concentrations stimulates carrier-mediated carbohydrate transport in PMNLs as measured by the uptake of 2-deoxy-D-[3H]glucose. Human PMNLs were preincubated at 37 degrees C for 15 min with zymosan-activated human serum or various purified preparations of human C5a.

Charles E. McCall; David A. Bass; Sue Cousart; Lawrence R. Dechatelet



Interaction of mammalian cells with polymorphonuclear leukocytes: Relative sensitivity to monolayer disruption and killing  

Microsoft Academic Search

Monolayers of murine fibrosarcoma cells that had been treated either with histone-opsonized streptococci, histone-opsonizedCandida globerata, or lipoteichoic acid-anti-lipoteichoic acid complexes underwent disruption when incubated with human polymorphonuclear leukocytes (PMNs). Although the architecture of the monolayers was destroyed, the target cells were not killed. The destruction of the monolayers was totally inhibited by proteinase inhibitors, suggesting that the detachment of the

Isaac Ginsburg; Douglas F. Gibbs; James Varani



Biochemical characteristics of ATP-induced secretion of lysosomal enzymes from rabbit polymorphonuclear leukocytes  

Microsoft Academic Search

ATP induces a release of the lysosomal enzymes,ß-glucuronidase and lysozyme, but not the cytoplasmic marker, lactic dehydrogenase, from rabbit peritoneal polymorphonuclear leukocytes. The release requires a low-ionic-strength medium but not divalent cations. It occurs to a greater extent in the presence of K+ than Na+, is greatly enhanced by cytochalasin B, is temperature dependent, and apparently requires a source of

Elmer L. Becker; Peter M. Henson



Suppression of respiratory burst of polymorphonuclear leukocytes by Azelastine hydrochloride (Azeptin)  

Microsoft Academic Search

The inhibitory action of azelastine hydrochloride (Azeptin) on the respiratory burst in peripheral polymorphonuclear leukocytes (PMN) and pulmonary alveolar macrophages (PAM) has been studied. Azeptin in vitro suppressed chemiluminescence and superoxide (O2-) generation by human PMN in a dose-and time-dependent manner. Phorbol myristyl acetate (PMA) and formyl-methionyl-leucyl-phenylalanine (FMLP)-induced O2-generation were strongly suppressed by 10-6M and 10-5M Azeptin, respectively. PMN and

E. Ueta; T. Osaki; N. Kawasaki; Y. Nomura



Morphometric cytochemistry of catalase and myeloperoxidase-containing granules in the rabbit polymorphonuclear leukocyte  

Microsoft Academic Search

Summary  Recently developed morphometric and statistical techniques were applied to the study of heterogeneity of the granule population of rabbit polymorphonuclear leukocytes. The cytochemical activities of myeloperoxidase and catalase were differentiated by incubation at pH 7.6, and pH 9.7 to 10.5, respectively. Each activity was found in more than one granule. Statistical evaluation suggested that in addition to the primary granule,

D. M. Zellmer; W. A. Shannon



Effect of Ciprofloxacin on Killing of Actinobacillus actinomycetemcomitans by Polymorphonuclear Leukocytes  

Microsoft Academic Search

Actinobacillus actinomycetemcomitans, a pathogen associated with aggressive periodontitis, resists phagocytic killing by polymorphonuclear leukocytes (PMNs). It is susceptible to ciprofloxacin, which PMNs actively accumulate. This study tested the hypothesis that ciprofloxacin-loaded PMNs are more effective at killing A. actinomycetemcomitans than control PMNs. Isolated human PMNs were loaded by brief incubation with 0.5 g of ciprofloxacin\\/ml. Opsonized bacteria (ATCC 43718) were

David A. Cacchillo; John D. Walters



Pulmonary accumulation of polymorphonuclear leukocytes in the adult respiratory distress syndrome  

Microsoft Academic Search

The polymorphonuclear leukocyte (PMN) plays an integral role in the development of permeability pulmonary edema associated with the adult respiratory distress syndrome (ARDS). This report describes 3 patients with ARDS secondary to systemic sepsis who demonstrated an abnormal diffuse accumulation of Indium (¹¹¹In)-labeled PMNs in their lungs, without concomitant clinical or laboratory evidence of a primary chest infection. In one




Characterization of serum-independent polymorphonuclear leukocyte chemotactic factors produced by Propionibacterium acnes  

Microsoft Academic Search

The size and production of polymorphonuclear leukocyte (PMN) chemotactic factors byPropionibacterium acnes has been studied. All eight strains ofP. acnes which were tested liberated PMN chemotactic factors in their growth culture media. The factor(s) produced by one strain, 6919, were studied in greater depth. The PMN response was proportional to the dose of culture supernatant and chemotactic activity increased with

Guy F. Webster; James J. Leyden



Effect of the lipopolysaccharide antagonist Planktothrix sp. FP1 cyanobacterial extract on human polymorphonuclear leukocytes.  


CyP is a lipopolysaccharide (LPS)-like molecule extracted from the freshwater cyanobacterium Oscillatoria planktothrix FP1, which has been reported to be a potent competitive inhibitor of bacterial LPS. In the present study the ability of CyP to affect human polymorphonuclear leukocyte (PMN) function was investigated. PMNs were isolated from venous blood by standard density-gradient centrifugation. Cell migration was measured by use of the Boyden chamber assay. Interleukin (IL)-8 and tumor necrosis factor (TNF)-? production was measured using a sandwich-type enzyme-linked immunosorbent assay. PMN intracellular reactive oxygen species (ROS) levels were assessed by the use of a fluorescent probe coupled to spectrophotometry. CyP 10-100 ?g/ml was chemotactic for PMNs without affecting the chemotactic response to either E. coli LPS or N-formyl-Met-Leu-Phe (fMLP). CyP per se did not affect PMN production of either IL-8 or TNF-?, but concentration-dependently reduced LPS-induced production of both cytokines. On the contrary, CyP had no effect either on fMLP-induced production of IL-8 or on PMN oxidative burst (at rest and after stimulation with fMLP), a response which is known to be independent from LPS-operated pathways. In human PMNs CyP behaves as a selective and effective LPS antagonist. These findings support the therapeutic potential of CyP in endotoxin-dependent disease. PMID:21115122

Maio, Ramòna Consuèlo; Cosentino, Marco; Rossetti, Carlo; Molteni, Monica; Lecchini, Sergio; Marino, Franca



Appearance of Fc receptors on polymorphonuclear leukocytes after migration and their role in phagocytosis.  

PubMed Central

The effect of the migration of bovine blood polymorphonuclear leukocytes (PMNs) in vitro on their phagocytic activity was studied. PMNs were examined before and after migration through various membranes for rosette formation with sensitized sheep erythrocytes to detect Fc receptors (FcRs), phagocytic activity mediated through FcRs with opsonized staphylococci (Smith strain), and phagocytic activity mediated through nonimmunological receptors with unopsonized staphylococci (strain 305). Migration of PMNs was observed from the upper to the lower compartment of the blind well chamber through Millipore and Nuclepore membranes; through Millipore, Nuclepore, and nylon membranes coated with collagen; and through collagen-coated Millipore, Nuclepore, and nylon membranes overlaid with MA-104, BHK-21, MDBK-99, TB, or FBHE cells. Random migration of PMNs toward the plain medium (the same medium in the upper and lower compartments) through the membranes with and without a monolayer of cells increased the percentage of PMNs forming rosettes. In contrast, migration toward the medium containing lipopolysaccharide (LPS), formyl-L-methionyl-L-leucyl-L-phenylalanine (FMLP), or zymosan-activated serum (Act. serum) did not change the percentage of PMNs forming rosettes. The increased percentage of PMNs forming rosettes was associated with the enhanced phagocytosis of opsonized staphylococci (mediated by FcRs). In contrast, migration of PMNs toward LPS, FMLP, or Act. serum did not enhance phagocytosis mediated through FcRs. However, PMNs after migration toward LPS, FMLP, Act. serum, and plain medium enhanced phagocytosis of unopsonized staphylococci (mediated through the nonimmunological receptors). Images

Targowski, S P; Niemialtowski, M



Phagocytosis and killing of Actinobacillus pleuropneumoniae by alveolar macrophages and polymorphonuclear leukocytes isolated from pigs.  

PubMed Central

To study the cellular response of phagocytic cells to Actinobacillus pleuropneumoniae, we investigated whether porcine alveolar macrophages (AM) and polymorphonuclear leukocytes (PMN) are able to phagocytize and intracellularly kill A. pleuropneumoniae in vitro. Bacterial cultivation methods of A. pleuropneumoniae were used to assess in vitro phagocytosis and the ability to kill. A specific-pathogen-free pig was killed, blood was collected, and PMN were isolated and counted. The AM were isolated by lung lavage of the same animal and counted. In addition, convalescent-stage serum was collected from a specific-pathogen-free pig that was infected with A. pleuropneumoniae. Both porcine AM and porcine PMN effectively phagocytized A. pleuropneumoniae in the presence of convalescent-stage pig serum. PMN killed 90 to 99% of the bacteria intracellularly, whereas AM did not. Because A. pleuropneumoniae produces exotoxins that kill porcine AM and porcine PMN, we incubated equal amounts of bacteria and phagocytic cells and tested the viability of the cells 120 min later. In the presence of convalescent-stage pig serum, A. pleuropneumoniae was toxic to AM but not to PMN. Probably in porcine AM, intracellular released toxins of A. pleuropneumoniae lessen the ability of the cell to kill the bacterium. Consecutive lysis of AM and release of viable A. pleuropneumoniae may initiate the characteristic porcine pleuropneumonia.

Cruijsen, T L; Van Leengoed, L A; Dekker-Nooren, T C; Schoevers, E J; Verheijden, J H



An evaluation of the role of leukocytes in the pathogenesis of experimentally induced corneal vascularization. III. Studies related to the vasoproliferative capability of polymorphonuclear leukocytes and lymphocytes.  

PubMed Central

Studies in the past have suggested that leukocytes are a prerequisite to corneal vascularization. To test this hypothesis further, experiments were conducted to determine whether the intracorneal instillation of polymorphonuclear leukocytes, lymphocytes, or components of leukocytes would induce a corneal vascular ingrowth. These cells or cellular fractions were injected intracorneally into Fisher albino rats whose circulating leukocytes had been depleted by total body x-irradiation. Polymorphonuclear leukocytes isolated from glycogen-induced peritoneal exudates caused a corneal vascular invasion, but lymphocytes obtained from thymus, spleen, and lymph nodes failed to do so. To learn whether an extractable factor could be isolated from polymorphonuclear leukocytes these cells were suspended in isotonic saline, ultrasonified and then centrifuged at 101,952g for 1 hour. Aliquots of the resulting sediment and supernatant were injected intracorneally into rats with radiation-induced leukopenia. The nonsedimentable supernatant caused corneal vascularization, but the sediment did not provoke the phenomenon. These studies not only provide further support for the hypothesis that leukocytes initiate corneal vascularization, possibly by the release of one or more heat labile chemical mediators, but directly implicate the polymorphonuclear leukocyte in this process. Images Figures 1-3 Figure 4 Figure 5 Figure 6

Fromer, C. H.; Klintworth, G. K.



The effect of taurine on polymorphonuclear leukocyte functions in endotoxemia.  


The aim of the present study was to measure MPO activity in PMN leukocytes after endotoxin administration, and to compare the levels of NO2- competing with taurine for reaction with HOCl. Furthermore we aimed to determine TauCl levels, a product of MPO-H2O2-Halide system, and to evaluate anti-inflammatory properties of PMN in endotoxemia. In addition, our second objective was to investigate the effect of taurine, an antioxidant amino acid, on anti-bactericidal and anti-inflammatory functions of PMN after administration of endotoxin together with taurine. All experiments were performed with four groups (control, taurine, endotoxemia, and taurine plus endotoxin) of ten guinea pigs. After endotoxin administration (4 mg/kg), MPO activities increased and taurine levels decreased. Therefore levels of TauCl, NO2*- increased. We observed the effects of taurine as conflicting. When taurine was administrated alone (300 mg/kg), all of these parameters decreased.Consequently, we suggested that taurine is influential in infected subjects but not on healthy ones as an antioxidative amino acid. In addition, we believe that in vivo effects of taurine may differ from those in vitro depending on its dosage. PMID:17520328

Erdamar, H; Türközkan, N; Ekremo?lu, M; Kurt, Y; Yaman, H



Effects of lead on polymorphonuclear leukocyte (PMN) functions in occupationally exposed workers.  


Previous in vitro experiments have shown that lead can inhibit PMN chemotaxis, phagocytosis and superoxide formation. Moreover, we have observed an inhibition of PMN chemotaxis in workers occupationally exposed to lead with a mean blood lead concentration of 3.06 mumol/l. The present study was carried out to evaluate locomotion and luminol assisted chemiluminescence (CL) of polymorphonuclear leukocytes (PMN) harvested from ten lead occupationally exposed workers with blood lead concentrations of 1.59 mumol/l (SD 0.27 mumol/l). Since lipids affect PMN activity and lipid composition is modified in erythrocytes of lead workers, PMN lipids were also studied. Ten healthy male subjects of the same age were taken as controls. Chemotaxis, i.e. locomotion stimulated through a specific membrane receptor, was impaired in the PMN of lead workers, but random migration, i.e. unstimulated cell locomotion, and respiratory burst were both unmodified. Cholesterol and phospholipids were not changed, but the percentage of arachidonic acid was significantly increased. The release of LTB4, generated by the oxidative metabolism of arachidonic acid, was increased. CL, which detects reactive oxygen species (ROS), was unmodified, but this lack of change could be the result of an increase in ROS, due to the augmentated percentage of arachidonic acid, and of a decrease in ROS, due to a direct inhibitory effect of lead on ROS generation. On the basis of the results from these ex vivo experiments, the conclusion that chemotaxis is the PMN function primarily affected by lead was confirmed. PMN are considered to be one of the first cellular targets for the action of lead; low exposure to lead modifies their activity and mainly modifies chemotaxis and LTB4 production. PMID:1660709

Valentino, M; Governa, M; Marchiseppe, I; Vison?, I



Polymorphonuclear leukocytes present laminin peptides in endocytic compartments.  


Rat and Human neutrophils presented cytoplasmic vacuoles immunoreactive for laminin at the electron microscopy level. Colocalization of the anti-laminin labeling with albumin-gold complexes and alkaline phosphatase activity in rat PMN suggest an endocytic nature for this compartment. Immunoblot analysis of human normal peripheral blood neutrophils revealed the presence of two laminin peptides around 100 kDa. PMID:8630048

Vannier-Santos, M A; de Souza, S J; Brentani, R R; de Souza, W



Are primed polymorphonuclear leukocytes contributors to the high heparanase levels in hemodialysis patients?  


Patients on chronic hemodialysis (HD) are at high risk for developing atherosclerosis and cardiovascular complications. Heparanase, an endoglycosidase that cleaves heparan sulfate (HS) side chains of proteoglycans, is involved in extracellular matrix degradation and, as such, may be involved in the atherosclerotic lesion progression. We hypothesize that heparanase is elevated in HD patients, partly due to its release from primed circulating polymorphonuclear leukocytes (PMNLs), undergoing degranulation. Priming of PMNLs was assessed by levels of CD11b and the rate of superoxide release. Heparanase mRNA expression in PMNLs was determined by RT-PCR. PMNL and plasma levels of heparanase were determined by immunoblotting, immunofluorescence, and flow cytometry analyses. The levels of soluble HS in plasma were measured by a competition ELISA. This study shows that PMNLs isolated from HD patients have higher mRNA and protein levels of heparanase compared with normal control (NC) subjects and that heparanase levels correlate positively with PMNL priming. Plasma levels of heparanase were higher in HD patients than in NC subjects and were further elevated after the dialysis session. In addition, heparanase expression inversely correlates with plasma HS levels. A pronounced expression of heparanase was found in human atherosclerotic lesions. The increased heparanase activity in the blood of HD patients results at least in part from the degranulation of primed PMNLs and may contribute to the acceleration of the atherosclerotic process. Our findings highlight primed PMNLs as a possible source for the increased heparanase in HD patients, posing heparanase as a new risk factor for cardiovascular complications and atherosclerosis. PMID:18032524

Cohen-Mazor, Meital; Sela, Shifra; Mazor, Rafi; Ilan, Neta; Vlodavsky, Israel; Rops, Angelique L; van der Vlag, Johan; Cohen, Hector I; Kristal, Batya



In vitro effect of tobacco smoke components on the functions of normal human polymorphonuclear leukocytes.  


The function of polymorphonuclear leukocytes (PMNs) has previously been shown to be impaired in smokers in comparison with healthy nonsmokers. Potent inhibition of PMN chemotaxis has been achieved with whole tobacco smoke, the gas phase of smoke, and a water-soluble extract of whole smoke. In the present work several aspects of PMN function were studied after exposure to water-soluble fraction of the particle phase of tobacco smoke collected on glass fiber filters. These tests included capillary tube random migration, chemotaxis under agarose, phagocytosis of yeasts, Nitro Blue Tetrazolium dye reduction, and whole-blood bactericidal activity. The water extract of the particle fraction of smoke had a high content of nicotine when compared with the levels achieved in plasma of smokers and a much lower concentration of aldehydes when compared with the gas phase of smoke. It had no cytotoxic effect and did not affect phagocytosis, oxygen consumption, or bactericidal activity. Nitro Blue Tetrazolium reduction of both resting and stimulated PMNs was significantly decreased only with the most concentrated solution. The tested solutions exerted a dose-related depressive effect on capillary tube random migration, whereas the random migration measured in the agarose chemotaxis test was normal. Nevertheless, the chemotactic response to a caseine solution was significantly decreased. The same tests were performed in the presence of several concentrations of a nicotine solution and the only test to be affected was the capillary tube random migration, and, that only at a very high concentration. The results of this study contribute to the more precise delineation of the extent of the dysfunction of PMNs exposed to tobacco smoke components and indicate that deleterious products are released from the particle phase of the smoke, which deposits all along the respiratory tree. PMID:7228386

Corberand, J; Laharrague, P; Nguyen, F; Dutau, G; Fontanilles, M; Gleizes, B; Gyrard, E



Production of reactive oxygen species by man-made vitreous fibres in human polymorphonuclear leukocytes.  


Human polymorphonuclear leukocytes (PMNL) or erythrocytes, isolated from human blood, were exposed to graded doses of asbestos (chrysotile), quartz, or man-made vitreous fibres (MMVF), i.e. refractory ceramic fibres (RCF), glasswool, or rockwool fibres. None of the MMVF affected either the viability of PMNL, as measured by trypan blue exclusion test, or induced haemolysis, whereas the positive controls, quartz and chrysotile, dose-dependently induced haemolysis in PMNL. MMVF did not increase the release of lactate dehydrogenase (LDH) from the PMNL, whereas the positive controls, chrysotile and quartz, induced a marked and dose-dependent release of LDH. When PMNL were exposed to MMVF, some of the fibre types slightly increased the levels of free intracellular calcium ([Ca2+]i) within the cells in a manner similar to that induced by chrysotile or quartz. All MMVF induced a dose-dependent production of reactive oxygen species (ROS) in PMNL, with RCF-induced production of ROS being the most marked. Production of ROS by MMVF seemed to depend on the availability of extracellular calcium because it could be attenuated with a Ca2+ channel blocker, verapamil, or a Ca2+ chelating agent, EGTA. Production of ROS may be a common pathway through which PMNL respond to MMVF-induced cell activation, but alterations of levels of free intracellular Ca2+ do not seem to be an absolute prerequisite for this effect. Fibre length seemed not to be an important factor in affecting the ability of MMVF to induce ROS production in PMNL. However, the balance between different elements in the fibre seemed importantly to affect the biological activity of a fibre. PMID:10413242

Ruotsalainen, M; Hirvonen, M R; Luoto, K; Savolainen, K M



Effects of stilbenes isolated from medicinal plants on arachidonate metabolism and degranulation in human polymorphonuclear leukocytes  

Microsoft Academic Search

Studies were made on the effects of stilbene derivatives isolated from medicinal plants on arachidonate metabolism and degranulation in human polymorphonuclear leukocytes (PMN-L). Resveratrol (3,4?,5-trihydroxystilbene) isolated from the roots of Reynoutria japonica was found to inhibit the 5-lipoxygenase products 5-hydroxy-6,8,11,14-eicosatetraenoic acid (5-HETE), 5,12-dihydroxy-6,8,10,14-eicosatetraenoic acid (5,12-diHETE) and leukotriene C4(LTC4); its concentrations for 50% inhibition (IC50) were 8.90 × 10?6 M, 6.70

Yoshiyuki Kimura; Hiromichi Okuda; Michinori Kubo



Calcium oxalate crystal-induced cytolysis in polymorphonuclear leukocytes and erythrocytes.  


Calcium oxalate microcrystals induce cytolysis of rabbit polymorphonuclear leukocytes (PMN's) and hemolysis of human erythrocytes. The effects on erythrocytes can be distinguished from those on PMN's because cytolysis of the latter is suppressed by substances such as cytochalasin A and N-naphthyl maleimide, known phagocytosis inhibitors. Polyvinylpyridine-N-oxide, a powerful hydrogen acceptor, has no protective effect. Cell injury of PMN's and erythrocytes is potentiated in the presence of cations, whereas poly-D-glutamic acid and other negatively charged compounds have an opposite effect. The results suggest that positive charges on the crystals play an essential role in calcium oxalate-induced cytolysis of PMN's and erythrocytes. PMID:7211577

Elferink, J G; Riemersma, R C



Interaction between human polymorphonuclear leukocytes (PMNL) and bacteria cultivated in aerobic and anaerobic conditions.  


A study was performed on one strain of Escherichia coli (E. coli K12D22) and on one strain of Salmonella braenderup (S. braenderup S2828). The physico-chemical surface properties of the bacteria were strongly influenced by oxygen supply, viz. anaerobic growth conditions resulted in increasing of hydrophobicity. Interaction between human polymorphonuclear leukocytes and bacteria, measured as chemiluminescence, was more efficient when bacteria had been cultivated anaerobically than when cultivated aerobically. The results show the importance of the surface hydrophobicity of bacteria in interaction with PMNL, and the role of the growth conditions of bacteria in that process. PMID:3893034

Maluszynska, G M; Stendahl, O; Magnusson, K E



Activation of the lipoxygenase pathway in the methionine enkephalin induced respiratory burst in human polymorphonuclear leukocytes  

SciTech Connect

In comparative studies of f-met-Leu-Phe (FMLP) and methionine enkephalin (ME) induced polymorphonuclear leukocyte (PMNL) stimulation the following results were obtained: (i) both FMLP and ME increased the intracellular killing (IK) capability of human PMNLs probably through NADPH oxidase activation, (ii) the ME-induced respiratory burst (RB) differed from the chemotactic peptide FMLP-triggered superoxide generation because the former was not accompanied by the activation of the glutathione system and the duration of the superoxide production was prolonged. The reaction was dependent on lipoxygenation, was potentiated by indomethacin (IM) and was inhibited by nordihidro-guairetic acid (NDGA), (iii) both /sup 14/C-arachidonic acid release and leukotriene B/sub 4/ (LTB/sub 4/) synthesis of ME-treated PMNLs were elevated as compared to those of FMLP triggered cells. Their results suggest that lipoxygenation and even an increased LTB/sub 4/ synthesis are involved in the ME-induced RB of leukocytes.

Nagy, J.T.; Foris, G.; Fulop, T. Jr.; Paragh, G.; Plotnikoff, N.P.



Enzyme release from polymorphonuclear leukocytes during interaction with calcium oxalate microcrystals.  


The interaction between polymorphonuclear leukocytes and calcium oxalate crystals results in enzyme release from the cells, due to exocytosis and plasma membrane damage. Positive charges on the crystals apparently play a predominant role in the interaction. Removal of negatively charged sialic acid from the cell surface has little effect on crystal-induced enzyme release. Glucose-loaded liposomes release glucose upon exposure to calcium oxalate crystals when the liposomes are negatively charged but not when the liposomes bear a positive charge. Cytoplasts, which are devoid of granules, are severely damaged by calcium oxalate crystals. As with intact cells the damaging effect of the crystals on liposomes and cytoplasts can be counteracted by a polyanion. The results are consistent with the view that calcium oxalate crystals can cause enzyme release from polymorphonuclear leukocytes, mainly by a direct effect on the plasma membrane. The release might be due to an interaction of positive charges on the crystals with negative countercharges on the cells, possibly located in the phospholipid bilayer. PMID:2439709

Elferink, J G; Deierkauf, M



High-performance liquid chromatography measurement of antimicrobial concentrations in polymorphonuclear leukocytes.  

PubMed Central

High-performance liquid chromatography was used to determine the penetration of 19 antimicrobial agents into human polymorphonuclear leukocytes. The ratios of the intracellular concentration to the extracellular concentration of ampicillin, piperacillin, cefazolin, ceftizoxime, cefpimizole, and ceftazidime were all less than 0.6. Lincomycin showed a high intracellular-to-extracellular ratio (3.0), while clindamycin achieved a ratio of 15.5, which was the highest ratio of all of the 19 tested antibiotics. Ratios for rifampin, isoniazid, chloramphenicol, and trimethoprim were 8.2, 1.1, 9.6, and 6.1, respectively. Six quinolone-class antimicrobial agents had ratios from 2.2 to 8.2. Flucytosine showed a penetration ratio of 4.6. Clindamycin uptake was temperature dependent and occurred best with live polymorphonuclear leukocytes; sodium fluoride, adenosine, and puromycin were inhibitory. The results obtained in this study correlate well with the results of other studies involving radioisotopic methods. This indicates that high-performance liquid chromatography is a useful method for determining the intracellular penetration of antimicrobial agents.

Koga, H



Demonstration of X chromatin in drumstick-like nuclear appendages of leukocytes by in situ hybridization on blood smears  

Microsoft Academic Search

An X chromosome specific nucleic acid probe was used to study the positions of the X chromosomes in leukocyte nuclei by in situ hybridization to smears of peripheral blood. This autoradiographic approach allowed the first direct demonstration of the presence of X chromosomal material in the drumstick-like structures of female polymorphonuclear leukocytes.

P. F. R. Hochstenbach; J. M. J. C. Scheres; T. W. J. Hustinx; B. Wieringa



Polymorphonuclear Leukocyte Lysosomal Release in Response to Propionibacterium acne in Vitro and its Enhancement by Sera from Inflammatory Acne Patients  

Microsoft Academic Search

Propionibacteriunt acnes cells were tested for the ability to trigger lysosomal hydrolase release from human polymorphonuclear leukocytes. Representative strains of P. acnes serotype I and II failed to stimulate lysosomal release in the absence of serum. P. acnes growth culture supernatants failed to trigger release under any test condition. Addition of fresh or heat-inactivated human serum resulted in lysosomal hydrolase

G. F. Webster; J. J. Leyden; C. C. Tsai; P. Baehni; W. P. McArthur



Clinically Effective Monoclonal Antibody 3F8 Mediates Nonoxidative Lysis of Human Neuroectodermal Tumor Cells by Polymorphonuclear Leukocytes1  

Microsoft Academic Search

Most studies of antibody-dependent cellular cytotoxicity (ADCC) by polymorphonuclear leukocytes (PMN) have supported oxidative lytic processes. This may be because the studies used nonhuman or nonneo- plastic cells that were highly sensitive to reactive oxygen species or were small enough to be phagocytosed by PMN. We therefore investigated whether oxygen radicals participate in PMN cytotoxicity toward human neuroectodermal solid tumor

Brian H. Kushner; Nai-Kong V. Cheung


Accumulation of polymorphonuclear leukocytes in reperfused ischemic canine myocardium: relation with tissue viability assessed by fluorine-18-2-deoxyglucose uptake  

SciTech Connect

Polymorphonuclear leukocytes may participate in reperfusion injury. Whether leukocytes affect viable or only irreversibly injured tissue is not known. Therefore, we assessed the accumulation of 111In-labeled leukocytes in tissue samples characterized as either ischemic but viable or necrotic by metabolic, histochemical, and ultrastructural criteria. Six open-chest dogs received left anterior descending coronary occlusion for 2 hr followed by 4 hr reperfusion. Myocardial blood flow was determined by microspheres and autologous 111In-labeled leukocytes were injected intravenously. Fluorine-18-2-deoxyglucose, a tracer of exogenous glucose utilization, was injected 3 hr after reperfusion. The dogs were killed 4 hr after reperfusion. The risk and the necrotic regions were assessed following in vivo dye injection and postmortem tetrazolium staining. Myocardial samples were obtained in the ischemic but viable, necrotic and normal zones, and counted for 111In and 18F activity. Compared to normal, leukocytes were entrapped in necrotic regions (111In activity: 207 +/- 73%) where glucose uptake was decreased (26 +/- 15%). A persistent glucose uptake, marker of viability, was mainly seen in risk region (135 +/- 85%) where leukocytes accumulation was moderate in comparison to normal zone (146 +/- 44%). Thus, the glucose uptake observed in viable tissue is mainly related to myocytes metabolism and not to leukocytes metabolism.

Wijns, W.; Melin, J.A.; Leners, N.; Ferrant, A.; Keyeux, A.; Rahier, J.; Cogneau, M.; Michel, C.; Bol, A.; Robert, A.



Cationic Proteins of Polymorphonuclear Leukocyte Lysosomes II. Composition, Properties, and Mechanism of Antibacterial Action  

PubMed Central

Zeya, H. I. (University of North Carolina, Chapel Hill), and J. K. Spitznagel. Cationic proteins of polymorphonuclear leukocyte lysosomes. II. Composition, properties, and mechanism of antibacterial action. J. Bacteriol. 91:755–762. 1966.—A basic proteins fraction from guinea pig polymorphonuclear (PMN) granules was obtained by acid extraction and precipitation with 20% (v/v) ethyl alcohol. The fraction accounted for most of the antibacterial activity of the PMN granules and corresponded to the antibacterial cationic components of intact granules (bands I, II, and III) resolved by zone electrophoresis. Absence from the fraction of components identical to the enzymatic components of intact lysosomes showed that the fraction was essentially free from lysosomal enzymes. The amino acid analysis of proteins in the fraction gave a preponderance of basic amino acids (25%), especially of arginine (16%). The comparative amino acid analysis showed that the lysosomal cationic proteins (LCP) fraction was markedly different from nuclear histones. The LCP fraction manifested antibacterial activity against certain gram-positive and gram-positive microorganisms, including Candida albicans, and exhibited stoichiometric relationship in its activity. Microorganisms treated with LCP fraction were agglutinated. Anionic substances such as nucleic acids, heparin, and endotoxin effectively blocked the antibacterial activity of the fraction. The LCP fraction caused suppression of oxygen uptake by bacterial cells and damaged the permeability barriers of cells as manifested by rapid release of P32 as well as ultraviolet-absorbing material at 260 m?, in the supernatant fluid. Images

Zeya, H. I.; Spitznagel, J. K.



Peripheral polymorphonuclear leukocyte activation as a systemic inflammatory response in ischemic stroke.  


Stroke is one of the leading causes of mortality and morbidity in the world. The activation of polymorphonuclear leukocytes (PMNL) plays an important role in the inflammatory response after ischemic stroke. However, in the current literature, there are few studies discussing the process and role of peripheral PMNL activation. Here, we give a comprehensive description of peripheral PMNL activation after ischemic stroke and discuss their potential roles in the process of ischemic injury. Based on our analysis, peripheral PMNL activation is supposed to be attributed to systemic inflammatory response to cerebral ischemic insult, not reflecting the activity of PMNL in local ischemic brain. Inhibiting peripheral PMNL activation in stroke animals has been effective in reducing infarction and improving behavioral outcome; thus, the same approach of inhibiting peripheral PMNL activation is a promising therapeutic strategy for stroke patients. PMID:23619532

Mo, Xiaoye; Li, Ting; Ji, Guang; Lu, Wei; Hu, Zhiping



Chemiluminescent and flow cytometric analysis of gamma interferon preincubation on neonatal and adult rat polymorphonuclear leukocytes.  

PubMed Central

Gamma interferon (IFN-gamma) has multiple immunomodulating effects and has been postulated as a possible immunopotentiating agent for the prevention or treatment of neonatal infections. This report describes the effect of rat recombinant IFN-gamma on the oxidative burst activity and CD11b expression of neonatal and adult rat polymorphonuclear leukocytes (PMNL). Oxidative burst activity was assessed by chemiluminescence and dihydrorhodamine flow cytometry. Neonatal PMNL exhibited significantly less oxidative burst activity than did adult PMNL. IFN-gamma mildly enhanced the chemiluminescence response of PMNL from both the rat pups and adults, but this effect was not statistically significant when analyzed by a multivariate model of repeated-measures analysis of variance for both chemiluminescence and dihydrorhodamine flow cytometry. CD11b expression was also not significantly enhanced by IFN-gamma.

Wittler, R R; Lieberman, M M; Paine, D D; Muehlbauer, S L; Lima, J E; Sachanandani, D M; Pinney, C A



Impairment of human polymorphonuclear leukocyte chemotaxis by 2,5-hexanedione.  


The effect of n-hexane metabolites on human polymorphonuclear leukocyte chemotaxis and luminol-dependent chemiluminescence was investigated. No effect was detected when 2-hexanol, 2-hexanone and gamma-valerolactone were used; 2,5-hexanedione at 75 micrograms/ml inhibited chemotaxis and a direct correlation between increasing the xenobiotic concentration and the degree of inhibition was found. Chemotactic peptide-induced chemiluminescence was not affected by 2,5-hexanedione. In order to clarify the phenomenon, plasma membrane fluidity was investigated by fluorescence polarization of the fluorescent probe trimethylammonium diphenylhexatriene. 2,5-hexanedione increased the membrane fluidity, while the other n-hexane metabolites did not change the degree of fluorescence polarization. Results suggest that the cellular functions modulated by membrane-cytoskeletal organization are affected by 2,5-hexanedione also at the low concentrations. PMID:3267444

Governa, M; Valentino, M; Visona, I; Rocco, M



Pulmonary accumulation of polymorphonuclear leukocytes in the adult respiratory distress syndrome  

SciTech Connect

The polymorphonuclear leukocyte (PMN) plays an integral role in the development of permeability pulmonary edema associated with the adult respiratory distress syndrome (ARDS). This report describes 3 patients with ARDS secondary to systemic sepsis who demonstrated an abnormal diffuse accumulation of Indium (/sup 111/In)-labeled PMNs in their lungs, without concomitant clinical or laboratory evidence of a primary chest infection. In one patient, the accumulation of the pulmonary activity during an initial pass suggested that this observation was related to diffuse leukoaggregation within the pulmonary microvasculature. A 4th patient with ARDS was on high-dose corticosteroids at the time of a similar study, and showed no pulmonary accumulation of PMNs, suggesting a possible reason for the reported beneficial effect of corticosteroids in human ARDS.

Powe, J.E.; Short, A.; Sibbald, W.J.; Driedger, A.A.



Mechanisms of lectin and antibody-dependent polymorphonuclear leukocyte-mediated cytolysis.  


The mechanisms of tumor lysis by polymorphonuclear leukocytes (PMNs) were investigated. In antibody-dependent PMN-mediated cytolysis (ADPC), sensitized tumor cells were specifically lysed via Fc receptors on PMNs. On the other hand, lectin-dependent PMN-mediated cytolysis (LDPC) caused nonspecific lysis of several murine tumors after recognition of carbohydrate moieties on the cell membrane of both PMNs and tumor cells. Both ADPC and LDPC depended on glycolysis, and cytotoxicity was mediated by reactive oxygen species; LDPC was dependent on superoxide and ADPC on the myeloperoxidase system. The participation of reactive oxygen species in PMN cytotoxicity was also demonstrated by pharmacological triggering with phorbol myristate acetate. These results indicate that reactive oxygen species have an important role In tumor killing by PMNs and that ADPC and LDPC have partly different cytolytic processes as well as different recognition steps. PMID:6862147

Tsunawaki, S; Ikenami, M; Mizuno, D; Yamazaki, M



Determination of phagocytosis of /sup 32/P-labeled Staphylococcus aureus by bovine polymorphonuclear leukocytes  

SciTech Connect

A procedure for the measurement of phagocytosis by bovine polymorphonuclear leukocytes (PMN) of /sup 32/P-labeled Staphylococcus aureus was modified so that a larger number of samples could be compared in a single run, and smaller volumes of sample, PMN, and /sup 32/P-labeled S aureus could be used. Results were highly reproducible, with a coefficient of variation between duplicate determinations of less than or equal to 2%. Lysostaphin was prepared from the supernatant of S staphylolyticus and was compared with a commercially available preparation. Effects of lysostaphin on PMN and influence of incubation media on release of /sup 32/P from /sup 32/P-labeled S aureus by lysostaphin were examined.

Dulin, A.M.; Paape, M.J.; Weinland, B.T.



Cytolytically inactive terminal complement complex causes transendothelial migration of polymorphonuclear leukocytes in vitro and in vivo.  


Intravital microscopy was used to monitor leukocyte traffic across rat mesenteric postcapillary venules induced by the inactive terminal complement (C) complex (iTCC) topically applied to ileal mesentery. Leukocytes started rolling within 15 minutes from the administration of iTCC, and by 1 hour they adhered almost completely to the endothelium emigrating from the vessels in the next 3 hours. C5a caused a similar, though less marked, effect, whereas boiled iTCC was inactive, excluding the contribution of contaminating lipopolysaccharide. The complex stimulated the migration of polymorphonuclear neutrophils (PMNs) across endothelial cells (ECs) in a transwell system after a 4-hour incubation of ECs with iTCC added to the lower chamber of the transwell, whereas a 30-minute incubation was sufficient for C5a and interleukin (IL)-8 to induce the passage of PMNs. C5a was not responsible for the effect of iTCC because this complex had no chemotactic activity and contained too small an amount of C5a to account for the transendothelial migration of PMNs. Similarly, the effect of iTCC was not mediated by IL-8 released by stimulated ECs because anti-IL-8 failed to inhibit the migration of PMNs induced by the complex. Unlike tumor necrosis factor-alpha, iTCC did not cause the redistribution of platelet-endothelial cell adhesion molecule-1 (PECAM-1), and PMN mobilization was partially blocked by anti-PECAM-1 antibodies. PMID:11756170

Dobrina, Aldo; Pausa, Mario; Fischetti, Fabio; Bulla, Roberta; Vecile, Elena; Ferrero, Elisabetta; Mantovani, Alberto; Tedesco, Francesco



Potent and selective inhibitors of leukotriene A4 hydrolase: effects on purified enzyme and human polymorphonuclear leukocytes.  


Leukotriene (LT) A4 hydrolase (EC is a bifunctional zinc metalloenzyme that catalyzes the hydrolysis of the unstable epoxide intermediate LTA4 into the proinflammatory substance LTB4 and also exhibits an amidase/peptidase activity toward synthetic substrates. Based on proposed reaction mechanisms for other zinc hydrolases, we have synthesized inhibitors of LTA4 hydrolase and evaluated their effects on the formation of LTB4 from LTA4 using both purified enzyme and intact polymorphonuclear leukocytes. The two most effective inhibitors, an alpha-keto-beta-amino ester (compound IV) and a thioamine (compound VIII), exhibited IC50 values of 1.9 +/- 0.9 and 0.19 +/- 0.12 microM (mean +/- SD, n = 4), respectively. Compounds IV and VIII were also potent inhibitors of LTB4 biosynthesis in ionophore stimulated polymorphonuclear leukocytes with IC50 < 200 nM. At higher concentrations, the biosynthesis of 5-hydroxy-eicosatetraenoic acid was also inhibited with IC50 approximately 10 microM for both substances. In contrast, leukocyte 15-lipoxygenase and platelet LTC4 synthase activity were not inhibited by these substances at the highest concentrations tested, 50 and 10 microM, respectively. Compounds IV and VIII thus exhibit selectivity among enzyme activities in the arachidonic acid cascade. In conclusion, we describe two compounds that are among the most potent and selective inhibitors of LTA4 hydrolase and LTB4 biosynthesis by intact polymorphonuclear leukocytes, described thus far. PMID:7562564

Wetterholm, A; Haeggström, J Z; Samuelsson, B; Yuan, W; Munoz, B; Wong, C H



Oxidative cross-linking of immune complexes by human polymorphonuclear leukocytes.  

PubMed Central

Incubation of human serum albumin-anti-human serum albumin immune complexes bound to a plastic surface, with human polymorphonuclear leukocytes for 1 h at 37 degrees C resulted in covalent cross-linking of 8.5% +/- 0.5 of the complexes, corresponding to a minimum rate of 700 antibody molecules per cell per minute. Similar results were obtained with IgG-anti-IgG and type II collagen-anticollagen II human antibodies. Cross-linking was defined as the antibody remaining attached to plastic-bound antigen after extraction with 3 M MgCl2 and 0.1 N HCl solutions. The effects of addition of oxygen radical scavengers, heme-enzyme inhibitors, and omission of Cl- indicated that the cross-linking process was mediated by the myeloperoxidase-H2O2-Cl- system. Cross-linking was also obtained with cell lysates, polymorphonuclear granules, and purified human myeloperoxidase in the presence of a steady flux of H2O2 provided by glucose oxidase-glucose. Cross-linking by the cell-free systems was also abolished by sodium azide or omission of chloride ions. Cross-linked immune complexes were also generated by incubation with 20 to 50 microM solutions of freshly distilled hypochlorous acid. Addition of 10 mM hypochlorous acid to soluble IgG resulted in the formation of protein precipitates insoluble in 5 M guanidine, 0.1 N HCl, or boiling 2.3 M sodium dodecyl sulfate-1.4 M 2-mercaptoethanol. The remaining soluble IgG contained fluorescent high molecular aggregates (ex: 360 nm; em: 454 nm). Oxidative cross-linking of antigen-antibody molecules, and of immune complexes to connective tissue macromolecules may play a pathogenic role in acute and chronic inflammatory processes.

Jasin, H E



Suppression of natural killing in vitro by monocytes and polymorphonuclear leukocytes: requirement for reactive metabolites of oxygen.  

PubMed Central

Natural killer cells spontaneously lyse certain tumor cells and may defend against malignancy. We have previously shown that natural killing (NK) by human peripheral blood mononuclear cells (PBMC) is suppressed in vitro by phorbol diester tumor promoters, including 12-O-tetradecanoylphorbol-13-acetate (TPA). We here demonstrate that suppression of NK is mediated by monocytes or polymorphonuclear leukocytes (PMN) and that suppression is dependent on the generation of reactive forms of molecular oxygen (RO), particularly hydrogen peroxide (H2O2). NK was suppressed not only by TPA but also by opsonized zymosan (yeast cell walls), which, like TPA, was not toxic to PBMC. Both TPA and zymosan stimulated the production of superoxide anion (O2-) and H2O2 by PBMC. Production of RO correlated with suppression of NK. When PBMC were depleted of monocytes, the production of RO and the suppression of NK were both markedly reduced. Suppression could be restored by monocytes or PMN, both of which produced RO in response to TPA or zymosan. Suppression of NK was dependent on RO. Monocytes or PMN from a patient with chronic granulomatous disease, whose cells cannot generate RO, did not mediate suppression of NK. Suppression was also reduced in glucose-free medium, which did not support the generation of RO. Suppression of NK by TPA was inhibited by catalase. Bovine superoxide dismutase had a limited effect on suppression, even in high concentration, and tyrosine-copper (II) complex, which also enhances dismutation of O2- to H2O2, had almost no effect on suppression. When H2O2 was directly generated enzymatically from glucose oxidase and glucose, NK was suppressed and suppression was reversed by catalase. NK was also suppressed by the enzymatic generation of O2- from xanthine oxidase and xanthine, but suppression under these conditions was again inhibited by catalase and not by superoxide dismutase, indicating that suppression was due to the secondary formation of H2O2 from O2-. These results indicate that H2O2 is important in suppression of NK. Myeloperoxidase did not appear to play a role in suppression because inhibition of this enzyme by sodium azide, cyanide, or aminotriazole did not prevent suppression of NK. Suppression of NK was reversible; after exposure to zymosan, NK could be partially restored by the addition of catalase and superoxide dismutase or by the removal of zymosan. These studies demonstrate cellular regulation of NK by monocytes or polymorphonuclear leukocytes and indicate a role for RO in immunoregulation.

Seaman, W E; Gindhart, T D; Blackman, M A; Dalal, B; Talal, N; Werb, Z



Stimulatory effect of some plant extracts used in homeopathy on the phagocytosis induced chemiluminescence of polymorphonuclear leukocytes.  


Some plant extracts on a large range of dilutions as used in Homeopathy were tested on the chemiluminescence emission produced by polymorphonuclear leukocytes. The high stimulatory action was noticed when extracts from Uvae Ursi and Saponaria were tested, as the classical effect exerted by zymosan was exceeded. A moderate stimulatory action comparable with that of zymosan was found when extracts from Echmaceea, Aleo and Prumis were used, as well as in the case of Propolis. The relationship between stimulatory effect and the concentration range is modulated as function of the extract source, several peaks being observed for some dilutions (Saponana), but generally no quantitative relations were obtained. By studying the time when a chemiluminescence peak was observed, it is possible to estimate wether the weight of the NADPH oxidase or myeloperoxidase pathways are involved in the stimulatory effect on polymorphonuclear leukocytes. PMID:11712436

Crocnan, D O; Greabu, M; Olinescu, R



Antifungal Triazoles and Polymorphonuclear Leukocytes Synergize To Cause Increased Hyphal Damage to Scedosporium prolificans and Scedosporium apiospermum  

Microsoft Academic Search

Scedosporium prolificans and Scedosporium apiospermum (Pseudallescheria boydii) cause pulmonary and dis- seminated infections refractory to most currently used antifungal agents in immunocompromised patients. We therefore investigated the potential antifungal activities of the triazoles itraconazole (ITC), voriconazole (VRC), and posaconazole (PSC) in combination with human polymorphonuclear leukocytes (PMNs) against the hyphae of these fungal pathogens. A colorimetric assay with (2,3-bis(2-methoxy-4-nitro-5-sulfophenyl)2H- tetrazolium-5-carboxanilide)

Cristina Gil-Lamaignere; Emmanuel Roilides; Juan Mosquera; Avgi Maloukou; Thomas J. Walsh



Biochemical and Functional Characterization of MCS2 Antigen (CD13) on Myeloid Leukemic Cells and Polymorphonuclear Leukocytes1  

Microsoft Academic Search

The antigen defined by MCS-2 monoclonal antibody (niAh) was char acterized by sodium dodecyl sulfate-polyacrylamide gel electrophoresis of surface and internally labeled cells. Surface iodination revealed that MCS-2 antigen on the surfaces of acute myelogenous leukemia cells, III 61)cells, and polymorphonuclear leukocytes (PMN) had the same molecular weight (.Vf, 150,000) and that their autoradiographic bands were also the same. Internal

Kenji Sakai; Toshio Hattori; Kimitaka Sagawa; M Usuo Yokoyama; Kiyoshi Takatsuki


Motility and oxidative metabolism of normal polymorphonuclear leukocytes in acute non-lymphoid leukemia in complete remission  

Microsoft Academic Search

Summary  Polymorphonuclear leukocytes (PMN) from 11 patients with acute non-lymphoid leukemia (ANLL) in complete remission and off\\u000a therapy for at least 22 months were studied for their oxidative metabolism and motility. In all patients, PMN motility tests\\u000a revealed no consistent abnormality, whereas cells from 2 patients showed a selective impairment of oxidative metabolism, as\\u000a assessed by the low capacity to reduce

Vincenzo Bottari; Andrea Fattorossi; Roberto Nisini; Maria Concetta Petti; Susanna Fenu; Luigi Fontana



Significance of Leukocytes in Endotoxin Shock.  

National Technical Information Service (NTIS)

Peripheral blood leukopenia and the sequestration of large numbers of polymorphonuclear leukocytes in pulmonary capillaries have been observed in experimental shock. It has been suggested that release of leukocytic lysosomal enzymes contributes to systemi...

W. W. Pingleton J. J. Coalson C. A. Guenter



Simultaneous measurement of phagocytosis and respiratory burst of leukocytes in whole blood from bottlenose dolphins ( Tursiops truncatus) utilizing flow cytometry  

Microsoft Academic Search

Phagocytic and respiratory burst activity was simultaneously measured by flow cytometry in polymorphonuclear leukocytes (PMN) and monocytes in whole blood from bottlenose dolphins (Tursiops truncatus). Blood was collected from 16 adult dolphins, 12 males (6–34 years of age) and 4 females (11–30 years) and subsequently incubated with a bacteria-to-leukocyte ratio of 25:1 and 10?l of 500?M 2?,7?-dichlorofluorescein diacetate for 70min

M. J. Keogh; T. Spoon; S. H. Ridgway; E. Jensen; W. Van Bonn; T. A. Romano


Intracellular penetration and activity of BAY Y 3118 in human polymorphonuclear leukocytes.  

PubMed Central

The penetration of a new quinolone (BAY Y 3118) into human polymorphonuclear leukocytes (PMNs) was evaluated by a fluorometric assay. The cellular concentration-to-extracellular concentration (C/E) ratio was higher than 6.3 at extracellular concentrations ranging from 2 to 100 mg/liter. The uptake of BAY Y 3118 was rapid, reversible and nonsaturable. The intracellular penetration of BAY Y 3118 was significantly affected by environmental temperature (C/E ratio at 4 degrees C, 5.4 +/- 0.5; control, 7.5 +/- 0.9; P < 0.05) and cell viability (C/E ratio in dead PMNs, 5.5 +/- 0.8; control 7.5 +/- 0.9; P < 0.05), but it was not affected by metabolic inhibitors. The ingestion of opsonized zymosan or opsonized Staphylococcus aureus significantly decreased the levels of PMN-associated BAY Y 3118. Cell stimulation by a membrane activator, however, significantly increased the intracellular concentration of this quinolone. At therapeutic extracellular concentrations (0.5, 2, and 5 mg/liter), BAY Y 3118 showed intracellular activity greater than that of ciprofloxacin against S. aureus in human PMNs. It was concluded that BAY Y 3118 reaches high intracellular concentrations within human PMNs and remains active intracellularly.

Garcia, I; Pascual, A; Perea, E J



Effects of adenosine on the functions of circulating polymorphonuclear leukocytes during hyperdynamic endotoxemia.  

PubMed Central

Endotoxin-activated polymorphonuclear leukocytes (PMNL) adhere to the vascular endothelium and cause damage by the release of toxic superoxide anions (O2-). Because adenosine is a potent inhibitor of PMNL in vitro, the present study investigates the effects of this nucleoside on the functions of circulating PMNL in a standardized porcine model of hyperdynamic endotoxemia. Ten anesthesized pigs received an intravenous (i.v.) 330-min infusion of endotoxin (5 microg/kg of body weight per h). Another 10 pigs were also infused with endotoxin plus adenosine (150 microg/kg/min [i.v.]); this treatment was begun 30 min prior to the beginning of endotoxin treatment. Control groups (five animals per group) received either adenosine or physiological saline. Infusion of endotoxin caused severe neutropenia, shedding of L-selectin, upregulation of beta2-integrins, increased binding of C3-coated zymosan particles, and subsequent phagocytosis by PMNL. While phagocytosis-induced production of oxygen radicals appeared to decrease, extracellular release of superoxide anions was strongly enhanced. Infusion of adenosine during endotoxemia had no effect on neutropenia, expression of adhesion molecules, C3-induced adhesion, phagocytosis, or intracellular production of oxygen radicals, whereas extracellular release of O2- was strongly inhibited. Thus, i.v. infusion of adenosine during endotoxemia could be useful in protecting from O2(-)-mediated tissue injury without compromising the bactericidal mechanisms of PMNL.

Thiel, M; Holzer, K; Kreimeier, U; Moritz, S; Peter, K; Messmer, K



Yersinia pestis Type III Secretion System-Dependent Inhibition of Human Polymorphonuclear Leukocyte Function?  

PubMed Central

Human polymorphonuclear leukocytes (PMNs, or neutrophils) are the primary innate host defense against invading bacterial pathogens. Neutrophils are rapidly recruited to sites of infection and ingest microorganisms through a process known as phagocytosis. Following phagocytosis by human PMNs, microorganisms are killed by reactive oxygen species (ROS) and microbicidal products contained within granules. Yersinia pestis, the causative agent of plague, is capable of rapid replication and dissemination from sites of infection in the host. Although Y. pestis survives in macrophages, the bacterial fate following interaction with human PMNs is less clear. The ability of Y. pestis to inhibit phagocytosis by human PMNs was assessed by differential fluorescence microscopy and was shown to be dependent on expression of the type III secretion system (TTSS). Previous studies have demonstrated that TTSS expression in enteropathogenic Yersinia spp. also inhibits the respiratory burst in PMNs and macrophages, and we show here that human PMN ROS production is similarly repressed by Y. pestis. However, exclusion of uningested TTSS-expressing Y. pestis with gentamicin revealed that intracellular bacteria are eliminated by human PMNs, similar to bacteria lacking the TTSS. In summary, our results suggest that the Y. pestis TTSS contributes to extracellular survival following interactions with human PMNs and that the intracellular fate is independent of TTSS inhibition of neutrophil ROS production.

Spinner, Justin L.; Cundiff, Jennifer A.; Kobayashi, Scott D.



Phorbol esters cause sequential activation and deactivation of complement receptors on polymorphonuclear leukocytes  

SciTech Connect

Phorbol myristate acetate (PMA) exerts a biphasic effect on receptors for C3b and C3bi of human polymorphonuclear leukocytes (PMN). The addition of PMA for 10 min enhances the capacity of these receptors to promote binding and phagocytosis of C3b- and C3bi-coated erythrocytes. Upon additional incubation for 60 min, the capacity of these receptors to bind and ingest ligand-coated erythrocytes decreases the levels below those of resting cells. Although PMA does cause increased expression of cell surface C3b and C3bi receptors, the sequential rise and fall of receptor activity cannot be accounted for the alterations in the number of surface receptors. It appears, rather, that PMA causes qualitative changes in these receptors, first an increase in receptor activity (activation) and then a decrease in receptor activity. To explore the role of phosphorylation in the sequential activation and deactivation of phagocytosis-promoting receptors, the authors loaded PMN with thiophosphate (thioP). This compound is incorporated into cellular nucleotides and proteins, and the resultant (thio)phosphorylated proteins are resistant to phosphatases. thioP-loaded cells show enhanced receptor activity, suggesting that activation of receptors is mediated by a phosphorylation event. Cells loaded with thioP and treated with PMA for 70 min do not deactivate C3 or Fc reactors, suggesting that the deactivation is the result of a dephosphorylation event.

Wright, S.D.; Meyer, B.C.



Role of YopK in Yersinia pseudotuberculosis Resistance against Polymorphonuclear Leukocyte Defense  

PubMed Central

The enteropathogen Yersinia pseudotuberculosis can survive in the harsh environment of lymphoid compartments that abounds in immune cells. This capacity is dependent on the plasmid-encoded Yersinia outer proteins (Yops) that are delivered into the host cell via a mechanism involving the Yersinia type III secretion system. We show that the virulence protein YopK has a role in the mechanism by which Y. pseudotuberculosis avoids the polymorphonuclear leukocyte or neutrophil (PMN) defense. A yopK mutant, which is attenuated in the mouse infection model, where it fails to cause systemic infection, was found to colonize Peyer's patches and mesenteric lymph nodes more rapidly than the wild-type strain. Further, in mice lacking PMNs, the yopK mutant caused full disease with systemic spread and typical symptoms. Analyses of effects on PMNs revealed that both the wild-type strain and the yopK mutant inhibited internalization and reactive oxygen species production, as well as neutrophil extracellular trap formation by PMNs. However, the wild-type strain effectively avoided induction of PMN death, whereas the mutant caused a necrosis-like PMN death. Taken together, our results indicate that YopK is required for the ability of Yersinia to resist the PMN defense, which is critical for the virulence of the pathogen. We suggest a mechanism whereby YopK functions to prevent unintended Yop delivery and thereby PMN disruption, resulting in necrosis-like cell death, which would enhance the inflammatory response favoring the host.

Thorslund, Sara E.; Ermert, David; Fahlgren, Anna; Erttmann, Saskia F.; Nilsson, Kristina; Hosseinzadeh, Ava; Urban, Constantin F.



Auranofin inhibits the activation pathways of polymorphonuclear leukocytes at multiple sites.  


In order to characterize the mechanism by which the anti-rheumatic gold complex auranofin (AF) affects the functions of resting and activated polymorphonuclear leukocytes (PMN) the following studies were performed: (1) The effect of AF on the major processes involved in the respiratory burst of PMN: glucose transport and phosphorylation; hexose monophosphate (HMP) shunt activity in intact cells and in a cell-free system; superoxide production by particulate fractions and intact PMN measured as lucigenin-dependent chemiluminescence. (2) A comparison of the effects of AF added to the PMN before, at the time of, or subsequent to the stimulants [N-formyl-methionyl-leucyl phenylalanine (FMLP), concanavalin A (ConA), calcium ionophore (A23187) and phorbol myristate acetate (PMA)]. (3) The effect of AF on PMN activated by two stimulates (PMA, ConA) added sequentially. AF (0.1-10 microM) caused a dose-dependent inhibition of lucigenin-dependent chemiluminescence regardless of the activator (FMLP, ConA, A23187, PMA) when AF was added before the activator. In contrast, when AF was added to PMN after stimulation, it inhibited only the chemiluminescence of PMN stimulated by PMA. Furthermore, the chemiluminescence was largely unaffected by AF in sequentially activated PMN. The relative sensitivity to AF of the various processes studied indicates that blockade of the activation signal appears to be responsible for inhibition of the respiratory burst of PMN. PMID:1645553

Rudkowski, R; Ziegler, J B; Graham, G G; Champion, G D



Role of YopK in Yersinia pseudotuberculosis resistance against polymorphonuclear leukocyte defense.  


The enteropathogen Yersinia pseudotuberculosis can survive in the harsh environment of lymphoid compartments that abounds in immune cells. This capacity is dependent on the plasmid-encoded Yersinia outer proteins (Yops) that are delivered into the host cell via a mechanism involving the Yersinia type III secretion system. We show that the virulence protein YopK has a role in the mechanism by which Y. pseudotuberculosis avoids the polymorphonuclear leukocyte or neutrophil (PMN) defense. A yopK mutant, which is attenuated in the mouse infection model, where it fails to cause systemic infection, was found to colonize Peyer's patches and mesenteric lymph nodes more rapidly than the wild-type strain. Further, in mice lacking PMNs, the yopK mutant caused full disease with systemic spread and typical symptoms. Analyses of effects on PMNs revealed that both the wild-type strain and the yopK mutant inhibited internalization and reactive oxygen species production, as well as neutrophil extracellular trap formation by PMNs. However, the wild-type strain effectively avoided induction of PMN death, whereas the mutant caused a necrosis-like PMN death. Taken together, our results indicate that YopK is required for the ability of Yersinia to resist the PMN defense, which is critical for the virulence of the pathogen. We suggest a mechanism whereby YopK functions to prevent unintended Yop delivery and thereby PMN disruption, resulting in necrosis-like cell death, which would enhance the inflammatory response favoring the host. PMID:23090955

Thorslund, Sara E; Ermert, David; Fahlgren, Anna; Erttmann, Saskia F; Nilsson, Kristina; Hosseinzadeh, Ava; Urban, Constantin F; Fällman, Maria



Lactoferrin: its role as a Ga-67-binding protein in polymorphonuclear leukocytes  

SciTech Connect

Gallium-67 bound to lactoferrin - an iron-binding protein found in high concentration in polymorphonuclear leukocytes - has been isolated from PMNs that have previously been incubated with Ga-67 citrate. Although the cell-labeling efficiency was highly variable (0.026-10%), much of the activity that did bind to the PMNs (74.8 +- 10%) was recovered in the supernatant after sonication and centrifugation. About half (approx. 47%) of the PMN-bound activity was retained after dialysis and was presumably bound to macromolecules in the supernatant. When this retained activity was placed on a column containing immobilized antilactoferrin antibody, almost three quarters of the activity was bound to the column. This bound activity was (36 +- 17%) of the total activity absorbed by the PMN. The addition to the antilactoferrin column of a known antigen-antibody-dissociating agent caused the dissolution of the complex. No significant activity was bound to a control column. The findings indicate that lactoferrin is a major Ga-67-binding protein present in PMNs and suggest that it may play a major role in Ga-67 localization in an abscess. These results support the contention that molecules binding ferric iron have an important effect on Ga-67 distribution in vivo.

Weiner, R.; Hoffer, P.B.; Thakur, M.L.



Neisseria gonorrhoeae-mediated inhibition of apoptotic signalling in polymorphonuclear leukocytes.  


The human pathogen Neisseria gonorrhoeae recruits and interacts extensively with polymorphonuclear leukocytes (PMNs) during infection. N. gonorrhoeae is able to survive the bactericidal activity of these innate immune cells and can actively modulate PMN functions in vitro. PMNs are short-lived cells which readily undergo apoptosis, and thus the effect of N. gonorrhoeae infection on PMN survival has implications for whether PMNs might serve as an important site of bacterial replication during infection. We developed and validated an HL-60 myeloid leukemia cell culture model for PMN infection and used both these cells and primary PMNs to show that N. gonorrhoeae infection alone does not induce apoptosis and furthermore that N. gonorrhoeae can inhibit both spontaneous apoptosis and apoptosis induced by the intrinsic and extrinsic apoptosis inducers staurosporine (STS) and tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL), respectively. N. gonorrhoeae infection also results in the activation of NF-?B signaling in neutrophils and induces secretion of an identical profile of proinflammatory cytokines and chemokines in both HL-60 cells and primary PMNs. Our data show that the HL-60 cell line can be used to effectively model N. gonorrhoeae-PMN interactions and that N. gonorrhoeae actively inhibits apoptosis induced by multiple stimuli to prolong PMN survival and potentially facilitate bacterial survival, replication, and transmission. PMID:21844239

Chen, Adrienne; Seifert, H Steven



Increased activity of 5-lipoxygenase in polymorphonuclear leukocytes from asthmatic patients  

SciTech Connect

The formation of 5-lipoxygenase products of arachidonic acid, 5-HETE and 5,12-diHETE, was determined in 100,000 x g supernatant of polymorphonuclear leukocytes from 17 healthy subjects, 17 patients with extrinsic asthma and 15 patients with intrinsic asthma. After the supernatant was incubated with /sup 14/C-arachidonic acid in the presence of calcium and indomethacin, the lipoxygenase products of arachidonic acid were separated by thin layer chromatography. The results were expressed as the percentage conversion of /sup 14/C-arachidonic acid into the product per 10/sup 7/ cells. The formation of 5,12-diHETE, but not of the 5-HETE, was significantly increased in the cells from the group of patients with extrinsic asthma (4.38 +/- 0.78%, mean +/- S.E.; p < 0.01) and intrinsic asthma (6.09 +/- 1.11%; p < 0.01), when compared to normal subjects (1.74 +/- 0.30%). Both extrinsic and intrinsic asthmatics had significantly enhanced 5-lipoxygenase activity, which was expressed as the sum of percentage conversion of /sup 14/C-arachidonic acid into 5-HETE and 5,12-diHETE. The percentage conversion in normal subjects was 4.19 +/- 0.39%, 6.24 +/- 0.84% for 17 patients with extrinsic asthma (p < 0.05), and 8.59 +/- 1.29% for 15 patients with intrinsic asthma (p < 0.01). There was no significant difference between these asthmatic groups. These results indicate that 5-lipoxygenase activity is increased in patients with bronchial asthma. 22 references, 3 figures.

Mita, H.; Yui, Y.; Taniguchi, N.; Yasueda, H.; Shida, T.



Effect of Fluconazole on Phagocytic Response of Polymorphonuclear Leukocytes in a Rat Model of Acute Sepsis  

PubMed Central

Recently, fluconazole (FLZ) has been shown to improve survival and reduce multiorgan failure in experimental and clinical septic shock. The mechanism by which FLZ affords protection against sepsis remains obscure. This study examines the effect of FLZ on phagocytic activity of polymorphonuclear leukocytes (PMNs) in a rat model of septic shock by inducing fecal peritonitis in male Wistar rats using intraperitoneal instillation (1?mL/kg) of fecal suspension in saline (1:1?w/v). Sham control rats received sterile fecal suspension and vehicle treatment. FLZ was administered in the doses of 0, 3, 10, and 30?mg/kg by gavage 30?minutes before fecal instillation. The samples of peritoneal fluid were collected 8?hours following fecal inoculation for the evaluation of phagocytic response of PMNs using zymosan-induced luminol-dependent chemiluminescence (CL). Fecal peritonitis caused massive infiltration of PMNs in the peritoneal cavity (ANOVA F4.45 = 6.322, P < .001). Although FLZ reduced the infiltration of PMNs, this effect was neither significant nor dose dependent. The actual CL response was significantly higher in the peritoneal fluid of rats subjected to peritonitis, which was significantly and dose-dependently attenuated by FLZ treatment (ANOVA F4.45 = 11.048, P < .001). Normalization of CL response for 1000?PMNs revealed that FLZ dose-dependently albeit insignificantly reduced the activity of PMNs. The high dose of FLZ caused 2.29-fold decrement in the area under curve (AUC) pertaining to cumulative CL response. The findings of this study suggest that FLZ protects rats against septic shock by inhibiting PMN-mediated inflammatory cascade without compromising their phagocytic activity.

Khan, Haseeb Ahmad



Possible role of calmodulin in the control of lysosomal enzyme release from human polymorphonuclear leukocytes.  


Human polymorphonuclear leukocytes (PMNs) were found to contain a mean +/- S.E.M. of 21.7 +/- 7.9 ng of immunoreactive calmodulin (CaM)/10(6) PMNs, which represents 0.032 +/- 0.001% of the total cellular protein. The functional role of CaM in the control of lysosomal enzyme release from human PMNs was investigated using several CaM antagonists. Trifluoperazine (TFP) (10(-6)-2 X 10(-5) M), pimozide (10(-6)-1.5 X 10(-5) M), chlorpromazine (CPZ) (10(-5)-10(-4) M) and promethazine (2 X 10(-5)-10(-4) M) inhibited in vitro lysosomal enzyme release from human PMNs induced by immunological (serum-treated zymosan, concanavalin A and formyl-L-methionyl-L-leucyl-L-phenylalanine) and nonimmunological (Ca++ ionophore A23187) stimuli. Trifluoperazine sulfoxide (TFP-S) and chlorpromazine sulfoxide (CPZ-S), which have very low affinity for CaM, had practically no inhibitory effect on lysosomal enzyme release. The inhibitory effect of TFP could be made irreversible by irradiating the cells with UV light. A sulfonamide derivative, W-7, N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide hydrochloride (10(-5)-2 X 10(-4) M), which selectively binds to CaM, inhibited the release of lysosomal enzymes from PMNs. In contrast, the chloride-deficient analog, W-5, N-(6-aminohexyl)-1-naphthalenesulfonamide hydrochloride, which interacts only weakly with CaM, had practically no inhibiting effect. The IC50 for enzyme release by a series of eight CaM antagonists was closely correlated (r = 0.89; P less than .001) with their affinity for binding to CaM, supporting the concept that these agents act by binding to CaM and thereby inhibiting lysosomal enzyme release.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:6502522

Marone, G; Poto, S; Columbo, M; Giugliano, R; Genovese, A; Condorelli, M



Lipopolysaccharide-induced pulmonary vascular sequestration of polymorphonuclear leukocytes is complement independent.  


Lipopolysaccharide (LPS) injected intravenously produces leukopenia and sequestration of polymorphonuclear leukocytes (PMN) in the pulmonary vascular bed. To evaluate the role of complement in this process, we used C5-sufficient (B10.D2/nSn) and C5-deficient (B10.D2/oSn) mice and Sprague-Dawley rats depleted of complement with Naja naja cobra venom factor (CVF). We found a comparable increase in the number of PMN in lung tissue of C5-sufficient and C5-deficient mice given Escherichia coli LPS (0127:B8, 3 mg/kg), revealing that LPS acts independently of C5 and its biologically active fragments. Intravenous injection of LPS (3 mg/kg) into rats caused significant intravascular complement activation as assessed by serum CH50 and resulted in an almost 10-fold increase in numbers of PMN in lung tissue. Pretreatment of rats with CVF (50 U) did not reduce LPS-induced PMN sequestration, suggesting that the process is independent of C3. As reported previously, we found large numbers of PMN in bronchoalveolar lavage samples of 24 h after injection of LPS (3 mg/kg). Complement depletion did not prevent LPS-induced migration of PMN. No PMN migration occurred 2, 6, 12, 24, or 48 h after injection of CVF alone, indicating that complement activation is not sufficient to cause PMN migration. In contrast to our findings in rats, no PMN migrated into airspaces of C5-sufficient and C5-deficient mice 24 or 48 h after injection of LPS (3 to 20 mg/kg). PMID:2064126

Cardozo, C; Edelman, J; Jagirdar, J; Lesser, M



Influence of inductive chemoradiotherapy on salivary polymorphonuclear leukocyte (SPMN) functions in oral cancer.  


Salivary polymorphonuclear leukocyte (SPMN) functions were examined in 18 patients with oral squamous cell carcinoma and in 20 healthy individuals. SPMN obtained from patients before therapy exhibited significantly less FMLP-stimulated chemotactic activity (132.4 +/- 17.5 cells/0.26 mm2) than that in SPMN from controls (177.1 +/- 11.6 cells/0.26 mm2), although no difference in phagocytosis was observed. When stimulated with PMA or FMLP, control SPMN generated superoxide (O2-) at levels of 50.3 +/- 10.5 pmol/min/10(4) cells and 88.4 +/- 15.4 pmol, respectively, while SPMN from untreated patients generated significantly reduced O2- in the presence of PMA or FMLP (24.3 +/- 3.5 pmol and 59.5 +/- 9.8 pmol, respectively). Only slightly lower chemiluminescence was observed in SPMN from untreated patients however, compared to controls, values being 68.0 +/- 18.9 vs 81.3 +/- 14.9 peak mV by PMA and 62.4 +/- 13.7 vs 64.4 +/- 12.9 peak mV by FMLP. Compared to Candida killing in control subjects (24.9 +/- 3.1%). SPMN from patients before treatment exhibited significantly reduced activity (18.7 +/- 4.9%). Further suppression of the SPMN functions examined was observed after chemoradio-therapy. Suppressed SPMN function in cancer patients, especially that associated with chemoradiotherapy, may therefore play a part in oral candidiasis. PMID:7823303

Ueta, E; Osaki, T; Yoneda, K; Yamamoto, T; Umazume, M



Role of Yersinia pestis Toxin Complex Family Proteins in Resistance to Phagocytosis by Polymorphonuclear Leukocytes.  


Yersinia pestis carries homologues of the toxin complex (Tc) family proteins, which were first identified in other Gram-negative bacteria as having potent insecticidal activity. The Y. pestis Tc proteins are neither toxic to fleas nor essential for survival of the bacterium in the flea, even though tc gene expression is highly upregulated and much more of the Tc proteins YitA and YipA are produced in the flea than when Y. pestis is grown in vitro. We show that Tc(+) and Tc(-) Y. pestis strains are transmitted equivalently from coinfected fleas, further demonstrating that the Tc proteins have no discernible role, either positive or negative, in transmission by the flea vector. Tc proteins did, however, confer Y. pestis with increased resistance to killing by polymorphonuclear leukocytes (PMNs). Resistance to killing was not the result of decreased PMN viability or increased intracellular survival but instead correlated with a Tc protein-dependent resistance to phagocytosis that was independent of the type III secretion system (T3SS). Correspondingly, we did not detect T3SS-dependent secretion of the native Tc proteins YitA and YipA or the translocation of YitA- or YipA-?-lactamase fusion proteins into CHO-K1 (CHO) cells or human PMNs. Thus, although highly produced by Y. pestis within the flea and related to insecticidal toxins, the Tc proteins do not affect interaction with the flea or transmission. Rather, the Y. pestis Tc proteins inhibit phagocytosis by mouse PMNs, independent of the T3SS, and may be important for subverting the mammalian innate immune response immediately following transmission from the flea. PMID:23959716

Spinner, Justin L; Carmody, Aaron B; Jarrett, Clayton O; Hinnebusch, B Joseph



Radiation-like modification of bases in DNA exposed to tumor promoter-activated polymorphonuclear leukocytes.  


Oxygen species generated by human polymorphonuclear leukocytes (PMNs) activated by 12-O-tetradecanoylphorbol-13-acetate (TPA) caused the formation of 5-hydroxymethyl-2'-deoxyuridine (HMdUrd), and (+) and (-) diastereoisomers of cis-thymidine glycol (dTG) in DNA that was exposed to them. There were 9 HMdUrds and 31 dTGs formed per 1 X 10(6) thymidine residues. When Fe(II)/ethylenediaminetetraacetic acid was added to TPA-activated PMNs at 0, 10, 15, and 20 min after TPA, HMdUrd formation increased 5-, 13-, 30-, and 35-fold. Although dTG was initially formed in larger amounts than HMdUrd, it eventually decreased but was still 5-, 6-, 5.5-, and 3-5-fold, respectively, higher than in the absence of iron. From 65 to 1800 times more HMdUrd was formed in DNA when autologous plasma was present during incubation of DNA with TPA-activated PMNs than in its absence. The levels of dTG also varied from about the same as HMdUrd to the nondetectable. Reconstituted human serum transferrin used instead of plasma or Fe(II) also supported the formation of HMdUrd and dTG. When DNA was treated with Fe(II)-reduced H2O2 in the absence of PMNs and TPA, both derivatives were formed. However, the same treatment of marker dTG of dTG-containing polydeoxyadenylic-thymidylic acid caused the decomposition of dTG. Thus, the reduction of hydrogen peroxide by Fe(II) complexed to either ethylenediaminetetraacetic acid or amino acids amy be responsible for the formation of HMdUrd and dTG and for subsequent decomposition of dTG in DNA exposed to the TPA-activated PMNs. PMID:3756901

Frenkel, K; Chrzan, K; Troll, W; Teebor, G W; Steinberg, J J



Altered polymorphonuclear leukocyte Fc gamma R expression contributes to decreased candicidal activity during intraabdominal sepsis  

SciTech Connect

We investigated the effects of untreated intraabdominal sepsis on polymorphonuclear leukocyte (PMN) candicidal activity. Two groups of swine were studied. Group I (n=6) underwent sham laparotomy, group II (n=7) underwent cecal ligation and incision. Untreated intraabdominal sepsis resulted in a progressive decrease in PMN candicidal activity. Concomitant rosetting and phagocytosis assays demonstrated a decrease in both the attachment and phagocytosis of Candida albicans opsonized with both normal and septic swine serum by PMNs in group II. Iodine 125-labeled swine immunoglobulin G (IgG) and fluorescein isothioalanate (FITC)-labeled swine IgG were used to investigate Fc gamma receptor ligand interactions. Scatchard analyses demonstrated a progressive decline in both the binding affinity constant and number of IgG molecules bound per PMN. Stimulation of the oxidative burst markedly reduced 125I-labeled IgG binding in both group I and group II, with a greater decrement being seen in animals with intraabdominal sepsis. Further, in group II, PMN recycling of the Fc gamma receptor to the cell surface after generation of the oxidative burst was reduced by postoperative day 4. Binding of monoclonal antibodies to Fc gamma receptor II, but not Fc gamma receptor I/III markedly reduced intracellular candicidal activity. Immunofluorescence studies revealed a homogeneous pattern of FITC-IgG uptake by nearly all group I PMNs, whereas by postoperative day 8 a substantial number of PMNs from group II failed to internalize the FITC-IgG. These studies suggest that untreated intraabdominal sepsis reduces PMN candicidal activity and that this is due, in part, to altered PMN Fc gamma receptor ligand interactions.

Simms, H.H.; D'Amico, R.; Monfils, P.; Burchard, K.W. (Rhode Island Hospital, Providence (USA))



Resistance of a Tn4351-generated polysaccharide mutant of Porphyromonas gingivalis to polymorphonuclear leukocyte killing.  

PubMed Central

In this study, we describe the development of an efficient transpositional mutagenesis system for Porphyromonas gingivalis using the Bacteroides fragilis transposon Tn4351. Using this system, we have isolated and characterized a Tn4351-generated mutant of P. gingivalis A7436, designated MSM-1, which exhibits enhanced resistance to polymorphonuclear leukocyte (PMN) phagocytosis and killing. P. gingivalis MSM-1 was initially selected based on its colony morphology; MSM-1 appeared as a mucoid, beige-pigmented colony. Analysis of P. gingivalis MSM-1 by electron microscopy and staining with ruthenium red revealed the presence of a thick ruthenium red-staining layer that was twice the thickness of this layer observed in the parent strain. P. gingivalis MSM-1 was found to be more hydrophilic than strain A7436 by hydrocarbon partitioning. Analysis of phenol-water extracts prepared from P. gingivalis A7436 and MSM-1 by Western (immunoblot) analysis and immunodiffusion with hyperimmune sera raised against A7436 and MSM-1 revealed the loss of a high-molecular-weight anionic polysaccharide component in extracts prepared from MSM-1. P. gingivalis MSM-1 was also found to be more resistant to PMN phagocytosis and intracellular killing than the parent strain, as assessed in a fluorochrome phagocytosis microassay. These differences were statistically significant (P < 0.05) when comparing PMN phagocytosis in nonimmune serum and intracellular killing in nonimmune and immune sera. P. gingivalis MSM-1 was also more resistant to killing by crude granule extracts from PMNs than was P. gingivalis A7436. These results indicate that the increased evasion of PMN phagocytosis and killing exhibited by P. gingivalis MSM-1 may result from alterations in polysaccharide-containing antigens.

Genco, C A; Schifferle, R E; Njoroge, T; Forng, R Y; Cutler, C W



Polymorphonuclear leukocyte lysosomal proteases, cathepsins B and D affect the fibrinolytic system in human umbilical vein endothelial cells.  


To clarify the physiological role played by neutrophil lysosomal protease in cultured human umbilical vein endothelial cells (HUVEC), we studied the effects of cathepsins B and D released from activated polymorphonuclear leukocytes on the fibrinolytic system in HUVEC. Cathepsins B and D reduced the antigens of tissue-type plasminogen activator, and they increased both the antigens and the activity of plasminogen activator inhibitor-1. These results suggest that cathepsins B and D are involved in the thrombotic tendency, since they inhibited the fibrinolytic system in cultured HUVEC. PMID:9244167

Kimura, Y; Yokoi-Hayashi, K



Polymorphonuclear leukocyte adhesion triggers the disorganization of endothelial cell-to-cell adherens junctions  

PubMed Central

Polymorphonuclear leukocytes (PMN) infiltration into tissues is frequently accompanied by increase in vascular permeability. This suggests that PMN adhesion and transmigration could trigger modifications in the architecture of endothelial cell-to-cell junctions. In the present paper, using indirect immunofluorescence, we found that PMN adhesion to tumor necrosis factor-activated endothelial cells (EC) induced the disappearance from endothelial cell-to-cell contacts of adherens junction (AJ) components: vascular endothelial (VE)-cadherin, alpha-catenin, beta-catenin, and plakoglobin. Immunoprecipitation and Western blot analysis of the VE- cadherin/catenin complex showed that the amount of beta-catenin and plakoglobin was markedly reduced from the complex and from total cell extracts. In contrast, VE-cadherin and alpha-catenin were only partially affected. Disorganization of endothelial AJ by PMN was not accompanied by EC retraction or injury and was specific for VE- cadherin/catenin complex, since platelet/endothelial cell adhesion molecule 1 (PECAM-1) distribution at cellular contacts was unchanged. PMN adhesion to EC seems to be a prerequisite for VE-cadherin/catenin complex disorganization. This phenomenon could be fully inhibited by blocking PMN adhesion with an anti-integrin beta 2 mAb, while it could be reproduced by any condition that induced increase of PMN adhesion, such as addition of PMA or an anti-beta 2-activating mAb. The effect on endothelial AJ was specific for PMN since adherent activated lymphocytes did not induce similar changes. High concentrations of protease inhibitors and oxygen metabolite scavengers were unable to prevent AJ disorganization mediated by PMN. PMN adhesion to EC was accompanied by increase in EC permeability in vitro. This effect was dependent on PMN adhesion, was not mediated by proteases and oxygen- reactive metabolites, and could be reproduced by EC treatment with EGTA. Finally, immunohistochemical analysis showed that VE-cadherin distribution was affected by PMN adhesion to the vessel wall in vivo too. This work suggests that PMN adhesion could trigger intracellular signals in EC that possibly regulate VE-cadherin /catenin complex disorganization. This effect could increase EC permeability and facilitate PMN transmigration during the acute inflammatory reaction.



Benidipine, an anti-hypertensive drug, inhibits reactive oxygen species production in polymorphonuclear leukocytes and oxidative stress in salt-loaded stroke-prone spontaneously hypertensive rats.  


Oxidative stress is associated with exacerbation of renal injuries in hypertension. In clinical studies benidipine hydrochloride (benidipine), a dihydropyridine calcium channel blocker with antioxidant activity, reduced oxidative stress. However, the mechanism of suppression of oxidative stress remains to be fully characterized. Reactive oxygen species production by polymorphonuclear leukocyte plays important pathological roles in hypertension. Therefore, we examined the effects of benidipine both on reactive oxygen species production of human polymorphonuclear leukocytes and oxidative stress of an animal model. Human peripheral polymorphonuclear leukocytes or polymorphonuclear leukocyte-like differentiated HL-60 cells were used to examine effects of benidipine (0.1-30 microM) on formyl-Met-Leu-Phe-induced reactive oxygen species production, calcium mobilization, NADPH oxidase activation and phosphorylation of protein kinase C substrates. High-salt (8% NaCl) loaded stroke-prone spontaneously hypertensive rats were treated with or without benidipine (1, 3, 10 mg/kg/day) for 2 weeks, and thiobarbituric acid reactive substances, a plasma oxidative stress marker, and renal expression of oxidative stress-induced genes were measured. Benidipine concentration-dependently suppressed formyl-Met-Leu-Phe-induced reactive oxygen species production in polymorphonuclear leukocytes more potently than other calcium channel blockers such as amlodipine, azelnidipine, nitrendipine and nifedipine. Benidipine partially inhibited all of intracellular Ca(2+) elevation, protein kinase C activation and NADPH oxidase activation. Salt loading in stroke-prone spontaneously hypertensive rats augmented plasma thiobarbituric acid reactive substances levels; renal dysfunction; and renal expression of transforming growth factor-beta, collagen I and collagen III mRNAs; which were attenuated by benidipine treatment. These results indicate that benidipine prevents the polymorphonuclear leukocyte-derived reactive oxygen species production, which is due at least in part to its antioxidant action and inhibition of Ca(2+)/protein kinase C/NADPH oxidase signaling. The attenuation of reactive oxygen species production might contribute to the drug's reduction of oxidative stress and renal injuries in hypertension. PMID:18048030

Matsubara, Masahiro; Akizuki, Osamu; Ikeda, Jun-ichi; Saeki, Koji; Yao, Kozo; Sasaki, Katsutoshi



Polymorphonuclear leukocytes as a significant source of tumor necrosis factor-alpha in endotoxin-challenged lung tissue.  

PubMed Central

The kinetic expression and potential cellular source of tumor necrosis factor-alpha (TNF-alpha) in lipopolysaccharide-(LPS) induced acute lung inflammation was investigated using a rat model by Northern blot analysis, in situ hybridization and immunohistochemistry. LPS induced a polymorphonuclear leukocyte infiltrate in the lung that peaked between 6 and 24 hours. TNF-alpha messenger (m)RNA was strongly induced by LPS in whole lung tissues shown by Northern analysis. Both alveolar macrophages and polymorphonuclear leukocytes (PMNs), purified from bronchoalveolar lavage fluids of LPS-treated rats, were shown to express TNF-alpha mRNA by Northern analysis. However, PMNs displayed several times more TNF-alpha mRNA, relative to actin mRNA, than alveolar macrophages at 6 and 12 hours. By in situ hybridization, most of the cells positive for TNF-alpha mRNA at 6 and 12 hours seemed to be PMNs located within the tissue near bronchioles or vessels. By immunohistochemistry, TNF-alpha protein was localized mainly to alveolar macrophages at early times (1 to 3 hours) after LPS challenge, and thereafter, PMNs seemed to be the predominant source of TNF-alpha protein as more than 90% of total intraalveolar positive cells at 6 and 12 hours were PMN. Thus, our data provide the first in vivo evidence that PMNs can serve as a significant source of TNF-alpha at sites of acute inflammation. Images Figure 2 Figure 3

Xing, Z.; Kirpalani, H.; Torry, D.; Jordana, M.; Gauldie, J.



Influence of He-Ne laser radiation on biogenic amines content and cytochemical parameters of polymorphonuclear leukocytes in short-term stress  

NASA Astrophysics Data System (ADS)

In experiments on white male rats short-term immobilization- sound stress was modelled. Decrease of glycogen content and myeloperoxidase activity, increase of lysosomal cationic proteins level and NBT-test parameters as well as fall of adrenaline, dopamine and 5-hydroxytryptamine amount in polymorphonuclear leukocytes were observed. Preliminary transcutaneous He-Ne laser irradiation modified metabolic reaction of leukocytes to stress and prevented stress- induced decrease of biogenic amines content in cells.

Brill, Grigory E.; Dobrovolsky, Gennady A.; Romanova, Tatyana P.; Porozova, Svetlana G.; Brill, Alexander G.



Time Course of the Effect of Nifedipine Therapy and Its Discontinuation on [Ca2+]i and Phagocytosis of Polymorphonuclear Leukocytes from Hemodialysis Patients  

Microsoft Academic Search

The abnormalities in [Ca2+]i and phagocytosis of polymorphonuclear leukocytes (PMNLs) from hemodialysis (HD) patients are significantly improved by their treatment with nifedipine. However, the rapidity with which this agent induces its benefical effect and whether these derangements re-emerge after cessation of therapy are not known. We studied 5 HD patients before, during and after treatment with nifedipine. Before treatment with

Jadwiga M. Alexiewicz; Miroslaw Smogorzewski; Sukhpal K. Gill; Mohammad Akmal; Shaul G. Massry



The effect of aurothiomalate on the oxidative burst of polymorphonuclear leukocytes varies with the quantity of drug in myocrisin ampoules.  


The antirheumatic drug, sodium aurothiomalate (GSTM), is not a well defined substance and chemical changes occur in the heat sterilization of the commercial ampoules (Myocrisin). In a comparison of the pharmacological properties of Myocrisin with freshly prepared solutions of GSTM, their effects on the chemiluminescence of polymorphonuclear leukocytes (PMN) activated by phorbol myristate acetate (PMA) were studied. Chemiluminescence was measured in the presence of GSTM from solid material and from Myocrisin ampoules. Myocrisin from 1 and 5 mg ampoules and GSTM in fresh solutions heated at 95 degrees C for 30 min inhibited chemiluminescence, whereas Myocrisin from the higher strength (10-50 mg) ampoules and GSTM in unheated solutions showed no effect at low concentrations and enhancement of chemiluminescence at higher concentrations. Since the gold complexes present in the different strength Myocrisin ampoules do not have identical biological effects, the use of GSTM in investigational studies should involve consideration of its source. PMID:1907663

Rudkowski, R; Graham, G G; Ziegler, J B; Champion, G D



Whole blood leukocytes isolation with microfabricated filter for cell analysis.  


The flow cytometric analysis of leukocytes in whole blood usually requires isolation of leukocytes from other components of whole blood. Density gradient centrifugation and red blood cell lysis are the most commonly used methods to separate leukocytes but come with significant limitations. We report the results of the evaluation of a microfabricated filtration device for blood preparation that separates erythrocytes from leukocytes based on their size and mechanical properties. The microfabricated filter evaluated here requires a rapid and simple procedure and results in high leukocytes recovery without introducing bias among the leukocyte subpopulations. The filter removes erythrocytes, platelets, plasma proteins, and unbound staining reagent. This gentle filtration process produces very clean stained leukocytes for cytometric analysis without any apparent damage to leukocytes. PMID:22110022

Yu, Liping; Warner, Patrick; Warner, Brian; Recktenwald, Diether; Yamanishi, Douglas; Guia, Antonio; Ghetti, Andrea



Promotion of DNA strand breaks in cocultured mononuclear leukocytes by protein kinase C-dependent prooxidative interactions of benoxaprofen, human polymorphonuclear leukocytes, and ultraviolet radiation  

SciTech Connect

At concentrations of 5 micrograms/ml and greater the nonsteroidal antiinflammatory drug benoxaprofen caused dose-related activation of lucigenin-enhanced chemiluminescence in human polymorphonuclear leukocytes (PMNL). Benoxaprofen-mediated activation of lucigenin-enhanced chemiluminescence by PMNL was increased by UV radiation and was particularly sensitive to inhibition by the selective protein kinase C inhibitor H-7. To identify the molecular mechanism of the prooxidative activity of benoxaprofen, the effects of the nonsteroidal antiinflammatory drug on the activity of purified protein kinase C in a cell-free system were investigated. Benoxaprofen caused a dose-related activation of protein kinase C by interaction with the binding site for the physiological activator phosphatidylserine, but could not replace diacylglycerol. When autologous mononuclear leukocytes (MNL) were cocultured with PMNL and benoxaprofen in combination, but not individually, the frequency of DNA strand breaks in MNL was markedly increased. UV radiation significantly potentiated damage to DNA mediated by benoxaprofen and PMNL. Inclusion of superoxide dismutase, H-7, and, to a much lesser extent, catalase during exposure of MNL to benoxaprofen-activated PMNL prevented oxidant damage to DNA. These results clearly demonstrate that potentially carcinogenic prooxidative interactions, which are unlikely to be detected by conventional assays of mutagenicity, may occur between phagocytes, UV radiation, and certain pharmacological agents.

Schwalb, G.; Beyers, A.D.; Anderson, R.; Nel, A.E.



Blood leukocyte and spleen lymphocyte immune response of spleen lymphocytes and whole blood leukocytes of hamsters  

SciTech Connect

This study was designed to evaluate the effects of chronic physical activity on the immune response of spleen lymphocytes and whole blood leukocytes of hamsters. Animals were kept sedentary or allowed to exercise spontaneously on running wheels for eight weeks. Physically active animals averaged 12 kilometers per day. The immune response of spleen lymphocytes whole blood leukocytes was evaluated by {sup 3}H-thymidine incorporation in response to Concanavalin A or lipopolysaccharide. There was no treatment effect between physically active and sedentary hamster in response of spleen lymphocytes. The immune response of whole blood leukocytes to these mitogens was significantly greater in physically active vs. sedentary hamsters. These results demonstrate that chronic physical activity has the capacity to modulate immunoresponses.

Peters, B.A.; Sothmann, M.; Wehrenberg, W.B. (Univ. of Wisconsin, Milwaukee (USA))



The interaction in vitro between human polymorphonuclear leukocytes and Neisseria gonorrhoeae cultivated in the chick embryo  

PubMed Central

Cultivation of Neisseria gonorrhoeae in the allantoic cavity of 10-day chick embryos ensured the following necessary properties for subsequent quantitive in vitro phagocytosis studies of viable gonococci: log phase of growth, resistance to the cidal effect of fresh human serum, maintenance of colonial type, and absence of clumping. Employing a modification of the Maaloe technique, phagocytosis of log-phase type 1 and 2 gonococci by human PMN leukocytes did not occur in the presence or absence of serum. These findings indicate that log-phase type 1 and 2 gonococci possess antiphagocytic surface factors Stationary-phase organisms of the same colonial type were ingested and rapidly killed by human PMN leukocytes under similar experimental conditions, thus emphasizing the necessity to employ log-phase gonococci in the study of phagocytosis and antiphagocytic surface factors. Log-phase type 4 gonococci were ingested and rapidly killed by human PMN leukocytes in the presence of fresh human serum but not heat-inactivated serum or in the absence of serum. Morphologic studies demonstrated that log-phase viable gonococci attach to the surface membrane of human PMN leukocytes. Interiorization of avirulent but not virulent organisms was observed in the presence of fresh human serum. Gonococci-human PMN leukocyte interactions thus provide a model for the investigation of the nonimmunologic and immunologic parameters associated with the attachment and ingestion stages of phagocytosis.



Fragmentation of gelatin-bound fibronectin (Fn) by inflammatory polymorphonuclear leukocytes (PMNL): A role for leukocyte elastase  

SciTech Connect

Fragmentation of lung matrix Fn by proteases released from activated leukocytes sequestered in the lung has been implicated in lung vascular injury. The authors determined if Fn bound to a denatured collagen (gelatin) surface was susceptible to degradation by inflammatory PMNLs. Tissue culture wells coated with 1.5% denatured collagen (2 ml/well) prior to the addition of rat peritonal exudate cells, harvested 16 hours after i.p. sterile casein. Inflammatory PMNLs (1 {times} 10{sup 6}) stimulated with zymosan (1 mg) released 3 times more {sup 125}I-Fn into the culture media (DMEM) during a 4 hour incubation as compared to unstimulated PMNLs (2885{plus minus}95 cpm vs 1027{plus minus}82 cpm/100 ul). {sup 125}I-Fn released by stimulated PMNLs was markedly blocked by addition of a leukocyte elastase inhibitor (AAPVCK), moderately blocked by a trypsin inhibitor (TLCK), and not blocked by a thrombin inhibitor (Hirudin). Western blot analysis demonstrated fragmentation of released Fn. Thus, Fn complexed with denatured collagen is susceptible to proteolysis by stimulated inflammatory PMNLs. This may contribute to lung vascular injury with sepsis and intravascular coagulation which elicit sequestration of activated PMNLs in the lung.

Daudi, I.; Gudewicz, P.W.; Saba, T.M.; Cho, E.; Lewis, M. (Albany Medical Coll., NY (United States))



Increased Production of Oxygen Free Radicals by Polymorphonuclear Leukocytes in Heart Failure Due to Aortic Stenosis  

Microsoft Academic Search

Oxygen free radicals have been linked to a wide variety of cellular damage in biological systems. Poly morphonuclear (PMN) leukocytes stimulation is one of the known sources for oxygen free radicals. It has been suggested that oxygen free radicals depress the excitation-con traction coupling in cardiac muscle. It is possible that a decrease in the myocardial contractility in heart fail

Kailash Prasad; Jawahar Kalra; K. Lorne Massey; Baikunth Bharadwaj



Nongenomic effect of thyroid hormone on free-radical production in human polymorphonuclear leukocytes  

Microsoft Academic Search

Over the past few years increasing evidence has suggested the nongenomic effects of thyroid hormone, such as the activation of the signal transduction pathways and the activation of nuclear factor-B by the induction of oxidat- ive stress. The present study was undertaken to investigate the effect of thyroid hormone on human polymorpho- nuclear leukocytes (PMNLs) which are known as import-

E Mezosi; J Szabo; E V Nagy; A Borbely; E Varga; G Paragh; Z Varga



Prevention of stromal ulceration in the alkali-burned rabbit cornea by glued-on contact lens. Evidence for the role of polymorphonuclear leukocytes in collagen degradation  

Microsoft Academic Search

Stromal ulceration of the alkali-burned rabbit cornea ivas found to be associated invariably with phagocytically active polymorphonuclear leukocytes (PMNs). A glued-on methylmethacry- late lens applied to corneas soon after burning, however, prevented re-epithelialization and also prevented PMN infiltration of the stroma and stromal ulceration. Subsequent partial detachment or complete removal of the lens residted in epithelial resurfacing of the stroma,

K. R. Kenyon; M. Berman



Degranulation of polymorphonuclear leukocytes is induced by multivalent cross-linking of wheat germ agglutinin binding site(S) on cell membrane  

Microsoft Academic Search

Polymorphonuclear leukocyte (PMN) surface membrane glycoproteins very likely are involved in the phenomenon of stimulus-response coupling. Previously, we have shown that subagglutinating concentrations of the plant lectin, wheat germ agglutinin (WGA) specifically and irreversibly inhibitedN-formyl-methionyl-leucyl-phenyl-alanine (FMLP)-mediated PMN chemotaxis. WGA did not affect the binding of [3H]FMLP to its receptor on the PMN plasma membrane. We have examined the possibility that

H. Daniel Perez; Richard R. Ong



Immunolocalization of CXC chemokine and recruitment of polymorphonuclear leukocytes in the rat molar periodontal tissue after topical application of lipopolysaccharide  

Microsoft Academic Search

This study investigated the recruitment of polymorphonuclear leukocytes (PMNs) and the immunolocalization of CXC chemokines, including macrophage inflammatory protein-2 (MIP-2) and cytokine-induced neutrophil chemoattractant-2 (CINC-2) in rat periodontal tissue after topical application of lipopolysaccharide (LPS; 5 mg\\/ml) from Escherichia coli into the rat molar gingival sulcus. In normal periodontal tissues, a small number of MIP-2- and CINC-2-positive cells were seen in junctional

Mutsumi Miyauchi; Shoji Kitagawa; Masae Hiraoka; Akihisa Saito; Sunao Sato; Yasusei Kudo; Ikuko Ogawa; Takashi Takata



Inhibitors of actin polymerisation stimulate arachidonic acid release and 5-lipoxygenase activation by upregulation of Ca 2+ mobilisation in polymorphonuclear leukocytes involving Src family kinases  

Microsoft Academic Search

Here, we show that actin polymerisation inhibitors such as latrunculin B (LB), and to a minor extent also cytochalasin D (Cyt D), enhance the release of arachidonic acid (AA) as well as nuclear translocation of 5-lipoxygenase (5-LO) and 5-LO product synthesis in human polymorphonuclear leukocytes (PMNL), challenged with thapsigargin (TG) or N-formyl-methionyl-leucyl-phenylalanine. The concentration-dependent effects of LB (EC50 ?200 nM)

Lutz Fischer; Daniel Poeckel; Eva Buerkert; Dieter Steinhilber; Oliver Werz




Microsoft Academic Search

At infection sites, synthesis of interleukin (IL-)1? by polymorphonuclear leukocytes (PMNs) facilitates the recruitment of inflammatory cells and enhances the inflammatory response. We investigated the role of protein kinase C (PKC) and Ca2+in the induction of PMN IL-1? gene expression by GM-CSF. The PKC inhibitors chelerythrine and H7 blocked induction of IL-1? mRNA expression in human PMNs. HA1004, an H7

Marilyn C Fernandez; Phillip T Marucha; Isolde G Rojas; John D Walters



beta. -Endorphin and related peptides suppress phorbol myristate acetate-induced respiratory burst in human polymorphonuclear leukocytes  

SciTech Connect

In the present study, the immunomodulatory effect of {beta}-endorphin ({beta}-E) and shorter pro-opiomelancortin (POMC) fragments was evaluated by assessing their influence on respiratory burst in human polymorphonuclear leukocytes (PMN). The effect of the peptides on phorbol myristate acetate (PMA)-stimulated production of reactive oxygen metabolites was measured in a lucigenin-enhanced chemiluminescence (CL) assay. Both POMC peptides with opiate-like activity and their non-opioid derivatives were tested. With the exception of {alpha}-E, PMA-stimulated respiratory burst was suppressed by all POMC fragments tested. A U-shaped dose-response relation was observed. Doses lower than 10{sup {minus}17}M and higher than 10{sup {minus}8}M were without effect. {beta}-E and dT{beta}E both suppressed PMA-induced oxidative burst in human PMN at physiological concentrations. {gamma}-E and dT{gamma}E proved to be less potent inhibitors, reaching maximal effect at higher concentrations. DE{gamma}E exerted an even less pronounced but still significant suppressive effect at the concentration of 10{sup {minus}10}M. None of the endorphins tested was shown to affect resting oxidative metabolism in the PMN. The modulatory effects of the opioid peptides could not be blocked by the opioid antagonist naloxone.

Diamant, M.; Henricks, P.A.J.; Nijkamp, F.P.; de Wied, D. (Univ. of Utrecht (Netherlands))



Effects of Acer okamotoanum sap on the function of polymorphonuclear neutrophilic leukocytes in vitro and in vivo.  


Sap is a plant fluid that primarily consists of water and small amounts of mineral elements, sugars, hormones and other nutrients. Acer mono (A. mono) is an endemic Korean mono maple which was recently suggested to have health benefits due to its abundant calcium and magnesium ion content. In the present study, we examined the effects of sap from Acer okamotoanum (A. okamotoanum) on the phagocytic response of mouse neutrophils in vivo and rat and canine neutrophils in vitro. We tested the regulation of phagocytic activity, oxidative burst activity (OBA) and the levels of filamentous polymeric actin (F-actin) in the absence and presence of dexamethasone (DEX) in vitro and in vivo. Our results showed that DEX primarily reduced OBA in the mouse neutrophils, and that this was reversed in the presence of the sap. By contrast, the phagocytic activity of the mouse cells was not regulated by either DEX or the sap. Rat and canine polymorphonuclear neutrophilic leukocytes (PMNs) responded in vitro to the sap in a similar manner by increasing OBA. However, regulation of phagocytic activity by the sap was different between the species. In canine PMNs, phagocytic activity was enhanced by the sap at a high dose, while it did not significantly modulate this activity in rat PMNs. These findings suggest that the sap of A. okamotoanum stimulates neutrophil activity in the mouse, rat and canine by increasing OBA in vivo and in vitro, and thus may have a potential antimicrobial effect in the PMNs of patients with infections. PMID:23165961

An, Beum-Soo; Kang, Ji-Houn; Yang, Hyun; Yang, Mhan-Pyo; Jeung, Eui-Bae



IgG Endopeptidase SeMac does not Inhibit Opsonophagocytosis of Streptococcus equi Subspecies equi by Horse Polymorphonuclear Leukocytes  

PubMed Central

The secreted Mac protein made by group A Streptococcus (GAS) inhibits opsonophagocytosis of GAS by human polymorphonuclear leukocytes (PMNs). This protein also has the endopeptidase activity against human immunoglobulin G (IgG), and the Cys94, His262 and Asp284 are critical for the enzymatic activity. The horse pathogen Streptococcus equi subspecies equi produces a homologue of Mac (SeMac). SeMac was characterized to determine whether SeMac has IgG endopeptidase activity and inhibits opsonophagocytosis of S. equi by horse PMNs. The gene was cloned and recombinant SeMac was overexpressed in Escherichia coli and purified to homogeneity. Mice with experimental S. equi infection and horses with strangles caused by S. equi seroconverted to SeMac, indicating that SeMac is produced in vivo during infection. SeMac has endopeptidase activity against human IgG. However, the protein just cleaves a small fraction, which may be IgG1 only, of horse IgG. Replacement of Cys102 with Ser or His272 with Ala abolishes the enzymatic activity of SeMac, and the Asp294Ala mutation greatly decreases the enzymatic activity. SeMac does not inhibit opsonophagocytosis of S. equi by horse PMNs but opsonophagocytosis of GAS by human PMNs. Thus, SeMac is a cysteine endopeptidase with a limited activity against horse IgG and must have other function.

Liu, Mengyao; Lei, Benfang



IgG Endopeptidase SeMac does not Inhibit Opsonophagocytosis of Streptococcus equi Subspecies equi by Horse Polymorphonuclear Leukocytes.  


The secreted Mac protein made by group A Streptococcus (GAS) inhibits opsonophagocytosis of GAS by human polymorphonuclear leukocytes (PMNs). This protein also has the endopeptidase activity against human immunoglobulin G (IgG), and the Cys94, His262 and Asp284 are critical for the enzymatic activity. The horse pathogen Streptococcus equi subspecies equi produces a homologue of Mac (SeMac). SeMac was characterized to determine whether SeMac has IgG endopeptidase activity and inhibits opsonophagocytosis of S. equi by horse PMNs. The gene was cloned and recombinant SeMac was overexpressed in Escherichia coli and purified to homogeneity. Mice with experimental S. equi infection and horses with strangles caused by S. equi seroconverted to SeMac, indicating that SeMac is produced in vivo during infection. SeMac has endopeptidase activity against human IgG. However, the protein just cleaves a small fraction, which may be IgG1 only, of horse IgG. Replacement of Cys102 with Ser or His272 with Ala abolishes the enzymatic activity of SeMac, and the Asp294Ala mutation greatly decreases the enzymatic activity. SeMac does not inhibit opsonophagocytosis of S. equi by horse PMNs but opsonophagocytosis of GAS by human PMNs. Thus, SeMac is a cysteine endopeptidase with a limited activity against horse IgG and must have other function. PMID:20556207

Liu, Mengyao; Lei, Benfang



Role of the Yersinia pestis Ail Protein in Preventing a Protective Polymorphonuclear Leukocyte Response during Bubonic Plague?  

PubMed Central

The ability of Yersinia pestis to forestall the mammalian innate immune response is a fundamental aspect of plague pathogenesis. In this study, we examined the effect of Ail, a 17-kDa outer membrane protein that protects Y. pestis against complement-mediated lysis, on bubonic plague pathogenesis in mice and rats. The Y. pestis ail mutant was attenuated for virulence in both rodent models. The attenuation was greater in rats than in mice, which correlates with the ability of normal rat serum, but not mouse serum, to kill ail-negative Y. pestis in vitro. Intradermal infection with the ail mutant resulted in an atypical, subacute form of bubonic plague associated with extensive recruitment of polymorphonuclear leukocytes (PMN or neutrophils) to the site of infection in the draining lymph node and the formation of large purulent abscesses that contained the bacteria. Systemic spread and mortality were greatly attenuated, however, and a productive adaptive immune response was generated after high-dose challenge, as evidenced by high serum antibody levels against Y. pestis F1 antigen. The Y. pestis Ail protein is an important bubonic plague virulence factor that inhibits the innate immune response, in particular the recruitment of a protective PMN response to the infected lymph node.

Hinnebusch, B. Joseph; Jarrett, Clayton O.; Callison, Julie A.; Gardner, Donald; Buchanan, Susan K.; Plano, Gregory V.



Role of the Yersinia pestis Ail protein in preventing a protective polymorphonuclear leukocyte response during bubonic plague.  


The ability of Yersinia pestis to forestall the mammalian innate immune response is a fundamental aspect of plague pathogenesis. In this study, we examined the effect of Ail, a 17-kDa outer membrane protein that protects Y. pestis against complement-mediated lysis, on bubonic plague pathogenesis in mice and rats. The Y. pestis ail mutant was attenuated for virulence in both rodent models. The attenuation was greater in rats than in mice, which correlates with the ability of normal rat serum, but not mouse serum, to kill ail-negative Y. pestis in vitro. Intradermal infection with the ail mutant resulted in an atypical, subacute form of bubonic plague associated with extensive recruitment of polymorphonuclear leukocytes (PMN or neutrophils) to the site of infection in the draining lymph node and the formation of large purulent abscesses that contained the bacteria. Systemic spread and mortality were greatly attenuated, however, and a productive adaptive immune response was generated after high-dose challenge, as evidenced by high serum antibody levels against Y. pestis F1 antigen. The Y. pestis Ail protein is an important bubonic plague virulence factor that inhibits the innate immune response, in particular the recruitment of a protective PMN response to the infected lymph node. PMID:21969002

Hinnebusch, B Joseph; Jarrett, Clayton O; Callison, Julie A; Gardner, Donald; Buchanan, Susan K; Plano, Gregory V



Solubilization and functional reconstitution of polymorphonuclear leukocyte formyl-Methionyl-Leucyl-Phenylalanine receptors and guanine nucleotide binding proteins  

SciTech Connect

Formyl-Methionyl-Leucyl-Phenylalanine (fMLP) binds to specific polymorphonuclear leukocyte plasma membrane receptors stimulating chemotaxis and bactericidal responses. One of the initial events of the ligand receptor interaction is a rise in inositol trisphosphate, which triggers intracellular calcium release. The generation of inositol trisphosphate is mediated by the fMLP-activated phospholipase C via a GTP-binding protein (G-protein). In analogy to the adrenergic stimulation of adenylate cyclase, the following signal transduction model has been proposed: The fMLP receptor activates a G-protein which then stimulates phospholipase C to hydrolyse phosphatidylinositol bisphosphate to inositol trisphosphate and diacylglycerol. This work has focused on characterizing the structural and functional coupling fMLP receptor and G-proteins in native membranes, detergent micelles and reconstituted phospholipid vesicles. Tight coupling between the fMLP receptor and G-protein has been demonstrated in both native and solubilized membranes by assaying quanine nucleotide-induced inhibition of (/sup 3/H)fMLP binding and fMLP stimulated GTPase activity.

Williamson, K.C.



Uptake and intracellular activity of sparfloxacin in human polymorphonuclear leukocytes and tissue culture cells.  

PubMed Central

The penetration of sparfloxacin into human neutrophils (PMN) and different tissue culture cells (HEp-2 and McCoy) was evaluated. The cellular to extracellular concentration ratios (C/E) of sparfloxacin were always higher than 4 at extracellular concentrations ranging from 0.5 to 25 mg/liter. The uptake of sparfloxacin by PMN was rapid, nonsaturable, reversible, not energy dependent, and significantly reduced at pH 8. The penetration of this agent into PMN was similar when viable and Formalin-killed cells were used and was not affected by environmental temperature. Ingestion of opsonized zymosan significantly increased the amount of PMN-associated sparfloxacin. Sparfloxacin at a concentration of 0.5 mg induced a significant reduction in the survival of intracellular Staphylococcus aureus. It is concluded that sparfloxacin reaches intracellular concentrations within leukocytic cells much higher than extracellular concentrations, while remaining active intracellularly.

Garcia, I; Pascual, A; Guzman, M C; Perea, E J



Impaired carbohydrate metabolism of polymorphonuclear leukocytes in glycogen storage disease Ib.  

PubMed Central

This study measures hexose monophosphate (HMP) shunt activity, glycolytic rate, and glucose transport in PMN and lymphocytes of patients with glycogen storage disease (GSD) type Ib as compared with controls and with GSD Ia patients. HMP shunt activity and glycolysis were significantly lower in intact PMN cells of GSD Ib patients as compared with GSD Ia patients and with controls. These activities were above normal levels in disrupted GSD Ib PMN. HMP shunt activity and glycolytic rates in lymphocytes were similar in all three groups studied. The rate of 2-deoxyglucose transport into GSD Ib PMN was 30% of that into cells of normal controls. In GSD Ib lymphocytes or in GSD Ia PMN and lymphocytes transport was normal. The striking limitation of glucose transport across the cell membrane of the PMN of GSD Ib patients may account for the impairment of leukocyte function that is characteristic of GSD Ib, but not found in GSD Ia patients.

Bashan, N; Hagai, Y; Potashnik, R; Moses, S W



Interleukin-8 production by polymorphonuclear leukocytes from patients with chronic infected leg ulcers treated with Lactobacillus plantarum.  


Bacterial infection impairs the healing process, promoting the chronicity of inflammation and wounds. Because antibiotics fail to eradicate bacteria, especially in biofilm form, new therapeutic modalities may be required. In the present study, the effectiveness of bacteriotherapy with Lactobacillus plantarum on infected chronic venous ulcers was investigated and its effects on interleukin (IL)-8 production by cells from the ulcer bed and neutrophils isolated from peripheral blood that were previously challenged in vitro with Pseudomonas aeruginosa and L. plantarum were studied. Topical application of L. plantarum culture to lesions (25-60 cm(2)) of 14 diabetic and 20 non-diabetic patients induced debridement, granulation tissue formation and total healing after 30 days in 43% diabetics and in 50% non-diabetics. No significant differences between the groups were observed. The cells from ulcer beds collected after treatment with L. plantarum for 10 days showed a decrease in the percentage of polymorphonuclear, apoptotic and necrotic cells and an enhancement of IL-8 production. IL-8 production by isolated neutrophils from these patients was compared with that in diabetics without ulcers, as well as normal subjects under basal conditions, and after infection of polymorphonuclear cells with P. aeruginosa preincubated either with or without L. plantarum. The basal values in diabetic and ulcer patients were higher than normal (p <0.001) and were increased by P. aeruginosa infection in normal, diabetics (p <0.001) and non-diabetics with ulcers (p <0.01). Preincubation with L. plantarum decreased IL-8 production in patients with ulcers non-diabetic and diabetic (p <0.001). Lactobacillus plantarum treatment reduced wound bacterial load, neutrophils, apoptotic and necrotic cells, modified IL-8 production and induced wound healing. PMID:19519855

Peral, M C; Rachid, M M; Gobbato, N M; Huaman Martinez, M A; Valdez, J C



The crucial role of polymorphonuclear leukocytes in resistance to Salmonella dublin infections in genetically susceptible and resistant mice.  


Macrophages are considered to be the mediators of resistance to extra-intestinal Salmonella infections. Nevertheless, the initial cellular response to Salmonella infections consists primarily of polymorphonuclear leukocytes (PMN). To determine whether PMN serve an important function for the infected host, we made mice neutropenic with the rat mAb to RB6-8C5 and infected them i.v. with approximately 10(3) Salmonella dublin or an isogenic derivative that lacks the virulence plasmid (LD842). We infected BALB/c mice, which have a point mutation in the macrophage-expressed gene Nramp1 that makes them susceptible to Salmonella, and BALB/c.D2 congenic mice, which have the wild-type Nramp1 gene that makes them resistant to Salmonella. Both mouse strains were resistant to LD842, and neutropenia made only the BALB/c strain susceptible to this infection. Neutropenic congenic mice, however, were susceptible only to wild-type S. dublin (plasmid+). These results show a complex interplay between plasmid-virulence genes in Salmonella, host macrophages, and PMN. Mice with normal macrophages need PMN to defend against nontyphoid Salmonella that carry a virulence plasmid but not against Salmonella without virulence plasmids. Mice with a mutant Nramp1 gene need PMN to defend against all Salmonella, even those that lack virulence plasmids. These results, plus the evidence that PMN kill Salmonella efficiently in vitro, suggest that Salmonella have adapted to grow inside macrophages where they are relatively sheltered from PMN. The adaptations that allow Salmonella to survive in macrophages do not protect them from PMN. PMID:9636209

Vassiloyanakopoulos, A P; Okamoto, S; Fierer, J



The crucial role of polymorphonuclear leukocytes in resistance to Salmonella dublin infections in genetically susceptible and resistant mice  

PubMed Central

Macrophages are considered to be the mediators of resistance to extra-intestinal Salmonella infections. Nevertheless, the initial cellular response to Salmonella infections consists primarily of polymorphonuclear leukocytes (PMN). To determine whether PMN serve an important function for the infected host, we made mice neutropenic with the rat mAb to RB6–8C5 and infected them i.v. with ?103 Salmonella dublin or an isogenic derivative that lacks the virulence plasmid (LD842). We infected BALB/c mice, which have a point mutation in the macrophage-expressed gene Nramp1 that makes them susceptible to Salmonella, and BALB/c.D2 congenic mice, which have the wild-type Nramp1 gene that makes them resistant to Salmonella. Both mouse strains were resistant to LD842, and neutropenia made only the BALB/c strain susceptible to this infection. Neutropenic congenic mice, however, were susceptible only to wild-type S. dublin (plasmid+). These results show a complex interplay between plasmid-virulence genes in Salmonella, host macrophages, and PMN. Mice with normal macrophages need PMN to defend against nontyphoid Salmonella that carry a virulence plasmid but not against Salmonella without virulence plasmids. Mice with a mutant Nramp1 gene need PMN to defend against all Salmonella, even those that lack virulence plasmids. These results, plus the evidence that PMN kill Salmonella efficiently in vitro, suggest that Salmonella have adapted to grow inside macrophages where they are relatively sheltered from PMN. The adaptations that allow Salmonella to survive in macrophages do not protect them from PMN.

Vassiloyanakopoulos, Antonis P.; Okamoto, Sharon; Fierer, Joshua



Functional role of mucoid exopolysaccharide (alginate) in antibiotic-induced and polymorphonuclear leukocyte-mediated killing of Pseudomonas aeruginosa.  

PubMed Central

We evaluated in vitro the functional role of mucoid exopolysaccharide (MEP) of Pseudomonas aeruginosa in blocking antibiotic-induced and polymorphonuclear leukocyte (PMN)-mediated pseudomonal killing. The serum-resistant P. aeruginosa isolates used were mucoid strain 144MR and its nonmucoid revertant, strain 144NM. By timed kill curves, early bacterial effects of amikacin against mucoid strain 144MR were substantially less than those observed with nonmucoid strain 144NM; this effect was reversible with enzymatic hydrolysis of MEP of strain 144MR by alginase. Also, early tobramycin uptake (15 to 30 min) by mucoid 144MR cells was less than that seen with nonmucoid strain 144NM; pretreatment of 144MR cells with alginase substantially enhanced early tobramycin uptake compared with untreated 144MR cells (P = 0.08). In strain 144NM (but not in strain 114MR) there was a notable postantibiotic leukocidal enhancement effect manifested by increased nonopsonic killing following brief exposure of these cells to supra-MIC amikacin; pretreatment of strain 144MR with alginase rendered these cells more susceptible to amikacin-induced postantibiotic leukocidal enhancement. Similarly, direct PMN-mediated nonopsonic killing of mucoid strain 144MR was significantly less than that observed with strain 144NM (P less than 0.05); pretreatment of 144MR cells with alginase rendered this strain equal to strain 144NM in susceptibility to nonopsonic killing. In addition, exogenous sodium alginate or extracted MEP of strain 144MR interfered with effective nonopsonic killing of strain 144NM by PMNs. Studies also indicated that mucoid strain 144MR was phagocytosed significantly less well than its nonmucoid mate (P less than 0.00001), an effect reversed by pretreatment of the mucoid cells with alginase. These data confirm that P. aeruginosa MEPs functionally decrease the uptake and early bactericidal effect of aminoglycosides in vitro and interfere with effective PMN-mediated nonopsonic phagocytosis and killing of mucoid strains.

Bayer, A S; Speert, D P; Park, S; Tu, J; Witt, M; Nast, C C; Norman, D C



Thyroid hormone and thyrotropin regulate intracellular free calcium concentrations in human polymorphonuclear leukocytes: in vivo and in vitro studies.  


Intracellular free calcium concentrations (Ca++i) were studied in polymorphonuclear leukocytes (PMNs) from 13 athyreotic patients who had been previously treated by total thyroidectomy and radioiodine therapy for differentiated thyroid carcinoma, and from age- and sex-matched euthyroid healthy controls. Patients were studied twice, when hypothyroid (visit 1) and after restoration of euthyroidism by L-T4 TSH-suppressive therapy (visit 2). PMNs from patients at visit 1 had significantly lower resting (Ca++)i levels compared to both visit 2 and controls. Values at visit 2 did not differ from those of the controls. Stimulus-induced (Ca++)i rise was also significantly blunted at visit 1 and normalized at visit 2, possibly through a differential contribution of distinct intracellular Ca++ stores, as suggested by the response pattern to the chemotactic agent, N-formyl-Met-Leu-Phe (fMLP), to the selective SERCA pump inhibitor, thapsigargine, and to the mitochondrial uncoupler, carbonyl cyanide p-trifluoromethoxyphenyl-hydrazone (FCCP). In vitro treatment of PMNs from healthy subjects with high TSH concentrations impaired intracellular Ca++ store function. Both resting (Ca++)i levels and fMLP-induced (Ca++)i rise increased in the presence either of low-concentration TSH or of T4, but effects of TSH and T4 were not additive. T3, rT3, and TRIAC had no effect. In conclusion, this study provides evidence for a direct relationship between thyroid status and (Ca++)i homeostasis in human PMNs, mainly related to direct actions of TSH and T4 on these cells. PMID:16569353

Marino, F; Guasti, L; Cosentino, M; De Piazza, D; Simoni, C; Bianchi, V; Piantanida, E; Saporiti, F; Cimpanelli, M G; Crespi, C; Vanoli, P; De Palma, D; Klersy, C; Frigo, G M; Bartalena, L; Venco, A; Lecchini, S


Gold complexes and activation of human polymorphonuclear leukocytes. Dissociation of changes in membrane potential and oxidative burst.  


The effects of the gold compounds on the alteration of membrane potential of polymorphonuclear leukocytes (PMN) in response to various stimulants have been compared with their effects on the oxidative burst. The present studies have shown that gold complexes [auranofin (AF), aurothiomalate (Autm), aurocyanide (Au(CN)2-)] have contrasting effects on the membrane potential of 3,3'-dipentyloxacarbocyanine [di-O-C5(3)] loaded PMN. Au(CN)2- at concentrations which inhibit the oxidative burst of PMN did not affect the membrane depolarization after activation of PMN by phorbol myristate acetate (PMA) and N-formyl-methionyl-leucyl phenylalanine (FMLP); Autm slightly stimulated the oxidative burst but had no effect on the depolarization of PMN. In contrast, AF inhibited the depolarization of stimulated PMN to an extent depending upon the concentration of AF, the time of preincubation and the stimulus. The membrane depolarization of PMN caused by PMA, FMLP and concanavalin A (ConA) was inhibited by AF (5 microM) but the depolarization induced by calcium ionophore (A23187) was not affected. AF at the same conditions inhibits the oxidative burst of PMN induced by all these single stimuli including the calcium ionophore. Dissociation of membrane depolarization and superoxide generation caused by AF was also seen in PMN activated by two stimuli. AF (5 microM) had little initial inhibitory effect on the oxidative burst of PMN stimulated by combinations of PMA and ConA or PMA and FMLP whereas it almost totally blocked the depolarization caused by these combinations. Preincubation of cells with 5 microM AF for less than 5 min prior to the addition of PMA allowed membrane depolarization which was followed rapidly by repolarization. None of the gold complexes studied had any effect on the resting membrane potential of PMN. PMID:1417933

Rudkowski, R; Ziegler, J B; Graham, G G; Joulianos, G



A Micro-Mixed Leukocyte Culture Technique using Whole Blood  

Microsoft Academic Search

Micro-mixed leukocyte cultures (MMLC) using whole human peripheral blood were compared with standard macro cultures using separated leukocytes. The MMLC were found to be of comparable sensitivity to the macro test, with the added advantages of requiring less blood and fewer technical manipulations in the performance of the test. One-way MMLC were performed using either X-irradiated whole blood (MMLC-X) or

Gail D. Stockman; D. M. Mumford; J. R. Wilbur



[Role of leukocyte adhesion in filterability of blood cell suspensions].  


We investigated blood cells suspension filterability of 16 donors. The filtration was performed trough 5 microns-pore nuclear filters at constant perfusion pressure 10(5) din/cm2. We estimated also the adherence of leukocytes and platelets on nylon. The adherence of platelets and mononuclear leukocytes reduced the level of the suspension filterability only by 21% (p > 0.05). The presence of nonadhesive polynuclear leukocytes in the suspensions did not change practically their filterability. The addition in the suspensions of adhesive polynuclear leukocytes reduced suspension filterability dramatically. PMID:1421261

Redchits, E G; Parfenov, A S; Rudenko, G R; Sokolovski?, E E; Guzeva, V O; Semavin, I E



Differential effect of exogenous interleukin-10 versus glucocorticoids on gene expression and pro-inflammatory cytokine release by polymorphonuclear leukocytes and monocytes of the newly born  

PubMed Central

Bronchopulmonary dysplasia (BPD) is one of the most common causes of mortality and morbidity in neonatal intensive care units. Persistent inflammation, with an abnormal influx of polymorphonuclear leukocytes (PMNs) followed by monocytes (MONOs), occurs early in the pathogenesis of BPD. Anti-inflammatory therapy with better efficacy and safety than dexamethasone (DEX) is needed. In the present study we determined cell-specific gene expression and cytokine release in response to glucocorticoids versus interleukin-10 (IL-10). Subsequently, we hypothesized that the insensitivity of MONOs to DEX was associated with a failure of the glucocorticoid receptor to translocate to the nucleus. PMNs and MONOs were isolated from umbilical cord blood at birth, and pretreated with PBS vehicle, IL-10 or glucocorticoids prior to endotoxin (LPS)-stimulation for 4 and 18h. Genome-wide gene expressions were determined by microarray and validated by RT-qPCR. Interleukin 8 release in cell culture supernatant was measured by ELISA. To examine the mechanism of monocyte insensitivity to glucocorticoids, nuclear translocation of the glucocorticoid receptor was determined by Western blots. MONOs had 6 times the number of genes changing expression with IL-10 compared to PMNs at 4h. DEX at the therapeutic level for neonates with BPD had no effect on gene expression in MONOs. The order of potency for inhibition of interleukin-8 release from MONOs was IL-10 >betamethasone >dexamethasone and hydrocortisone. Glucocorticoid potency in MONOs was directly related to glucocorticoid receptor translocation to nucleus. Gene expression profiling for IL-10 versus glucocorticoids indicates there may be major differences in therapeutic efficacy for BPD.

Davidson, Dennis; Patel, Hardik; Degoy, Ana C; Gershkovich, Irina; Vancurova, Ivana; Miskolci, Veronika



Differential effect of exogenous interleukin-10 versus glucocorticoids on gene expression and pro-inflammatory cytokine release by polymorphonuclear leukocytes and monocytes of the newly born.  


Bronchopulmonary dysplasia (BPD) is one of the most common causes of mortality and morbidity in neonatal intensive care units. Persistent inflammation, with an abnormal influx of polymorphonuclear leukocytes (PMNs) followed by monocytes (MONOs), occurs early in the pathogenesis of BPD. Anti-inflammatory therapy with better efficacy and safety than dexamethasone (DEX) is needed. In the present study we determined cell-specific gene expression and cytokine release in response to glucocorticoids versus interleukin-10 (IL-10). Subsequently, we hypothesized that the insensitivity of MONOs to DEX was associated with a failure of the glucocorticoid receptor to translocate to the nucleus. PMNs and MONOs were isolated from umbilical cord blood at birth, and pretreated with PBS vehicle, IL-10 or glucocorticoids prior to endotoxin (LPS)-stimulation for 4 and 18h. Genome-wide gene expressions were determined by microarray and validated by RT-qPCR. Interleukin 8 release in cell culture supernatant was measured by ELISA. To examine the mechanism of monocyte insensitivity to glucocorticoids, nuclear translocation of the glucocorticoid receptor was determined by Western blots. MONOs had 6 times the number of genes changing expression with IL-10 compared to PMNs at 4h. DEX at the therapeutic level for neonates with BPD had no effect on gene expression in MONOs. The order of potency for inhibition of interleukin-8 release from MONOs was IL-10 >betamethasone >dexamethasone and hydrocortisone. Glucocorticoid potency in MONOs was directly related to glucocorticoid receptor translocation to nucleus. Gene expression profiling for IL-10 versus glucocorticoids indicates there may be major differences in therapeutic efficacy for BPD. PMID:23390570

Davidson, Dennis; Patel, Hardik; Degoy, Ana C; Gershkovich, Irina; Vancurova, Ivana; Miskolci, Veronika



Chemotactic activity of human polymorphonuclear leukocytes and industrial xenobiotics: a brief review.  


This review deals with some of our contributions to the use of chemotaxis, as a tool in evaluating effects of industrial xenobiotics on PMN, either in vitro or ex vivo. In vitro experiments have shown that lead, arsenic, styrene and 2,5-hexanedione, a major neurotoxicant metabolite of n-hexane, reduce chemotaxis. The most important results of ex vivo experiments have confirmed those obtained in vitro with styrene and 2,5-hexanedione: a significant reduction of chemotaxis was indeed observed in PMN harvested from workers exposed to low levels of n-hexane or styrene who did not show any sign of biochemical or clinical alteration. After 3 weeks under non-exposed conditions, the chemotactic indexes were markedly increased in most of the workers which were exposed to styrene and in all the workers exposed to n-hexane, all of whom have shown a reduced chemotaxis at the first blood sampling. Moreover chemotaxis was found to be significantly reduced at relative low levels of lead: results of the in vitro and ex vivo experiments show a comparable ranking of midpoint inhibition concentrations. We are only at the dawn of the understanding of the relation between occupational xenobiotics and PMN chemotaxis. This research area is still promising for the future, since PMN chemotaxis seems to be adequate and it must therefore enter in well-defined study protocols for investigating the potential immunotoxicity of occupational chemicals to which humans are exposed at low levels. PMID:8059440

Governa, M; Valentino, M; Visonà, I



Roles of p38 MAPK, PKC and PI3-K in the signaling pathways of NADPH oxidase activation and phagocytosis in bovine polymorphonuclear leukocytes  

Microsoft Academic Search

Stimulation of bovine polymorphonuclear leukocytes (PMN) with serum-opsonized zymosan (sOZ) induced the activation of p38 mitogen-activated protein kinase (MAPK), protein kinase C (PKC) and phosphatidylinositol 3-kinase (PI3-K) and sOZ-induced O2? production was significantly attenuated by their inhibitors (SB203580 for p38 MAPK, GF109203X for PKC and wortmannin for PI3-K). They caused significant attenuation of sOZ-induced phosphorylation of p47phox as well. Flow

Tohru Yamamori; Osamu Inanami; Hajime Nagahata; Yu-Dong Cui; Mikinori Kuwabara



Leukotriene B/sub 4/ modulates human polymorphonuclear leukocyte (PMN) phospholipid (PL) methylation  

SciTech Connect

Formation of phosphatidylcholine (PC) by methylation of phosphatidylethanolamine (PE) is required for transduction of chemotactic factor messages in phagocytic cells, including the development of a motile configuration and release of arachidonic acid (AA) from PL. The authors examined PL methylation in human peripheral blood PMN following stimulation by leukotriene B/sub 4/ (LTB/sub 4/), a potent chemotactic lipid metabolite of AA that is produced by human PMN. (/sup 3/H)-methionine, in the presence or absence of LTB/sub 4/, was added directly to the cells without a preloading period. (/sup 3/H)-methyl mono- and di-methylated PE, PC and lyso PC were separated by thin layer chromatography. PL methylation in human PMN is dependent on time of incubation, LTB/sub 4/ concentration and methionine concentration. The optimal LTB/sub 4/ concentration is 10/sup -7/ M, the same concentration that induces a maximal chemotactic response in PMN. At early time points (2-10 min), formation of methylated PL is enhanced following LTB/sub 4/ stimulation. In contrast, at later time points (20-60 min), methylated PL synthesis in LTB/sub 4/ treated cells is depressed or the same as control cells. Synthesis of PC by the choline transferase pathway is not affected by LTB/sub 4/ stimulation. This data shows that, at early time points following LTB/sub 4/ stimulation, PL methylation is enhanced. This correlates with other reports of calcium mobilization and chemotaxis in PMN at early time points following LTB/sub 4/ stimulation.

Bomalaski, J.S.; Clark, M.A.; Dundee, D.



Separation of leukocytes from blood using spiral channel with trapezoid cross-section.  


Inertial microfluidics has recently drawn wide attention as an efficient, high-throughput microfluidic cell separation method. However, the achieved separation resolution and throughput, as well as the issues with cell dispersion due to cell-cell interaction, have appeared to be limiting factors in the application of the technique to real-world samples such as blood and other biological fluids. In this paper, we present a novel design of a spiral inertial microfluidic (trapezoidal cross-section) sorter with enhanced separation resolution and demonstrate its ability in separating/recovering polymorphonuclear leukocytes (PMNs) and mononuclear leukocytes (MNLs) from diluted human blood (1-2% hematocrit) with high efficiency (>80%). PMNs enriched by our method also showed negligible activation as compared to original input sample, while the conventional red blood cell (RBC) lysis method clearly induced artificial activation of the sensitive PMNs. Therefore, our proposed technique would be a promising alternative to enrich/separate sensitive blood cells for therapeutic or diagnostic applications. PMID:23025404

Wu, Lidan; Guan, Guofeng; Hou, Han Wei; Bhagat, Ali Asgar S; Han, Jongyoon



Adhesomes: specific granules containing receptors for laminin, C3bi/fibrinogen, fibronectin, and vitronectin in human polymorphonuclear leukocytes and monocytes  

PubMed Central

We have localized several major extracellular matrix protein receptors in the specific granules of human polymorphonuclear (PMN) and monocytic leukocytes using double label immunoelectron microscopy (IEM) with ultrathin frozen sections and colloidal-gold conjugates. Rabbit antibodies to 67-kD human laminin receptor (LNR) were located on the inner surface of the specific granule membrane and within its internal matrix. LNR antigens co-distributed with lactoferrin, a marker of specific granules, but did not co-localize with elastase in azurophilic granules of PMNs. Further, CD11b/CD18 (leukocyte receptor for C3bi, fibrinogen, endothelial cells, and endotoxin), mammalian fibronectin receptor (FNR), and vitronectin receptor (VNR) antigens were also co- localized with LNR in PMN specific granules. A similar type of granule was found in monocytes which stained for LNR, FNR, VNR, CD18, and lysozyme. Activation of PMNs with either PMA, f-met-leu-phe (fMLP), tumor necrosis factor (TNF), or monocytic leukocytes with lipopolysaccharide (LPS), induced fusion of specific granules with the cell membrane and expression of both LNR and CD18 antigens on the outer cell surface. Further, stimulation led to augmented PMN adhesion on LN substrata, and six- to eightfold increases in specific binding of soluble LN that was inhibited by LNR antibody. These results indicate that four types of extracellular matrix receptors are located in leukocyte specific granules, and suggest that up-regulation of these receptors during inflammation may mediate leukocyte adhesion and extravasation. We have thus termed leukocyte specific granules adhesomes.



Characterization of leukocytes by filtration of diluted blood.  


From the known numbers and properties of normal blood cells, it is apparent that the analysis of the flow of diluted blood, through micropore filters, should allow calculation of the properties of leukocytes without the need for their prior purification. The number and transit time of a "slow" leukocyte population can be deduced by fitting the flow, of diluted blood through 5 microns membranes over about 150 s, to the appropriate mathematical model, which is chosen by the use of a suitable statistical test--the runs test. This population of leukocytes equates numerically with the monocyte population of normal blood; cells have a transit time, through 5 microns pores, of 27.7 +/- 10.9 s. The remaining "fast" leukocytes represent the sum of granulocytes and lymphocytes; their flow properties can be deduced from the measured initial flow rate of diluted blood and the estimated properties of the red blood cells. The properties of the red cells can be assessed from filtration of purified suspensions with any concentration of cells from 0.52 x 10(9)/ml to 3.0 x 10(9)/ml. The transit times for red cells and granulocytes/lymphocytes, in blood diluted with about an equal volume of buffer, are 1.36 +/- 0.17 ms and 1.48 +/- 0.33 s, respectively. The transit times of blood cells, through 5 microns pores, are therefore inversely related to their numbers in blood. PMID:7696635

Adams, R A; Evans, S A; Jones, J G


Effect of Ceftriaxone on Pseudomonas aeruginosa and Staphylococcus aureus in Broth, Serum, and in Combination with Human Polymorphonuclear Leukocytes  

Microsoft Academic Search

We investigated the antibacterial activity of ceftriaxone at concentrations of ¼ × minimum inhibition concentration (MIC), 1 × MIC and 4 × MIC against a serum-resistant Pseudomonas aeruginosa and a serum-resistant Staphylococcus aureus strain in broth, serum, and in combination with leukocytes. Killing effect of ceftriaxone in broth was significantly better than in serum; ceftriaxone improved leukocyte bactericidal activity without

M. Bossier; H. Blaschke; M. Just; F. D. Daschner



Analysis of cytokines and chemokines produced by whole blood, peripheral mononuclear and polymorphonuclear cells.  


Cytokines are immunomodulating proteins involved in cellular communication. The levels of different cytokines reflect the immune capabilities of a person. In literature both whole blood and peripheral blood mononuclear cells (PBMCs) are used, which might lead to different results. The choice between these different sources is not always explained. The goal of our experiments is to determine the cytokine response of whole blood, PBMCs and polymorphonuclear cells (PMNs) after stimulation with lipopolysaccharide (LPS). We used a multiplex analysis to determine a difference in cytokine secretion patterns. In general, PBMCs demonstrated the highest cytokine production and PMNs have an overall low cytokine production. CCL11 and interleukin-23 (IL-23) (and IL-12p40) were exclusively expressed in whole blood. IL-20, VEGF and GM-CSF were expressed only by PBMCs. This difference in expression could be explained by the bioactive components in serum, presence and interaction with granulocytes or platelets in whole blood, the anticoagulant heparin in whole blood and others. The expression of cytokines by cells is dependent on the microenvironment. Different conditions lead to different results. We recommend a thorough examination of the conditions before performing experiments. PMID:23994257

van Dooren, Faas H; Duijvis, Nicolette W; Te Velde, Anje A



Blood leukocyte profile in different phases of menstrual cycle.  


The optimal availability of immune cells in the peripheral blood stream of women plays a critical role in their response to disease and therapeutic interventions. Interaction between the reproductive and immune system plays an important immunoregulatory role. This study was designed to examine the impact of different phases of menstrual cycle on the blood leukocytes. Twenty-four healthy women in their reproductive age group and having regular menstrual cycle were studied during menstrual, proliferative and secretary phases of menstrual cycle. Total leukocyte count, absolute and differential counts of neutrophils, lymphocytes and mixed cells (includes eosinophils, basophils and monocytes) were analyzed. Results showed that the variations in the different types of leukocytes during different phases of menstrual cycle were not statistically significant. No significant inter group difference, except for the significant decrease in differential lymphocyte percentage in proliferative phase as compared to menses were observed. PMID:19130867

Tikare, Swati N; Das, Kusal K; Dhundasi, Salim A


Oxidative Stress in Leukocytes Is a Possible Link Between Blood Pressure, Blood Glucose, and C-Reacting Protein  

Microsoft Academic Search

Because oxidative stress and inflammation are believed to play roles in the pathogenesis of cardiovascular diseases, oxidative stress in polymorphonuclear leukocytes (PMNs) and mononuclear cells (MNCs) has been measured. A total of 529 subjects participated this study. Intracellular oxidative stress in PMNs and MNCs was measured by gated flow cytometry using carboxyfluorescin diacetate bis-acetoxymethyl ester. C-reacting protein (CRP), insulin action

Kenichi Yasunari; Kensaku Maeda; Munehiro Nakamura; Junichi Yoshikawa


Leukocyte transport by red blood cells in a microvessel  

NASA Astrophysics Data System (ADS)

A simulation model is used to study the transport of relatively large, spherical, and stiff white blood cells (leukocytes) by the relatively smaller and highly flexible red cell as they flow in the microcirculation. Their interaction dynamics are thought to be an important component of the inflammation response, in which leukocytes bind to the walls of blood vessels. The red cells are modeled in the simulations as highly deformable three-dimensional shells encasing a Newtonian fluid, and the viscous-flow equation is solved via a boundary integral formulation in which the cell shapes discretized by global spectral basis functions. For slow flow rates, it is found that the leukocyte is predominantly adjacent the vessel walls, whereas for faster flow rates this configuration appears to become unstable and the leukocyte traverses the whole vessel in a seemingly random fashion. For the straight round tubes simulated thus far, the stable leukocyte stand-off distance is always beyond the range of the binding molecules that capture it, which suggests that vessel inhomogeneities or interactions with other white cells are needed to create contact and thereby binding with the vessel walls.

Freund, Jonathan



Peripheral blood leukocyte counts in welcome swallow nestlings.  


It is unclear whether developmental trends in total leukocyte (WBC) and differential lymphocyte (PropL) counts in peripheral blood of altricial birds typically mirror the known ontogenetic increase in immunocompetence. We documented the development of leukocyte and lymphocyte numbers in peripheral blood of wild, altricial Welcome Swallow (Hirundo neoxena) nestlings. Nestlings had a mass-overshoot-recession growth profile. Hatchlings' mean WBC (7.94 x 10(9) cells/l) and PropL (0.65) were respectively 4x and approximately 1.7x the mean adult value. Both variables declined at a steady rate throughout nestling development and were 1.3x the mean adult value at fledging. Hatching WBC values that substantially exceeded those of adults could have reflected the parasite- and pathogen-rich nest environment of this species. The developmental declines in peripheral blood WBC and PropL were not inconsistent with an ontogenetic increase in specific immunocompetence; they are likely to have resulted mainly from an increase in the rate of leukocyte trafficking to vulnerable tissues and organs. PMID:19901398

Sindik, Antonette; Lill, Alan



Computational fluid dynamic studies of leukocyte adhesion effects on non-Newtonian blood flow through microvessels  

Microsoft Academic Search

The study of the effect of leukocyte adhesion on blood flow in small vessels is of primary interest to understand the resistance changes in venular microcirculation. Available computational fluid dynamic studies provide information on the effect of leukocyte adhesion when blood is considered as a homogeneous Newtonian fluid. In the present work we aim to understand the effect of leukocyte

B. Das; P. C. Johnson; A. S. Popel


Granulocyte-macrophage colony-stimulating factor stimulates JAK2 signaling pathway and rapidly activates p93fes, STAT1 p91, and STAT3 p92 in polymorphonuclear leukocytes.  


Granulocyte-macrophage colony-stimulating factor (GM-CSF), supports proliferation, differentiation, and functional activation of hemopoietic cells by its interaction with a heterodimeric receptor. Although GM-CSF receptor is devoid of tyrosine kinase enzymatic activity, GM-CSF-induced peripheral blood polymorphonuclear leukocytes (PMN) functional activation is mediated by the phosphorylation of a large number of intracellular signaling molecules. We have previously shown that JAK2 becomes tyrosine-phosphorylated in response to GM-CSF in PMN. In the present study we demonstrate that also the signal transducers and activators of transcription (STAT) family members STAT1 p91 and STAT3 p92 and the product of the c-fps/fes protooncogene become tyrosine-phosphorylated upon GM-CSF stimulation and physically associated with both GM-CSF receptor beta common subunit and JAK2. Moreover GM-CSF was able to induce JAK2 and p93fes catalytic activity. We also demonstrate that the association of the GM-CSF receptor beta common subunit with JAK2 is ligand-dependent. Finally we demonstrate that GM-CSF induces a DNA-binding complex that contains both p91 and p92. These results identify a new signal transduction pathway activated by GM-CSF and provide a mechanism for rapid activation of gene expression in GM-CSF-stimulated PMN. PMID:8631962

Brizzi, M F; Aronica, M G; Rosso, A; Bagnara, G P; Yarden, Y; Pegoraro, L



The two-step conversion of big endothelin 1 to endothelin 1 and degradation of endothelin 1 by subcellular fractions from human polymorphonuclear leukocytes.  

PubMed Central

The metabolism of big endothelin 1 (bET) and endothelin 1 (ET-1) by subcellular fractions from human polymorphonuclear leukocytes (PMNs) was investigated by bioassay and reversed-phase high-performance liquid chromatography. More than 80% of endothelin-converting activity was recovered from the cytosolic fraction, which in addition to ET-1 generated other peptides from bET. The processing of bET to all its metabolites including ET-1 was prevented by the serine protease inhibitor 3,4-dichloroisocoumarin (DCI; 50 microM) or the elastase inhibitor ONO-5046 (100 microM) but not by phenylmethylsulfonyl fluoride (PMSF; 143 microM), another serine protease inhibitor. Paradoxically, human leukocyte elastase, despite generating a bET fragmentation pattern similar to that of PMN cytosol, produced very little ET-1. However, subsequent treatment of the elastase-derived metabolites of bET with PMN cytosol in the presence of ONO-5046 dramatically increased the amount of ET-1 formed. The generation of ET-1 following this intervention was inhibited by DCI. The PMN membrane preparation degraded ET-1 to a major metabolite, similar to that produced from ET-1 by elastase, and several minor products, paralleled by a loss of its smooth muscle contracting activity. The degradation of ET-1 by PMN microsomes was prevented by DCI, PMSF, or ONO-5046. Our results suggest that an elastase-initiated serine protease cascade is responsible for the sequential conversion of bET to ET-1 by the PMN cytosol. Elastase also partly accounts for the ET-metabolizing properties of PMN microsomes. Images

Kaw, S; Hecker, M; Vane, J R



Angiotensin Type 1a Receptor Signaling Is Not Necessary for the Production of Reactive Oxygen Species in Polymorphonuclear Leukocytes  

PubMed Central

Background. Although angiotensin II (Ang II) has inflammatory effects, little is known about its role in polymorphonuclear leucocytes (PMLs). To elucidate the role of Ang II in PMLs ROS production, we examined hydrogen peroxide (H2O2), one of the ROS, and NO production in AT1a receptor knockout (AT1KO) mice. Methods and Results. PMLs were analyzed from Ang II type 1a receptor knockout mice (AT1KO) and C57BL/6 wild type mice. Using flow cytometry, we studied hydrogen peroxide (H2O2) production from PMLs after Staphylococcus aureus phagocytosis or phorbol myristate acetate (PMA) stimulation. Nitric oxide (NO) production in the AT1KO was low at basal and after phagocytosis. In the AT1KO, basal H2O2 production was low. After PMA or phagocytosis stimulation, however, H2O2 production was comparable to wild type mice. Next we studied the H2O2 production in C57BL/6 mice exposed to Ang II or saline. H2O2 production stimulated by PMA or phagocytosis did not differ between the two groups. Conclusions. AT1a pathway is not necessary for PMLs H2O2 production but for NO production. There was a compensatory pathway for H2O2 production other than the AT1a receptor.

Yamato, Fumiko; Tsuji, Shoji; Hasui, Masafumi; Kaneko, Kazunari



Relationship between zinc malnutrition and alterations in murine peripheral blood leukocytes  

SciTech Connect

Studies using a murine model have shown that the immune system responds rapidly and adversely to zinc deficiency. The extent of alteration of peripheral blood leukocytes (PBL) and immunoglobulin levels were investigated in four zinc dietary groups: zinc adequate (ZA); restricted fed zinc adequate (RZA); marginal zinc deficient (MZD, 72-76% of ZA mouse weight); and severely zinc deficient. The peripheral white blood cell count was 3.66 {plus minus} 1.08 {times} 10{sup 6} cells/ml for ZA mice decreasing by 21%, 28% and 54% for RZA, MZD and SZD mice respectively. An equally dramatic change in the flow cytometric light scatter profile was found. ZA mice had 66% lymphocytes and 21% polymorphonuclear granulocytes (PMN) in their peripheral blood while MZD and SZD mice contained 43% and 30% lymphocytes and 40% and 60% PMNs respectively. Analysis of the phenotypic distribution of specific classes of lymphocytes revealed ZA blood contained 25% B-cells and 40% T-cells (CD5{sup +}). B-cells decreased 40-50% for RZA and MZD mice and 60-70% for SZD mice. The decline in CD5{sup +} T-cells was more modest at 30% and 45% for MZD and SZD mice. A nearly 40% decline in both T{sub h} and T{sub c/s} cells was noted for both MZD and SZD mice. Radioimmunoassay of serum for changes in IgM and IgG content revealed no change among dietary groups while serum zinc decreased 10% for RZA mice and 50% for both MZD and SZD mice. The authors conclude that peripheral blood differential counts in concert with total B and T-cell phenotype may serve as indicators of zinc status while serum zinc and Ig will not.

King, L.E.; Morford, L.A.; Fraker, P.J. (Michigan State Univ., East Lansing (United States))



Reduced redox potential during growth of some gram-negative bacteria. Effect on the biochemical and physicochemical surface properties and phagocytosis by polymorphonuclear leukocytes.  


When cultivated at reduced redox potential the physico-chemical surface properties were altered in strains of E. coli, Salmonella and Yersinia bacteria. In particular, strains which showed hydrophilic surface properties under normal aerobic cultivation became more hydrophobic when exposed to anaerobic conditions (e.g. E. coli K12, E. coli K12D21, E. coli K12D22, S. minnesota S99, S. typhimurium 395MS, S. braenderup 2828 and Yersinia enterocolitica). Moreover, there were qualitative as well as quantitative differences in the protein profiles of whole bacterial lysates and membrane preparations analysed in SDS-PAGE. There were no qualitative differences in the lipopolysaccharide (LPS) bands. However, when E. coli K12D22 were cultivated aerobically, remarkably more high molecular temperature-sensitive (70 degrees C for 45 min) carbohydrate material was produced (weight about 360 KD and 660 KD). Interaction between polymorphonuclear leukocytes (PMNL) and the E. coli K12D22 strain, measured as chemiluminescence, showed that the anaerobically cultivated bacteria induced a chemiluminescence that was mainly of intracellular origin, while the aerobically cultivated induced an extracellular response. Phagocytosis and killing-studies showed that only anaerobically-grown E. coli were effectively inactivated by the PMNL. PMID:2901846

Maluszynska, G M; Magnusson, K E; Stendahl, O; Lock, R; Kniola, B



Myristic Acid, A Side Chain of Phorbol Myristate Acetate (PMA), Can Activate Human Polymorphonuclear Leukocytes to Produce Oxygen Radicals More Potently than PMA  

PubMed Central

Myristic acid (MyA), which is a saturated fatty acid (C14:0) and a side chain of phorbol 12-myristate 13-acetate (PMA), was examined if MyA stimulates human polymorphonuclear leukocytes (PMNs) to release oxygen radicals comparable to PMA by applying electron paramagnetic resonance (EPR)-spin-trapping method. When MyA was added to isolated human PMNs, spin adducts of 5,5-dimethyl-1-pyrroline-N-oxide (DMPO)-OH and DMPO-OOH were time-dependently observed. The amounts of these spin adducts were larger than those of PMNs stimulated by PMA. These results clearly show that MyA is more potent agent to prime human PMNs than PMA, in a point of view of not only O2·? but also ·OH production. This fact calls attention that too much intake of MyA that is known to be contained vegetable oils can lead to crippling effect through uncontrolled production of reactive oxygen species.

Tada, Mika; Ichiishi, Eiichiro; Saito, Rumiko; Emoto, Natsumi; Niwano, Yoshimi; Kohno, Masahiro



Human polymorphonuclear leukocyte chemotaxis as a tool in detecting biological early effects in workers occupationally exposed to low levels of n-hexane.  


Human polymorphonuclear leukocytes (PMN) were chosen to measure two cellular end points--chemotaxis and respiratory burst--and to verify whether they could function as biomarkers of early effect in detecting occupational exposure to n-hexane of apparently healthy shoe workers, without any electroneuromyographic (ENMG) abnormality. Chemotaxis, but not respiratory burst, was found to be impaired. A negative linear correlation between chemotaxis of PMN of those workers that had been exposed to n-hexane versus 2,5-hexanedione (2,5-HD) urinary concentrations were found. This negative trend is consistent with our previous in vitro experimental findings: it was observed that the progressive addition of 2,5-HD to PMN suspensions inhibited chemotaxis in a dose-dependent mode, while chemiluminescence was not modified. Now we have confirmed in vivo that chemotaxis is more sensitive than the respiratory burst response to 2,5-HD. Such results justify the interest in the behaviour of PMN harvested from workers exposed to n-hexane. Since significant inhibition of chemotactic activity was observed in some workers whose urinary 2,5-HD levels were lower than 5 mg l-1, which is the biological exposure index suggested by ACGIH, this study suggests that PMN chemotaxis may be proposed as a useful biomarker in detecting occupational exposure to low level of n-hexane. PMID:7826683

Governa, M; Valentino, M; Visonà, I; Monaco, F



Influence of polyclonal immunoglobulins on the polymorphonuclear leukocyte response to lipopolysaccharide of Salmonella enteritidis as measured with luminol-enhanced chemiluminescence.  

PubMed Central

In gram-negative sepsis, the activation of polymorphonuclear leukocytes (PMN) by lipopolysaccharide (LPS) and the resulting production of superoxide and other oxygen radicals may be an important cause of tissue damage. A suppression of the PMN response to LPS stimulation would be therapeutically beneficial. The aim of this study was to determine whether different polyclonal immunoglobulins (Igs; 5S-Ig, 7S-Ig, and 19S-Ig) influence the PMN response to LPS of Salmonella enteritidis in vitro. The respiratory burst activity of PMN was measured with luminol-enhanced chemiluminescence. After addition of a 5S-Ig solution containing F(ab')2 fragments of IgG and a 19S-Ig solution containing 12% polyclonal IgM, luminol-enhanced chemiluminescence was reduced by 27% (P < 0.05) and 46% (P < 0.005), respectively. However, after addition of a 7S-Ig solution containing polyclonal IgG, luminol-enhanced chemiluminescence was increased fourfold (P < 0.05). The results suggest that the influence of polyclonal Igs on PMN response to LPS stimulation is dependent on the Ig class, F(ab')2 fragments of IgG and IgM leading to LPS neutralization and IgG leading to the production of potentially toxic oxygen radicals.

Wagner, D R; Heinrich, D



In vitro effects of aqueous seeds extract of Acacia cyanophylla on the opsonized zymosan-induced superoxide anions production by rat polymorphonuclear leukocytes.  


In vitro studies were carried out in rat pleural polymorphonuclear leukocytes (PMNs) activated by opsonized zymosan (OZ) to investigate the effects of aqueous extract from Acacia cyanophylla seeds on superoxide anions generation. PMNs were collected, after induction of an acute inflammatory reaction, by injection in the rat pleural cavity, of a suspension of calcium pyrophosphate (CaPP) crystals (pleurisy with CaPP) or serum (pleurisy with serum). The results obtained indicate that Acacia cyanophylla aqueous seeds extract had, in vitro, a significant stimulatory effect, in a dose dependent manner, on the PMN superoxide anions generation. It also corrected the diminution of superoxide anions production induced by diclofenac pre-treated PMNs. It could be concluded from the results of this study that the stimulatory properties of Acacia cyanophylla seeds aqueous extract may at least be due to the presence of polyphenols such tannins and/or lignins. Further investigations are needed to determine clearly the mechanisms mediating the generation of superoxide radicals in this phenomenon. PMID:15036483

El Abbouyi, Ahmed; Toumi, Mina; El Hachimi, Youssef; Jossang, Akino



C3a activates the respiratory burst in human polymorphonuclear neutrophilic leukocytes via pertussis toxin-sensitive G-proteins.  


In contrast to C5a, which represents a well-established potent activator of the respiratory burst in polymorphonuclear neutrophilic granulocytes (PMN), the functional role of C3a in the activation of PMN is, so far, poorly understood. Herein, the potential role of human C3a in the activation of the respiratory burst in human PMN was investigated. The release of reactive oxygen species (ROS) of PMN from healthy donors was measured by lucigenin-dependent chemiluminescence. C3a dose-dependently induced the production of ROS in human PMN in the range between 10 ng/mL and 1,000 ng/mL, whereas C3a-desArg was inactive. Flow cytometric measurement of H2O2 by dihydrorhodamine-123 labeling of anti-CD16-stained PMN showed that predominantly neutrophilic PMN are responsible for the C3a-induced activation of the respiratory burst. To exclude that C3a stimulation was caused by contamination with C5a, the specificity of C3a-induced activation of PMN was shown using monoclonal antibodies (MoAbs). Accordingly, the effect of C3a was completely abolished in the presence of Fab fragments of a blocking anti-C3a MoAb. In addition, blockade of the C5a receptor by the anti-C5a receptor (anti-C5aR) MoAb, S5/1, totally inhibited the C5a-induced production of ROS, whereas the C3a response in the presence of the anti-C5aR MoAb was unaffected. The specificity of the response was further confirmed by homologous desensitization after restimulation with C3a. In contrast, no cross-desensitization was observed upon stimulation with C5a. The C3a-induced ROS production by PMN was inhibited by pertussis toxin, indicating the involvement of guanine nucleotide-binding proteins (Gi proteins) in the signal transduction process initiated by C3a. In addition, stimulation of PMN by C3a resulted in a transient increase in the cytosolic free calcium concentration ([Ca2+]i) in a dose-dependent manner. In contrast to C3a-induced ROS production, C3a did not induce a chemotactic response in PMN, indicating functional qualitative differences as compared with C5a. In summary, these results show that C3a is a potent activator of the respiratory burst in human PMN. Therefore, these findings point to a novel role of C3a in the pathogenesis of inflammatory diseases associated with increased C3a levels and PMN activation. PMID:8193368

Elsner, J; Oppermann, M; Czech, W; Kapp, A



Guidance for Industry: Pre-Storage Leukocyte Reduction of Whole Blood and Blood Components Intended for Transfusion.  

National Technical Information Service (NTIS)

We, FDA, are issuing this guidance document to provide you, blood establishments, with recommendations for pre-storage leukocyte reduction of Whole Blood and blood components intended for transfusion, including recommendations for validation and quality c...



Protein nitration in cutaneous inflammation in the rat: essential role of inducible nitric oxide synthase and polymorphonuclear leukocytes  

PubMed Central

We have examined the relationship between neutrophil accumulation, NO• production and nitrated protein levels in zymosan-mediated inflammation in rat skin in vivo. Rats were anaesthetized and cutaneous inflammation was induced by zymosan (injected intradermally, i.d.). Experiments were carried out up to 48 h, in recovery procedures as appropriate. Assays for neutrophil accumulation (measurement of myeloperoxidase), nitric oxide (assessment of NO2?/NO3?) and nitrated proteins (detected by ELISA and Western blot) were performed in skin extracts. The results demonstrate a close temporal relationship between these parameters. Samples were assayed at 1, 4, 8, 24 and 48 h after i.d. injection of zymosan. The highest levels measured of each parameter (P<0.001 compared with vehicle) were found at 4–8 h, with a reduction towards basal levels by 24 h. Selective depletion of circulating neutrophils with anti-neutrophil antibody abolished neutrophil accumulation and protein nitration. In addition substantially decreased NO levels were found. A selective inducible nitric oxide synthase (iNOS) inhibitor, N-3-aminomethyl-benzyl-acetamidine-dihydrochloride (1400W) also significantly reduced neutrophil levels and NO production and substantially inhibited protein nitration. We conclude that the neutrophil leukocyte plays an essential role in the formation of iNOS-derived NO and nitrated proteins in inflammation, in a time-dependent and reversible manner. The NO-derived iNOS also has a role in stimulating further neutrophil accumulation into skin. This suggests a close mechanistic coupling between neutrophils, NO production and protein nitration.

Greenacre, S A B; Rocha, F A C; Rawlingson, A; Meinerikandathevan, S; Poston, R N; Ruiz, E; Halliwell, B; Brain, S D



Derivative of wheat germ agglutinin specifically inhibits formyl-peptide-induced polymorphonuclear leukocyte chemotaxis by blocking re-expression (or recycling) of receptors  

SciTech Connect

The mechanism of action of a derivative of wheat germ agglutinin (WGA-D) which specifically and irreversibly inhibits N-formyl-methionyl-leucyl-phenylalanine (FMLP)-induced polymorphonuclear leukocyte (PMN) chemotaxis was examined. At a concentration that completely inhibited PMN chemotaxis, WGA-D had no effect on either the uptake or release of (/sup 3/H)-FMLP by PMN. Similarly, WGA-D did not affect either the short-term binding to, or internalization by, PMN of a fluoresceinated FMLP analog. WGA-D did interfere, however, with the re-expression (or recycling) of FMLP receptors by PMN that had been preincubated with 1 FMLP for 10 min at 4/sup 0/C. This effect was specific for WGA-D, because it was not observed when concanavalin A was used. Scatchard plot analysis of FMLP binding to PMN after receptor re-expression demonstrated that WGA-D-treated PMN had a significant diminution in the number of high affinity receptors. WGA-D-mediated inhibition of FMLP receptor re-expression was associated with inhibition of FMLP-induced PMN chemotaxis, but had no effect on either FMLP-induced PMN superoxide anion generation or degranulation. Studies using (/sup 12/%I)-WGA-D demonstrated that PMN did not internalize WGA-D spontaneously. The data indicate that WGA-D perhaps by binding to the FMLP receptor, inhibits FMLP-induced PMN chemotaxis by blocking the re-expression (or recycling) of a population of receptors required for continuous migration.

Perez, H.D.; Elfman, F.; Lobo, E.; Sklar, L.; Chenoweth, D.; Hooper, C.



Abnormalities of polymorphonuclear leukocyte function associated with a heritable deficiency of high molecular weight surface glycoproteins (GP138): common relationship to diminished cell adherence.  

PubMed Central

Investigations of polymorphonuclear leukocyte (PMN) function were performed in a 5-yr-old white female with delayed umbilical cord separation, impaired pus formation, and a severe defect of PMN chemotaxis. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis demonstrated an almost total deficiency of a high molecular weight glycoprotein(s) (GP138) in the granule and membrane fractions of the patient's cells, and NaB3H4-galactose oxidase labeling demonstrated the absence of a major glycoprotein complex on the surface of her PMNs. Monoclonal antibodies (MAb) were employed in flow cytometry experiments to demonstrate that two previously characterized glycoproteins (Mo1 and LFA1) were undetectable on the surface of the patient's PMNs and monocytes. Immunoprecipitation of 125I-labeled patient cells with subunit specific MAbs confirmed that the alpha-subunits of Mo1 (155 kD) and LFA1 (177 kD) and their common beta-subunit (94 kD) were totally deficient. Functional analyses of patient PMNs demonstrated severe impairment of adherence- and adhesion-dependent cell functions including spreading, aggregation, orientation in chemotactic gradients, antibody-dependent cellular cytotoxicity, and phagocytosis of particles (Oil-Red-0-paraffin, zymosan) selectively opsonized with C3-derived ligands. Patient PMNs demonstrated a normal capacity to rosette with IgG or C3b-coated sheep erythrocytes, but rosette formation with C3bi-coated erythrocytes was profoundly diminished. Adhesion-independent functions including shape change, N-formyl-methionyl-leucyl-3H-phenylalanine binding, and O-2 generation or secretion elicited by soluble stimuli were normal. Membrane fluidity, surface charge, and microtubule assembly were also normal. These findings provide new evidence that critical PMN surface glycoproteins are required to facilitate multiple adhesion-dependent cellular functions of the inflammatory response. Images

Anderson, D C; Schmalstieg, F C; Arnaout, M A; Kohl, S; Tosi, M F; Dana, N; Buffone, G J; Hughes, B J; Brinkley, B R; Dickey, W D



?3/4 Fucosyltransferase 3-Dependent Synthesis of Sialyl Lewis A on CD44 Variant Containing Exon 6 Mediates Polymorphonuclear Leukocyte Detachment from Intestinal Epithelium during Transepithelial Migration.  


Polymorphonuclear leukocyte (PMN) migration across the intestinal epithelium closely parallels disease symptoms in patients with inflammatory bowel disease. PMN transepithelial migration (TEM) is a multistep process that terminates with PMN detachment from the apical epithelium into the lumen. Using a unique mAb (GM35), we have previously demonstrated that engagement of the CD44 variant containing exon 6 (CD44v6) blocks both PMN detachment and cleavage of CD44v6. In this article, we report that PMN binding to CD44v6 is mediated by protein-specific O-glycosylation with sialyl Lewis A (sLe(a)). Analyses of glycosyltransferase expression identified fucosyltransferase 3 (Fut3) as the key enzyme driving sLe(a) biosynthesis in human intestinal epithelial cells (IECs). Fut3 transfection of sLe(a)-deficient IECs resulted in robust expression of sLe(a). However, this glycan was not expressed on CD44v6 in these transfected IECs; therefore, engagement of sLe(a) had no effect on PMN TEM across these cells. Analyses of sLe(a) in human colonic mucosa revealed minimal expression in noninflamed areas, with striking upregulation under colitic conditions that correlated with increased expression of CD44v6. Importantly, intraluminal administration of mAb GM35 blocked PMN TEM and attenuated associated increases in intestinal permeability in a murine intestinal model of inflammation. These findings identify a unique role for protein-specific O-glycosylation in regulating PMN-epithelial interactions at the luminal surface of the intestine. PMID:24068663

Brazil, Jennifer C; Liu, Renpeng; Sumagin, Ronen; Kolegraff, Keli N; Nusrat, Asma; Cummings, Richard D; Parkos, Charles A; Louis, Nancy A



Polymorphonuclear Leukocyte Accumulation in Brain Regions with Low Blood Flow during the Early Postischemic Period.  

National Technical Information Service (NTIS)

In an anesthetized canine model in which ischemia was induced by incremental air embolism, 16 animals were exposed to 1 hr of ischemia and monitored for 10 min (n = 4), 60 min(n = 6), or 240 min (n = 6). Fourteen animals were observed for corresponding pe...

J. M. Hallenbeck A. J. Dutka T. Tanishima P. M. Kochanek K. K. Kumaroo



Continuous infusion of Escherichia coli endotoxin in vivo primes in vitro superoxide anion release in rat polymorphonuclear leukocytes and Kupffer cells in a time-dependent manner.  

PubMed Central

Continuous infusion of a nonlethal dose of Escherichia coli lipopolysaccharide (LPS) (0.5 mg/kg) induced early (3 h) accumulation of polymorphonuclear leukocytes (PMNL) in rat liver followed by later (30 h) greater extravasation of mononuclear phagocytes (MNP) (E. B. Rodriguez de Turco and J. A. Spitzer, J. Leukocyte Biol. 48:488-494, 1990). Nonparenchymal liver cells from rats treated for 3 and 30 h with LPS were recovered by centrifugal elutriation, yielding a 23-ml/min fraction (endothelial cells) and a 45-ml/min fraction (PMNL, Kupffer cells, and MNP), and compared for their capacity for basal and agonist-stimulated superoxide (O2-) production. Stimulation with phorbol myristate acetate and opsonized zymosan caused a dose-dependent release of O2- from the 45-ml/min fraction derived from rats treated for 3 h with saline, but not from the 23-ml/min fraction. Further purification of the 45-ml/min fraction by discontinuous density gradient centrifugation into a Kupffer and a PMNL fraction revealed that most of the agonist-induced O2- release was generated by infiltrating PMNL at this early time point of LPS infusion. By 30 h of LPS infusion, although enhancement of the phorbol-12-myristate-13-acetate- and opsonized zymosan-stimulated release of O2- was observed in the 45-ml/min fraction, but not in the 23-ml/min fraction, the maximum release of O2- was smaller than that observed in the rats treated for 3 h. Our results support the following conclusions: (i) after a 3-h LPS infusion, PMNL found in the liver in increased numbers are also highly primed for agonist-stimulated release of O2-, while Kupffer cell priming is of a lesser extent; (ii) after a 30-h infusion of LPS, infiltrating MNP found in the liver in increased numbers are primed for agonist-induced O2- release, while priming of PMNL has diminished; (iii) at both 3 and 30 h of LPS infusion, liver endothelial cells are not significantly primed for agonist-stimulated O2- release; and (iv) in vivo priming by LPS infusion at both 3 and 30 h was not reversed by the experimental method used for cell recovery (ca. 3 h), thus suggesting that in vivo LPS priming of O2- release may ultimately lead to severe impairment of liver function and metabolism observed during endotoxemia and sepsis if not therapeutically blocked at an early time point.

Mayer, A M; Spitzer, J A



Computational fluid dynamic studies of leukocyte adhesion effects on non-Newtonian blood flow through microvessels.  


The study of the effect of leukocyte adhesion on blood flow in small vessels is of primary interest to understand the resistance changes in venular microcirculation. Available computational fluid dynamic studies provide information on the effect of leukocyte adhesion when blood is considered as a homogeneous Newtonian fluid. In the present work we aim to understand the effect of leukocyte adhesion on the non-Newtonian Casson fluid flow of blood in small venules; the Casson model represents the effect of red blood cell aggregation. In our model the blood vessel is considered as a circular cylinder and the leukocyte is considered as a truncated spherical protrusion in the inner side of the blood vessel. The cases of single leukocyte adhesion and leukocyte pairs in positions aligned along the same side, and opposite sides of the vessel wall are considered. The Casson fluid parameters are chosen for cat blood and human blood and comparisons are made for the effects of leukocyte adhesion in both species. Numerical simulations demonstrated that for a Casson fluid with hematocrit of 0.4 and flow rate Q = 0.072 nl/s, a single leukocyte increases flow resistance by 5% in a 32 microns diameter and 100 microns long vessel. For a smaller vessel of 18 microns, the flow resistance increases by 15%. PMID:11026943

Das, B; Johnson, P C; Popel, A S



Evaluation of antitularaemia immunity via whole human blood leukocyte cytofluorometric analysis  

NASA Astrophysics Data System (ADS)

Tularaemia is followed by development of a long-lasting protective immunity. Peripheral blood leukocytes of vaccinated individuals partially lysed in vitro after exposure to killed Francisella tularensis cells and this leukocytolis reaction is used for evaluation of antitularaemia immunity. Here, results from cytofluorometric characterization of human whole blood leukocyte response following exposure to F. tularensis are reported. Leukocytes were stained in whole blood by acridine orange. The green (nuclear chromatin) and red (lysosomal granules) fluorescent signals from individual cells were measured by flow cytometry to detect the decrease of leukocyte concentration in whole human blood (the leukocytolis intensity) and to calculate the damaged leukocytes with changed chromatin structure and lysosomal granules per cell content. Our data indicate that flow cytometry offers a rapid and more informative technique for evaluation of human antitularaemia immunity in vitro.

Firstova, Victoria V.; Kravtsov, Alexander L.; Schmelkova, Tatyana P.; Shchukovskaya, Tatyana N.



Obesity related methylation changes in DNA of peripheral blood leukocytes  

PubMed Central

Background Despite evidence linking obesity to impaired immune function, little is known about the specific mechanisms. Because of emerging evidence that immune responses are epigenetically regulated, we hypothesized that DNA methylation changes are involved in obesity induced immune dysfunction and aimed to identify these changes. Method We conducted a genome wide methylation analysis on seven obese cases and seven lean controls aged 14 to 18 years from extreme ends of the obesity distribution and performed further validation of six CpG sites from six genes in 46 obese cases and 46 lean controls aged 14 to 30 years. Results In comparison with the lean controls, we observed one CpG site in the UBASH3A gene showing higher methylation levels and one CpG site in the TRIM3 gene showing lower methylation levels in the obese cases in both the genome wide step (P = 5 × 10-6 and P = 2 × 10-5 for the UBASH3A and the TRIM3 gene respectively) and the validation step (P = 0.008 and P = 0.001 for the UBASH3A and the TRIM3 gene respectively). Conclusions Our results provide evidence that obesity is associated with methylation changes in blood leukocyte DNA. Further studies are warranted to determine the causal direction of this relationship as well as whether such methylation changes can lead to immune dysfunction. See commentary:



A computational study of the effects of leukocyte adhesion on non-Newtonian blood flow in microvessels  

Microsoft Academic Search

The study of the effects of leukocyte adhesion on blood flow in microvessels is of significant interest for understanding the resistance changes in microcirculation. Several computational fluid dynamic studies have been done to understand flow resistance and drag forces due to adhering leukocytes by considering blood as a Newtonian fluid. In the present work we investigate the effect of leukocyte

Bigyani Das; Aleksander S. Popel



Blood Memo - Collection of Human Leukocytes for Furter ...  

Center for Biologics Evaluation and Research (CBER)

Text Version... of Source Leukocytes before they are shipped for ... available prior to shipping, the procedures to ... establishments desiring to collect and ship Source ... More results from


Effect of glucose concentration, osmolality, and sterilization process of peritoneal dialysis fluids on cytokine production by peripheral blood mononuclear cells and polymorphonuclear cell functions in vitro  

Microsoft Academic Search

We sought to investigate the effects of high glucose concentration, osmolality, and heat sterilization of peritoneal dialysis fluids on tumor necrosis factor-alpha (TNF-alpha) production by peripheral blood mononuclear cells (PBMC) and polymorphonuclear cell (PMN) functions. Blood samples were obtained from eight healthy volunteers. PBMCs and PMNs were harvested by centrifugation with Ficoll-Hypaque (Sigma, St Louis, MO). PBMC were incubated with

M Cendoroglo; S Sundaram; BL Jaber; BJ Pereira



LeukoCatch, a quick and efficient tool for the preparation of leukocyte extracts from blood  

PubMed Central

Background Whole-protein extracts from peripheral blood leukocytes are ideal for basic and clinical research. However, lack of a simple preparation technique has limited the use of such extracts. The aim of this study is to develop a simple and easy system that can selectively obtain leukocyte extracts without hemoglobin. Methods A filter that captures the leukocytes but not RBCs was set at the bottom of a 10-mL medical syringe by sandwiching it between plastic stoppers. The capturing efficiency of leukocytes with this tool, called LeukoCatch, was examined using human macrophage cells (MONO-MAC-6). The abilities of LeukoCatch system to capture the leukocyte proteins and to remove the hemoglobin from RBCs were tested by western blot analysis using human blood samples. Results This study presents the development of LeukoCatch, a novel tool that allows the preparation of leukocyte extracts from blood samples within 3 min without centrifugation. Tissue-cultured human macrophage cells were tested to determine the optimal filter numbers and pass-through frequencies of LeukoCatch, which was then applied to 2-mL blood samples. Samples were passed 2~5 times through a LeukoCatch equipped with 5 filters, washed twice with phosphate-buffered saline for red cell removal, and leukocyte proteins were extracted with 0.5 mL of elution buffer. Western blot analysis of the purified extract indicated that more than 90% of hemoglobin was removed by the LeukoCatch and that the protein recovery rate of leukocytes was at least 4 times better than that of the conventional centrifugation method. Conclusion We conclude that LeukoCatch is useful not only for diagnosis at the bedside but also for basic research using blood samples or tissue culture cells.



Effect of an Intramuscular Injection of Human Leukocyte Interferon on Blood Leukocyte Counts and Proportions of Lymphocytes Forming E, EA and EAC Rosettes  

Microsoft Academic Search

Following an intramuscular injection of human leukocyte interferon blood leukocyte counts decreased in most patients as tested 24 h after the injection. This decrease comprised lymphocytes, monocytes and granulocytes. 24 h after interferon injection, the proportion of lymphocytes forming E rosettes was increased in most patients, whereas the proportion of lymphocytes forming EA and EAC rosettes was not significantly changed.Copyright

Stefan Einhorn; Henric Blomgren; Hans Strander



Changes in peripheral blood leukocyte populations in pigs with naturally occurring exudative epidermitis  

Microsoft Academic Search

The objective of the study was to analyze changes in peripheral blood leukocyte subsets in cases of naturally occurring exudative epidermitis (EE) in pigs. Five of ten piglets developed the chronic clinical form of EE 2–5 days after weaning (PW). Blood samples were obtained at 7, 14 and 21 days from both normal and clinically affected piglets for routine haematology

M. Nofrarías; J. Pujols; J. Segalés; X. Gibert; N. Majó



Improved survival of newborns receiving leukocyte transfusions for sepsis  

SciTech Connect

To determine the role of polymorphonuclear (PMN) leukocyte transfusions in neonates with sepsis, 23 consecutive newborns were prospectively randomly selected during an 18-month period in a treatment plan to receive polymorphonuclear leukocyte transfusions with supportive care or supportive care alone. Thirteen neonates received transfusions every 12 hours for a total of five transfusions. Each transfusion consisting of 15 mL/kg of polymorphonuclear leukocytes was subjected to 1,500 rads of radiation. The polymorphonuclear leukocytes were obtained by continuous-flow centrifugation leukapheresis and contained 0.5 to 1.0 X 10(9) granulocytes per 15 mL with less than 10% lymphocytes. Positive findings on blood cultures were obtained in 14/23 patients and seven were randomly selected for each treatment group. Absolute granulocyte counts were less than 1,500/microL in 13 patients but tibial bone marrow examinations revealed that the neutrophil supply pool was depleted in only three patients. The survival was significantly greater in the treatment group compared with the group that did not receive transfusions.

Cairo, M.S.; Rucker, R.; Bennetts, G.A.; Hicks, D.; Worcester, C.; Amlie, R.; Johnson, S.; Katz, J.



Cryopreservation of blood mononuclear leukocytes and stem cells suspended in a large fluid volume  

Microsoft Academic Search

Summary It was the purpose of this study to establish and evaluate a freezing and-thawing method for preservation of hemopoietic stem cells from the peripheral blood. Blood leukocytes collected by means of an IBM Blood-Cell-Separator were frozen in plastic bags using 10% DMSO and controlled cooling rates. Thawing was performed rapidly, and DMSO was diluted and removed prior to the

T. M. Fliedner; M. Körbling; W. Calvo; Ch. Bruch; E. Herbst



Rapid isolation of leukocyte subsets from fresh and cryopreserved peripheral blood mononuclear cells in clinical research.  


Isolation and processing blood into leukocyte subsets are important processes in research. Although methods have been developed to fractionate small volumes of blood, optimizing the methods and balancing the underlying costs are often necessary. The need for such optimization is particularly critical when processing larger volumes of blood. We describe a simple and reproducible method for processing larger volumes of fresh blood rapidly and consistently, which yields peripheral blood mononuclear cells (PBMCs) and leukocyte subsets with high purity (81-96%; n=13) and higher yields relative to stored blood. RNA isolated from these cells was found to be suitable for downstream applications. Blood stored for 24 hours (n=4) before processing resulted in significantly lower yields of PBMCs (58 percent lower), T cells (52 percent lower), B cells (21 percent lower) and monocytes (25 percent lower) compared to fresh blood. However, the purity of the fractionated cells was comparable to that obtained with fresh blood. Furthermore, we report that the yield and purity of the leukocyte subsets isolated from cryopreserved PBMCs (n=4) were not compromised. PMID:23224370

Arimilli, S; Damratoski, B E; Chen, P; Jones, B A; Prasad, G L


Significance of leukocyte concentration in the performance of the quantitative cytomegalovirus (CMV) antigenemia assay  

Microsoft Academic Search

Background: The quantitative cytomegalovirus-antigenemia (CMV-Ag) assay is an important technology in the regimen of tests utilized in the care and management of acquired immunodeficiency syndrome (AIDS) and other immunocompromised patient groups. Performance of this assay is contingent upon the appropriate processing of the polymorphonuclear leukocyte (PMNL) compartment of the peripheral blood. However, a cell input standard in the performance of

Steven M Lipson; Ana Toro; Madhavi Lotlikar; Mark E Match; Mark H Kaplan; David H Shepp; Jerry Gong



[Effect of different altitudes on telomere length of rat peripheral blood leukocyte].  


The present study was aimed to investigate the effect of different altitudes on telomere length of rat peripheral blood leukocyte and possible mechanism. Sixty male rats were randomly divided into three groups, lower altitude control group (10 m), moderate altitude group (2 260 m) and very high altitude group (simulated 5 000 m). The moderate altitude group and very high altitude group rats were transported to Xining and hypobaric chamber in Qinghai University, respectively. The peripheral blood specimens were extracted 30 d after the transportation. By means of real-time PCR, automatic blood cell analyzer, ELISA, TBA and WST-1 methods, the telomere lengths of blood leukocyte, the hemoglobin (Hb) contents, the plasma levels of telomerase reverse transcriptase (TERT) and hypoxia-inducible factor 1? (HIF-1?), the plasma content of malondialdehyde (MDA) and superoxide dismutase (SOD) activity were measured, respectively. The results showed that the telomere lengths of peripheral blood leukocyte in moderate altitude group were longer than those in control group and very high altitude group. The changes of TERT were compatible with the telomere length of peripheral blood leukocyte under different altitudes. The levels of HIF-1? in moderate altitude group and very high altitude group were higher than that of control group. The very high altitude group showed decreased SOD activities and increased level of MDA, compared with the other two groups. These results suggest that the telomere lengths of rat peripheral blood leukocyte in moderate altitude are elongated, and that the telomere-elongating effect is lost under very high altitude. The changes of HIF-1?, TERT and oxidative stress damage are the main mechanisms of telomere length changes. Moderate altitude living might be beneficial to increasing the life span in mammals. PMID:24129736

Wang, Ya-Ping; Yang, Ying-Zhong; Ma, Lan; Zhao, Yan-Xia; Ge, Ri-Li



[Selectivity of accumulation of chlorine e6 derivatives in blood leukocytes].  


The uptake of chlorine e6 derivatives by peripheral blood leukocytes was studied using a fluorescence-activated sorter. The analysis showed that the order of pigment uptake by leukocyte populations is the following: granulocytes > or = monocytes > lymphocytes. It was found that the accumulation of the pigments in the cell significantly varied. The level of chlorine e6 dimethyl ester accumulated by cells was found to be 15 times higher than that of chlorine e6. It was assumed that the differences in pigment uptake by different types of blood cells are due to structural and morphological features of leukocytes. The data obtained may be useful in developing the new methods of photodynamic therapy. PMID:12630115

Savitski?, V P; Zorin, V P


Analysis of blood processing conditions to obtain high-quality total RNA from human leukocyte concentrate.  


Blood samples are used as a biological source to discover biomarkers of hematological and non-hematological disorders. The present study shows the impact of different experimental conditions associated with cell lysis buffer, TRI-reagent protocol and blood cell storage buffer and their correlation with the quantity, quality and Adrenomedullin gene expression levels of total RNA when RT-PCR technique is used. A leukocyte cell bank protocol is also proposed for further mRNA expression analysis using RNAlater as storage buffer. There is evidence that total RNA isolated from leukocyte concentrate stored for 1 month at -70 degrees C did not show significant differences concerning quality, purity and Adrenomedullin gene expression compared with the freshly processed leukocyte sample. PMID:17573660

Pérez, I A G; Santana, S P; Argudin, T D; Gardon, D O P




Microsoft Academic Search

Ichthyohematology is a common tool in the clinic analysis of economically important fish, and used less to understand fish physiology in relation to the environment. In this context and because of the scarce information about the condrictian hematology, we characterize the leukocytes of the Chilean catshark Schroederichtys chilensis and determine the quan- titative reference values. Blood was obtained from fish

Ariel Valenzuela; Ciro Oyarzún; Víctor Silva


Leukocyte-depletion of blood components does not significantly reduce the risk of infectious complications  

Microsoft Academic Search

Allogeneic blood transfusions are claimed to be an independent risk factor for postoperative infections in open colorectal surgery due to immunomodulation. Leukocyte-depletion of erythrocyte suspensions has been shown in some open randomized studies to reduce the rate of postoperative infection to levels observed in nontransfused patients. Using a double-blinded, randomized design, we studied the postoperative infection rate in patients undergoing

Ingrid Louise Titlestad; Liselotte S. Ebbesen; Alan P. Ainsworth; Søren T. Lillevang; Niels Qvist; Jørgen Georgsen




Technology Transfer Automated Retrieval System (TEKTRAN)

The composition and functional capacity of peripheral blood mononuclear leukocyte populations from dairy cows are altered substantially during the peripartal period. These changes are associated with a heightened susceptibility of the mammary gland to infection. It has been postulated that the met...


Ischemia\\/reperfusion injury in rat mesenteric venules: Red blood cell velocity and leukocyte rolling  

Microsoft Academic Search

The authors determined the effects of 15 (n = 9) and 30 (n = 12) minutes of warm ischemia on the rat mesentery and compared the results with those of a sham-operated group (n = 10). Red blood cell velocity and number of rolling leukocytes were assessed before ischemia as well as 10, 20, 30, 60, 90, and 120 minutes

Roland J. Beuk; Mirjam G. A. oude Egbrink; Harrie A. J. M. Kurvers; Harm-Jan Bonke; Geert-Jan Tangelder; Erik Heineman



Phagocytic and bactericidal activities of leukocytes in whole blood from atomic bomb survivors  

SciTech Connect

This study evaluated the phagocytic and bactericidal activities of peripheral blood leukocytes from Hiroshima and Nagasaki atomic bomb survivors for Staphylococcus aureus. The data were analyzed by multiple linear regression for age, sex, radiation exposure, city of exposure, and neutrophil counts. No significant radiation effect was observed for either blood phagocytic or bactericidal activities. The only significant variable for these functions was the neutrophil count.

Sasagawa, S.; Yoshimoto, Y.; Toyota, E.; Neriishi, S.; Yamakido, M.; Matsuo, M.; Hosoda, Y.; Finch, S.C. (Hiroshima Univ. (Japan))



A computational study of the effects of leukocyte adhesion on non-Newtonian blood flow in microvessels.  

NASA Astrophysics Data System (ADS)

The study of the effects of leukocyte adhesion on blood flow in microvessels is of significant interest for understanding the resistance changes in microcirculation. Several computational fluid dynamic studies have been done to understand flow resistance and drag forces due to adhering leukocytes by considering blood as a Newtonian fluid. In the present work we investigate the effect of leukocyte adhesion on the non-Newtonian flow of blood in microvessels. In the model the blood vessel is considered as a circular cylindrical tube and the leukocyte is considered as a truncated spherical protrusion inward from the blood vessel wall. The cases of single leukocyte adhesion and adhesion of a pair of leukocytes in the aligned and opposite configurations are considered. Blood (red blood cell suspension) is modeled as a Casson fluid and the parameters of the Casson fluid model are chosen both for cat blood and human blood. Comparisons are made for the resistance changes due to leukocyte adhesion for different hematocrits and for microvessels of different size.

Das, Bigyani; Popel, Aleksander S.



Laser-induced priming of human blood leukocytes  

NASA Astrophysics Data System (ADS)

We investigated the influence of He-Ne ((lambda) equals 632.8 nm) laser irradiation (LI) on a functional activity of human blood leucocytes. The method of luminol-dependent chemiluminescence with the zymosan-activated phagocytes was used. The leucocytes were irradiated without and in the presence of autologic human blood plasma, containing of the endogenous (porphyrins) and/or exogenous (phthalocyanine) photosensitizers. The LI initiated a priming of the leucocytes. Priming revealed itself after the activation of the phagocytes by zymosan. The changes of the calcium concentration in leucocytes cytoplasm were studied too. Fluorimetric method with Fura-2AM was used for this. The laser irradiation initiated the changes of the calcium concentration in the leucocytes cytoplasm. All the investigating parameters depended on the irradiation dose and on the concentration of photosensitizers. The results of this work allowed to formulate the main theses of the free radical mechanism of the low intensive laser irradiation action on human blood leucocytes.

Chichuk, Tatyana V.; Stranadko, Evgueni P.; Strashkevich, I. A.; Klebanov, Gennady I.



Leukocyte adhesion in angiogenic blood vessels. Role of E-selectin, P-selectin, and beta2 integrin in lymphotoxin-mediated leukocyte recruitment in tumor microvessels.  

PubMed Central

Interaction of circulating leukocytes with tumor microvasculature is a critical event in the recruitment of effector cells into the tumor stroma. We have examined the ability of lymphotoxin (TNF-beta), to stimulate rolling, adhesion, and transmigration of leukocytes in angiogenic blood vessels induced by tumor spheroids of Lewis lung carcinoma (LLC) implanted in dorsal skinfold chambers of nude mice. In the absence of cytokine stimulation, circulating leukocytes failed to appreciably interact with tumor microvessels (TMV), although significant rolling and adhesion was observed in normal vessels. However, stimulation with lymphotoxin (LT) resulted in a rapid increase in the number of fast and slow rolling leukocytes in TMV. Treatment with anti-P-selectin mAb 5H1 resulted in inhibition of fast rollers alone, while combination treatment with anti-P-selectin and anti-E-selectin (9A9) mAbs effectively blocked slow rolling of leukocytes. Superfusion of the lymphotoxin-stimulated neovasculature with leukotriene B4 (LTB4) resulted in stable cell adhesion followed by emigration of leukocytes into the tumor stroma. LTB4-mediated adhesion and transmigration was significantly inhibited by treatment with anti-beta2 mAb 2E6. These studies delineate a multistep cascade of leukocyte adhesion in TMV and demonstrate that stimulation of the neovasculature with cytokines and chemoattractants can result in P- and E-selectin-dependent rolling and beta2-dependent stable adhesion followed by transmigration into the tumor stroma.

Borgstrom, P; Hughes, G K; Hansell, P; Wolitsky, B A; Sriramarao, P



In vivo lipopolysaccharide exposure of human blood leukocytes induces cross-tolerance to multiple TLR ligands.  


In vitro and in vivo experiments in mice have shown that exposure of cells to the TLR4 ligand LPS induces tolerance toward a second exposure to LPS and induces cross-tolerance to certain other TLR ligands. Recently, we found that LPS tolerance in experimental human endotoxemia and Gram-negative sepsis is associated with elevated levels of IL-1R-associated kinase M, an intracellular negative regulator of MyD88-dependent TLR signaling. In the present study, we investigated whether in vivo exposure of humans to LPS induces tolerance in circulating leukocytes to other TLR agonists that rely either on MyD88- dependent or on MyD88-independent signaling. Analysis of TNF, IL-1beta, IL-6, and IL-10 levels in whole blood demonstrated that leukocytes were hyporesponsive to ex vivo LPS restimulation 3-8 h after i.v. LPS injection (4 ng/kg). Reduced cytokine release during the same interval was also observed in whole blood further stimulated with MyD88-dependent ligands for TLR2, TLR5, and TLR7 or with whole bacteria. Strikingly, blood leukocytes were also tolerant to a ligand for TLR3, which signals solely through a MyD88-independent (Toll IL-1R domain-containing adaptor-inducing IFN-beta (TRIF)-dependent) pathway. The hyporesponsiveness of leukocytes to TLR3 ligation was associated with reduced rather than increased levels of the recently identified TRIF inhibitor SARM. Taken together, these data indicate that systemic LPS challenge of human volunteers induces cross-tolerance to multiple TLR ligands that signal in a MyD88-dependent or MyD88-independent manner and suggest that LPS exposure of human blood leukocytes may hamper the inflammatory response to various microbial components. PMID:19542464

de Vos, Alex F; Pater, Jennie M; van den Pangaart, Petra S; de Kruif, Martijn D; van 't Veer, Cornelis; van der Poll, Tom



Levosimendan inhibits release of reactive oxygen species in polymorphonuclear leukocytes in vitro and in patients with acute heart failure and septic shock: a prospective observational study  

PubMed Central

Introduction Levosimendan is an extensively investigated inodilator showing also cardioprotective and antiinflammatory effects. The aim of our study was to explore the influence of levosimendan on polymorphonuclear leucocytes (PMN), a main source of reactive oxygen species, in vitro and in patients with acute heart failure or septic myocardial depression. Methods PMN isolated from healthy volunteers were incubated with levosimendan in vitro. After stimulation with N-formyl-Met-Leu-Phe (fMLP) or phorbol 12-myristate 13-acetate (PMA) respiratory burst was quantified using a fluorescent dye. Apoptosis and expression of cell adhesion molecules of PMN were measured by flow cytometry. For determination of in vivo effects patients with acute heart failure (n = 16) or septic cardiac failure (n = 9) receiving levosimendan treatment were enrolled consecutively. PMN were isolated to measure respiratory burst activity before treatment as well as one and two hours after initiation of levosimendan administration. Furthermore inflammatory, hemodynamic and renal function parameters were obtained. Results In vitro, levosimendan suppressed respiratory burst activity in fMLP or PMA stimulated PMN in a dose dependent manner by 30 ± 11% (P < 0.001) at 100 ng/mL and by 27 ± 17% (P < 0.001) at 1000 ng/mL respectively. Markers of apoptosis and PMN cell adhesion molecule expression remained unaffected by levosimendan treatment. In vivo, levosimendan treatment for two hours resulted in a significant reduction of PMA stimulated oxidative burst by 45% (P < 0.01) and fMLP stimulated oxidative burst by 49% (P < 0.05) in patients with acute heart failure. In patients suffering from septic shock levosimendan treatment decreased oxidative burst activity in unstimulated, fMLP and PMA stimulated PMN by 48% (P < 0.05), 46% (P < 0.01) and 43% (P < 0.01) respectively. Conclusions Levosimendan appears to exert distinct immunomodulatory effects by decreasing oxidative burst activity of PMN. This property might contribute to the previously described cardioprotective effects of the drug.



Comparison of sample fixation and the use of LDS-751 or anti-CD45 for leukocyte identification in mouse whole blood for flow cytometry  

Microsoft Academic Search

Flow cytometry methods used to measure leukocyte function often entail sample preparation procedures that cause artifactual cell activation. To avoid leukocyte activation by isolation techniques, some preparation methods use fluorescent markers to discriminate leukocytes from erythrocytes in whole blood. One of these markers, laser dye styryl-751(LDS-751), has been used to distinguish leukocytes by staining nucleic acid, but has been found

Melissa L. Maes; Lisa B. Davidson; Paul F. McDonagh; Leslie S. Ritter



Galactose-1-phosphate uridyltransferase and galactokinase activity in cultured human diploid fibroblasts and peripheral blood leukocytes  

PubMed Central

The specific activities of galactokinase and galactose-1-phosphate uridyltransferase were determined in peripheral blood leukocytes directly after separation from whole blood, and in cultured skin fibroblasts at various times during the subculture growth period. Growth curves were obtained for fibroblasts based on three different parameters: direct cell counts, total protein, and total deoxyribonucleic acid (DNA) content. At the time in culture when the specific activity of both enzymes was maximal and least variable, the ratio of transferase to galactokinase correlated well with the transferase genotypes of the original tissue donors. Leukocyte transferase: galactokinase ratios gave a similar distribution pattern. Whereas transferase activity in both fibroblasts and leukocytes was similar, galactokinase was approximately three times as active in fibroblasts as in leukocytes. All fibrobast cell strains tested had similar galactokinase activity regardless of transferase genotype. The kinetic properties of fibroblast galactokinase were examined. Galactose-1-phosphate inhibits galactokinase activity in both normal and galactosemic cell strains, whereas other glycolytic intermediates have no effect. There was no detectable transferase activity in eight galactosemic (GtG/GtG) cell strains when transferase activity was maximal in cell strains of other transferase genotypes. Inhibitors responsible for the absence of transferase activity could not be demonstrated. In addition, transferase activity in galactosemic cell lysates was not observed in cells during logarithmic growth; measurable uridine diphosphate galactose (UDPgal) pyrophosphorylase activity was found in human diploid fibroblast cultures, as well as significant levels of endogenous uridine triphosphate (UTP) in lysates of fibroblast cultures. Images

Tedesco, Thomas A.; Mellman, William J.



[Morpho-functional analysis of blood leukocytes after isoniazid administration].  


The functional-metabolic response peculiarities of the rat blood cells to isoniazid (one of the main antituberculosis medications) were studied. Isoniazid or its complex with pyridoxine were administrated orally for 3 months in therapeutic (5 mg/kg) and the toxic (100 mg/kg) doses (based on isoniazid). Isoniazid in toxic dose was found to inhibit the phagocytosis and the myeloperoxidase activity in neutrophils, the hydrolase activity in neutrophils and lymphocytes, but, at the same time, it activated dehydrogenase activity in lymphocytes. Pyridoxine modified the toxic effect of isoniazid on the metabolic processes (prevented the inhibition of phagocytosis and peroxidase activity in neutrophils, stimulated acid phosphatase activity in lymphocytes, decreased in the level of the lysosomal cationic proteins in neutrophils). It is suggested that the response of the blood cells to isoniazid, demonstrated in this work, was caused by the capacity of the medication to functionally activate the adrenal cortex, while the cellular-metabolic effects of pyridoxine could be associated with the modulation of glucocorticosteroid activity. PMID:23236893

Dolgushin, M V; Sobolev, V G; Gushchin, A S



In vitro evaluation of feline leukocytes radiolabeled in whole blood with 99mTc stannous colloid.  


Technetium-99m stannous colloid (9mTcSnC) has been used to radiolabel human leukocytes to investigate various inflammatory disorders. We investigated the in vitro behavior of feline leukocytes labeled in whole blood with 99mTcSnC. Heparinized blood samples were collected from healthy cats and divided into control and test aliquots. The latter were labeled with 99mTcSnC using a standard procedure. Leukocyte viability was determined for each sample using a trypan blue exclusion test. Labeling efficiency was determined for test aliquots. Test aliquots were layered onto Histopaque-1077 and centrifuged before measurement of radioactivity of the blood components. Leukocytes from radiolabeled and control samples were washed and incubated with opsonized zymosan particles to allow assessment of phagocytic function. Aliquots were taken from radiolabeled feline leukocyte samples at 1, 3, 4, and 7 h postlabelling. After centrifugation of each aliquot, radioactivity of the supernatant and pellet was measured and the labeling retention determined. Leukocyte viability in both radiolabeled and control samples was > 98%. The labeling efficiency was 95.2 +/- 0.14%. The distribution of radioactivity in feline blood was found to be 3.4 +/- 0.18% in plasma, 39.0 +/- 0.37% in erythrocytes, and 57.6 +/- 0.38% in leukocytes. Labeled feline leukocytes had phagocytic activity of 90.9 +/- 0.18% (control 91.3 +/- 0.15%). The radiolabeled leukocytes retained 93.4 +/- 0.19% of the radioactivity up to 7h postlabeling. 99TcSnC efficiently labeled feline leukocytes with no effect on viability and minimal effect on phagocytic function. The percentage retention of radioactivity by the leukocytes was still high at 7h postlabeling. PMID:19788042

Abushhiwa, Mohamed H; Salehi, Nouria S; Whitton, Robert C; Charles, Jennifer A; Finnin, Peter J; Lording, Peter M; Parry, Bruce W


Neutrophils Are Not Less Sensitive Than Other Blood Leukocytes to the Genomic Effects of Glucocorticoids  

PubMed Central

Background Neutrophils are generally considered less responsive to glucocorticoids compared to other inflammatory cells. The reported increase in human neutrophil survival mediated by these drugs partly supports this assertion. However, it was recently shown that dexamethasone exerts potent anti-inflammatory effects in equine peripheral blood neutrophils. Few comparative studies of glucocorticoid effects in neutrophils and other leukocytes have been reported and a relative insensitivity of neutrophils to these drugs could not be ruled out. Objective We assessed glucocorticoid-responsiveness in equine and human peripheral blood neutrophils and neutrophil-depleted leukocytes. Methods Blood neutrophils and neutrophil-depleted leukocytes were isolated from 6 healthy horses and 4 human healthy subjects. Cells were incubated for 5 h with or without LPS (100 ng/mL) alone or combined with hydrocortisone, prednisolone or dexamethasone (10?8 M and 10?6 M). IL-1?, TNF-?, IL-8, glutamine synthetase and GR-? mRNA expression was quantified by qPCR. Equine neutrophils were also incubated for 20 h with or without the three glucocorticoids and cell survival was assessed by flow cytometry and light microscopy on cytospin preparations. Results We found that glucocorticoids down-regulated LPS-induced pro-inflammatory mRNA expression in both cell populations and species. These drugs also significantly increased glutamine synthetase gene expression in both equine cell populations. The magnitude of glucocorticoid response between cell populations was generally similar in both species. We also showed that dexamethasone had a comparable inhibitory effect on pro-inflammatory gene expression in both human and equine neutrophils. As reported in other species, glucocorticoids significantly increase the survival in equine neutrophils. Conclusions Glucocorticoids exert genomic effects of similar magnitude on neutrophils and on other blood leukocytes. We speculate that the poor response to glucocorticoids observed in some chronic neutrophilic diseases such as severe asthma or COPD is not explained by a relative lack of inhibition of these drugs on pro-inflammatory cytokines expression in neutrophils.

Hirsch, Gaelle; Lavoie-Lamoureux, Anouk; Beauchamp, Guy; Lavoie, Jean-Pierre



Increased Thymidylate Synthase mRNA Concentration in Blood Leukocytes following an Experimental Stressor  

Microsoft Academic Search

Background: While it is well documented that immune responses, e.g. proliferative responses, can be influenced by psychosocial factors, e.g. stress, less is known about the biological mechanisms mediating such influences. The aim of the present investigation was to study the effect of an experimental stressor on mRNA levels in peripheral blood leukocytes of thymidylate synthase (TS), a gene necessary for

Eva Ehrnrooth; Robert Zacharia; Gunner Svendsen; Michael M. Jørgensen; Maya Yishay; Boe S. Sørensen; Jørgen Hjelm Poulsen; Hans von der Maase



Red blood cell and leukocyte alloimmunization in patients awaiting kidney transplantation  

PubMed Central

Objective To determine the rates of red blood cell and leukocyte alloimmunization in patients with chronic kidney disease awaiting kidney transplantation. Methods In this cross-sectional and prospective study, the serum of 393 chronic kidney disease patients on a transplant waiting list in Ceará, Northeastern Brazil were tested for red cell and leukocyte antibodies. In addition, demographic, clinical and laboratory data were collected. Results The average age in the sample of 393 patients was 34.1 ± 14 years. Slightly more than half (208; 52.9%) were male. The average numbers of transfusions and gestations were 3.1 ± 3.3 and 1.6 ± 6, respectively. One third (33.6%) were alloimmunized: 78% with leukocyte antibodies, 9.1% with red cell antibodies and 12.9% with both. Red cell antibodies were detected in 29 cases (7.4%), 17 of whom were women, who had received more transfusions than the males (p-value < 0.0001). The most frequently detected red cell antibodies belonged to the Rh (24.1%) and Kell (13.8%) blood group systems. Leukocyte antibodies were detected in 30.5% of cases, 83 of whom were women, who had received more transfusions than the males (p-value < 0.0001) and were more reactive to panel reactive antibodies (p-value < 0.0001). The mean alloreactivity to panel reactive antibodies was 47.7 ± 31.2%. Conclusion Chronic kidney disease patients on the transplant waiting list in Ceará, Brazil, display high rates of red cell (7.4%) and leukocyte (30.5%) alloimmunization. In this sample, alloimmunization was significantly associated with the number of transfusions and gender.

da Silva, Silvia Fernandes Ribeiro; Ferreira, Glaucia Maria; da Silva, Sonia Leite; Alves, Tania Maria de Oliveira; Ribeiro, Ilana Farias; Ribeiro, Thyciana Rodrigues; Cavalcante, Maria do Carmo Serpa



Altered gene expression pattern in peripheral blood leukocytes from patients with arterial hypertension.  


The role of various inflammatory mechanisms and oxidative stress in the development of atherosclerosis and arterial hypertension (AH) has been increasingly acknowledged during recent years. Hypertension per se or factors that cause hypertension along with other complications lead to infiltration of activated leukocytes in the vascular wall, where these cells contribute to the development of vascular injury by releasing cytokines, oxygen radicals, and other toxic mediators. However, molecular mechanisms underlying leukocyte activation at transcriptional level in AH are still far from being clear. To solve this problem we employed cDNA microarray technology to reveal the differences in gene expression in peripheral blood leukocytes from patients with AH compared with healthy individuals. The microarray data were verified by a semi-quantitative RT-PCR method. We found 25 genes with differential expression in leukocytes from AH patients among which 21 genes were upregulated and 4 genes were downregulated. These genes are implicated in apoptosis (CASP2, CASP4, and CASP8, p53, UBID4, NAT1, and Fte-1), inflammatory response (CAGC, CXCR4, and CX3CR1), control of MAP kinase function (PYST1, PAC1, RAF1, and RAFB1), vesicular trafficking of molecules among cellular organelles (GDI-1 and GDI-2), cell redox homeostasis (GLRX), cellular stress (HSPA8 and HSP40), and other processes. Gene expression pattern of the majority of genes was similar in AH patients independent of the disease stage and used hypotensive therapy, but was clearly different from that of normotensive subjects. PMID:17341625

Timofeeva, A V; Goryunova, L E; Khaspekov, G L; Kovalevskii, D A; Scamrov, A V; Bulkina, O S; Karpov, Yu A; Talitskii, K A; Buza, V V; Britareva, V V; Beabealashvilli, R Sh



Peripheral Blood Leukocyte Phagocytosis and Respiratory Response to Certain Macromolecular Substances in the ABCC-JNIH Adult Health Study, Hiroshima.  

National Technical Information Service (NTIS)

The functional integrity of the peripheral blood leukocytes of 10 heavily exposed subjects and 10 matched controls was evaluated by measuring oxygen consumption following the addition of latex particles and E. coli endotoxin, and measuring sensitized and ...

R. F. Barerras S. C. Finch



Maintenance of ?(1)-antitrypsin activity by means of co-application of hypochlorous acid-scavengers in vitro and in the supernatant of polymorphonuclear leukocytes: as a basis for a new drug delivery approach.  


Tissue destruction, pain and loss of function in chronically inflamed tissues can result from noxious agents released from myeloperoxidase (MPO) and its highly reactive product hypochlorous acid (HOCl) or proteases such as neutrophil elastase (NE). Currently there exists a high demand for medications that provide gentle treatments, free from side effects inherent in those prescribed today. One method to circumvent side effects is through the use of locally applied drug delivery. In contrast to systemic therapy, the main advantages of transport systems are the low dosages of drug with a time-controlled delivery. The aim of this study was to ascertain interactions of NE and its inhibitor ?(1)-antitrypsin (AT), the influence of hypochlorous acid (HOCl), as well as its scavengers, in order to define an effective mixture of drugs acting in a synergistic way which can be applied by means of drug delivery systems. These investigations determine the effective amounts of AT/HOCl-scavengers that drug mixtures need for delivery under inflammatory conditions in order to prevent tissue damage. AT was shown to inhibit NE in a dose-dependent manner, whereas a physiological concentration of 1.14 µM AT caused a significant NE inhibition (78%, pH 7.5). The concomitant existence of MPO/HOCl inactivated AT in a dose-dependent manner as well. To regain AT efficacy, HOCl-scavengers, such as L-methionine, ?-aminosalicylic acid and cefoperazone were additionally applied. Finally, AT was assembled as surface layer onto layer-by-layer biopolymer-coated microcarriers and carrier phagocytosis by polymorphonuclear leukocytes could be shown. PMID:23507783

Schönberg, Maria; Reibetanz, Uta; Rathmann, Sophie; Lessig, Jacqueline


Comparison of Methods for Assessing Chemotaxis of Monocytes and Polymorphonuclear Leukocytes Isolated From Patients With AIDS or AIDS-Related Conditions  

Microsoft Academic Search

We evaluated the ability of normal human peripheral blood monocytes and polymorpho- nuclear leucocytes (PMNL) isolated from patients with acquired immunodeficiency syndrome (AIDS) or AIDS-related conditions (ARC) to migrate toward a chemoattractant. Migration in blind-weii chambers was compared to that under agarose. Chemotaxis results obtained from both assays for PMNL were similar, however there was a difference in the results

Linda S. Martin; Thomas J. Spira; Sherry L. Orloff; Robert C. Holman


Effects of resistance exercise and protein ingestion on blood leukocytes and platelets in young and older men.  


This study investigated, in a multi-experiment design, the acute effects of milk protein ingestion, aging [50 young (approximately 26 years) vs. 45 older (approximately 61 years) men] and training state for the blood leukocyte and platelet responses acutely after a single bout of resistance exercise (RE). Moreover, basal effects of 21 weeks of resistance training (RT) were examined. The single bout of RE rapidly increased all blood leukocytes and platelets (P < 0.05). Protein ingestion before or before and after the RE bout did not have an effect on this response. However, younger men had a larger immediate exercise-induced response in leukocytes and platelets than older men. Basal fasting levels of leukocytes and platelets remained unchanged after 21 weeks of RT and this RT period did not change the acute RE-induced leukocyte and platelet response. The long-term RT was, however, able to slightly increase blood hematocrit. Blood platelet counts were consistently higher in the younger men when compared to the older men. Blood lymphopenia occurred only after a larger volume of exercise. In conclusion, the acute increase in blood leukocytes and platelets may be smaller in the older as when compared to the younger men. However, the number of immune cells and thus probably their function may not be affected by milk protein ingestion or months of resistance training. PMID:20101405

Hulmi, Juha J; Myllymäki, T; Tenhumäki, M; Mutanen, N; Puurtinen, R; Paulsen, G; Mero, A A



Occurrence of nonlymphoid leukocytes that are not derived from blood islands in Xenopus laevis larvae.  


Previous immunohistochemical observations using the monoclonal antibody (XL-1) which recognizes all types of leukocytes in Xenopus laevis revealed the occurrence of XL-1+ cells in the mesenchyme throughout the early larval body, before the appearance of any lymphocytes. The present experiments were performed to determine whether these leukocytes originate, like lymphocytes and red blood cells (RBCs), in the ventral blood islands (VBI) or the dorsolateral plate (DLP). For tracing the derivation of cells, a specific staining by quinacrine to nuclei of X. laevis and Xenopus borealis hybrid (LB) cells was used to distinguish them from X. laevis (LL) cells. Orthotopic graftings of VBI tissue from st.22-23 LB embryos to the stage-matched LL embryos and examinations at st.44-45 before differentiation of the lymphocytes showed that the proportion of XL-1+ LB cells was always significantly lower than that of RBCs with the same marker in all experimental larvae. The head (LB)-body (LL) chimeras from st.22-23 embryos and culture of the head-portions as VBI- and DLP-free explants from st.14-23 embryos both demonstrated that a significant number of XL-1+ cells which had originated in the head portions had begun to differentiate by st.42-43. These results indicate that there is a significant population of larval nonlymphoid leukocytes (mostly macrophages) that do not originate from either the VBI or DLP region, and are distributed in the mesenchyme throughout the body. PMID:2202604

Ohinata, H; Tochinai, S; Katagiri, C



Sensitivity and specificity of blood leukocyte counts as an indicator of mortality in horses after colic surgery.  


The objectives of this study were to describe and relate perioperative changes in blood leukocyte counts to the outcome of surgical colic horses, determine a cut-off value in the early postoperative period to obtain an indicator of the outcome, and compare the obtained value to a validation population of horses. Fifty-three horses undergoing colic surgery were included in the descriptive part of the study. Total leukocyte counts were performed before, during and serially after surgery. A receiver operating characteristic analysis was performed on the leukocyte counts of 45 of these horses to determine a cut-off value for the outcome. The results obtained were validated on a second set of 50 horses that underwent colic surgery in similar conditions. The kinetics of blood leukocytes in survivors was higher than in non-survivors during the first days. Non-survivor horses were more likely to have at least one blood leukocyte count ?3.9×10(3)/mm(3) between 28 and 60 hours after surgery than survivor horses. This cut-off value was confirmed in the validation population. These results suggest that routine values of blood leukocyte counts can be used as an additional prognostic indicator after colic surgery alongside other predictors previously associated with the outcome. PMID:23939753

Salciccia, A; Sandersen, C; Grulke, S; de la Rebière de Pouyade, G; Caudron, I; Serteyn, D; Detilleux, J



TNF{alpha} release from peripheral blood leukocytes depends on a CRM1-mediated nuclear export  

SciTech Connect

Tumor necrosis factor-{alpha} (TNF{alpha}) is a potent pro-inflammatory cytokine that plays a major role in the pathogenesis of acute and chronic inflammatory disorders such as septic shock and arthritis, respectively. Leukocytes stimulated with inflammatory signals such as lipopolysaccharide (LPS) are the predominant producers of TNF{alpha}, and thus control of TNF{alpha} release from stimulated leukocytes represents a potential therapeutic target. Here, we report that leptomycin B (LMB), a specific inhibitor of CRM1-dependent nuclear protein export, inhibits TNF{alpha} release from LPS-stimulated human peripheral blood neutrophils and mononuclear cells. In addition, immunofluorescence confocal microscopy and immunoblotting analysis indicate that TNF{alpha} is localized in the nucleus of human neutrophils and mononuclear cells. This study demonstrates that the cellular release of TNF{alpha} from stimulated leukocytes is mediated by the CRM1-dependent nuclear export mechanism. Inhibition of CRM1-dependent cellular release of TNF{alpha} could thus provide a novel therapeutic approach for disorders involving excessive TNF{alpha} release.

Miskolci, Veronika [Department of Biology, St. John's University, NY 11439 (United States); Department of Pediatrics, Feinstein Institute for Medical Research at the North Shore-Long Island Jewish Health System, New Hyde Park, NY 11040 (United States); Ghosh, Chandra C. [Department of Biology, St. John's University, NY 11439 (United States); Rollins, Janet [Department of Biology, St. John's University, NY 11439 (United States); Romero, Carlos [Department of Biology, St. John's University, NY 11439 (United States); Vu, Hai-Yen [Department of Biology, St. John's University, NY 11439 (United States); Robinson, Staci [Department of Biology, St. John's University, NY 11439 (United States); Davidson, Dennis [Department of Pediatrics, Feinstein Institute for Medical Research at the North Shore-Long Island Jewish Health System, New Hyde Park, NY 11040 (United States); Vancurova, Ivana [Department of Biology, St. John's University, NY 11439 (United States) and Department of Pediatrics, Feinstein Institute for Medical Research at the North Shore-Long Island Jewish Health System, New Hyde Park, NY 11040 (United States)]. E-mail:



Cisplatin-DNA damage and repair in peripheral blood leukocytes in vivo and in vitro.  

PubMed Central

We have extended our studies on the relationship between cisplatin/carboplatin-induced DNA damage in readily accessible tissue(s) and clinical response to therapy. Such an approach may assist in the study of cancer drug resistance and in establishing parameters for assessing human populations for sensitivity to DNA damaging agents in the environment. Platinum-DNA adduct levels were measured by atomic absorbance spectrometry. DNA repair capacity was assessed in human T-lymphocytes by the ability to repair cisplatin lesions in cellular DNA or in transfected plasmid DNA. In a "blinded" study of 21 patients receiving combination cisplatin/carboplatin drug therapy, there was a direct relationship between DNA damage in leukocytes and disease response (summary two-sided p = 0.00011). The cohort of patients had 15 different tumor types, suggesting that blood tissue and tumor tissue of an individual may process platinum-DNA damage similarly regardless of the tissue of origin of the tumor. In leukocytes in vivo, persistence and accumulation were prominent features of the cisplatin-DNA adduct profile. Functional DNA repair capacity has been studied in eight human leukocyte cell lines in vitro (three, T-cells; three, B-cells; one, monocytic; one, promyelocytic), using a host cell reactivation assay with cisplatin-damaged pRSVcat. In the three T cell lines studied, host cell reactivation efficiency was directly related to the cells' abilities to repair cisplatin-damaged cellular DNA (correlation coefficient = 0.993).(ABSTRACT TRUNCATED AT 250 WORDS)

Dabholkar, M; Bradshaw, L; Parker, R J; Gill, I; Bostick-Bruton, F; Muggia, F M; Reed, E



Blood transfusion with autologous and leukocyte-depleted or standard allogeneic red blood cells and the immune response to open heart surgery.  


Allogeneic blood transfusions have been associated with impaired outcome in surgical patients. This effect may be mediated by leukocytes. Animal experiments have shown that at least some of the effect can be modified by removal of leukocytes from transfused blood. Therefore, we compared the effects of autologous + leukocyte-depleted against standard allogeneic red blood cell transfusion on postoperative immunosuppression in 24 men undergoing coronary artery bypass surgery. In the autologous + leukocyte-depleted red blood cell transfusion group, patients received 800 +/- 200 mL (mean +/- SD) autologous blood and 2.2 +/- 2.0 units (mean +/- SD) of leukocyte-depleted saline-adenine-glucose-mannitol (SAGM) red blood cells. In the standard red blood cell transfusion group, patients were transfused with 5.5 +/- 1.4 units (mean +/- SD) of SAGM red blood cells. Leukocyte and differential counts; percentages of lymphocyte subpopulations (CD3-, CD4-, CD8-, CD16-, CD20-, CD25-, and B5-positive lymphocytes) and monocytes (CD14); phytohemagglutinin-, concanavalin A-, and pokeweed mitogen-induced and unstimulated proliferation of separated lymphocytes; unstimulated and pokeweed mitogen-stimulated production of IgG, IgM, or IgA; and serum interleukin-6, interleukin-1 beta, and serum C-reactive protein concentrations were measured preoperatively and on postoperative Days 1, 7, and 21. Significant changes were seen in these variables, but there were no differences between the groups. Three of the 12 patients in the allogeneic leukocyte-containing red blood transfusion group became human lymphocyte antigen (HLA) alloimmunized. No infections or other complications occurred in any patients. We conclude that HLA alloimmunization was the only effect that could be modified by use of autologous blood. PMID:7943771

Perttilä, J T; Salo, M S; Jalonen, J R; Kuttila, K T; Viinamäki, O; Pulkki, K J



Differential processing of amyloid precursor protein in brain and in peripheral blood leukocytes.  


Because amyloid precursor protein (APP) fragments exist in many tissues throughout the body, including the fluid compartments of blood, they have been the focus of numerous investigations into their potential as a biomarker of Alzheimer's disease. Using immunohistochemistry, immunoelectron microscopy, Western blot, and quantitative real-time-polymerase chain reaction (qRT-PCR) analysis we examined whether APP processing in leukocytes is analogous to APP processing in the brain. We show APP immunoreactivity at light and electron microscopic levels in the cytoplasm and nucleus of peripheral blood leukocytes (PBL) yet our Western blot analysis data demonstrated that brain and PBL contain different APP fragments and differentially expressed APP processing enzymes. A Disintegrin and Metalloproteinase domain 10 (ADAM10), nicastrin, and beta-secretase 2 (BACE2) were present in brain but were undetected in PBL. Presenilin 1 and beta-secretase 1 (BACE1) were detected in both tissues but showed different patterns in Western blots. Quantitative PCR results identified Neprilysin as the only processing enzyme we interrogated in which Western and quantitative PCR data coincided. Although our data on differential processing of APP in brain and PBL point to exercising caution when generalizing between blood and brain with regard to mechanisms, they have no implications regarding utility as biomarkers. PMID:23298733

Delvaux, Elaine; Bentley, Karen; Stubbs, Victoria; Sabbagh, Marwan; Coleman, Paul D



Comparative rheology of the adhesion of platelets and leukocytes from flowing blood: why are platelets so small?  


We investigated rheological adaptation of leukocytes and platelets for their adhesive functions in inflammation and hemostasis, respectively. Adhesion and margination of leukocytes or platelets were quantified for blood perfused through capillaries coated with P-selectin or collagen, when flow rate, suspending phase viscosity, red cell aggregation, or rigidity was modified. Independent variation of shear rate and shear stress indicated that the ability of platelets to attach at higher levels than leukocytes was largely attributable to their smaller size, reducing their velocity before attachment, and, especially, drag after attachment. Increasing red cell aggregation increased the number of marginated and adhering leukocytes but inhibited platelet adhesion without effect on the number marginated. Increasing red cell rigidity tended to inhibit leukocyte adhesion but promote platelet adhesion. The effects on platelets may be explained by changes in the depth of the near-wall, red cell-depleted layer; broadening (or narrowing) this layer to greater (or less) than the platelet diameter would decrease (or increase) the normal force applied by red blood cells and make attachment less (or more) efficient. Thus different adhesive capabilities of leukocytes and platelets may arise from their differences in size, both directly because of influence on cell velocity and force experienced at the wall and indirectly through effects of size on margination in the bloodstream and interaction with the cell-free layer. In addition, red cell aggregation (of hitherto uncertain physiological significance) may be useful in promoting leukocyte adhesion in inflamed venules but inhibiting unwanted platelet deposition in veins. PMID:23585130

Watts, Tim; Barigou, Mostafa; Nash, Gerard B



Enhanced Susceptibility of Escherichia coli Pretreated with RP7293 to Leukocyte Activity: Comparison with Other Antimicrobial Agents  

Microsoft Academic Search

Antimicrobial agents may interact with polymorphonuclear leukocytes and directly modify the leukocyte functions, or bacteria can be modified by the antimicrobial agents causing them to be treated differently by phagocytic cells. Brief exposure of bacteria to high levels of antimicrobials can affect their interaction with leukocytes. The susceptibility of Escherichia coli to phagocytosis and killing by human polymorphonuclear leukocytes in

P. Pérez; I. Herrera; M. Martín; P. Martínez; M. L. Gómez-Lus



Dengue viruses and mononuclear phagocytes. II. Identity of blood and tissue leukocytes supporting in vitro infection  

PubMed Central

Studies were made on the identity of human and monkey mononuclear leukocytes permissive to antibody-enhanced dengue 2 virus (D2V) infection. In cultures of peripheral blood leukocytes (PBL) inoculated immediately after separation, it was concluded that only mononuclear phagocytes support dengue infection. This is based upon observations that D2V-permissive cells were resistant to 1,200 rads, were both plastic adherent and nonadherent, were removed when passed through nylon wool columns in 10 percent fetal bovine serum or 100 percent autologous serum, and were destroyed by incubation with 100 ?g/ml particulate silica. On direct immunofluorescence staining, perinuclear dengue antigen was visualized at 24 h, becoming maximal at 60 h. Antigen-containing cells had ample cytoplasm, ruffled cytoplasmic membrane, and 73 percent were actively phagocytic. As further evidence of the infection of mononuclear phagocytes, antibody-enhanced D2V replication was observed in bone marrow cultures from five of five rhesus monkeys, but not in cell cultures of spleen, thymus, or lymph nodes prepared from the same animals. It is hypothesized that dengue virus complexed with non-neutralizing antibody is internalized by immune phagocytosis in a mononuclear phagocyte with a defective virus-destroying mechanism. Dengue permissiveness may depend upon cellular immaturity since bone marrow leukocytes could be infected even when held for 4 days before infection while PBL held for this time decreased in permissiveness. In vitro antibody-dependent infection of mononuclear phagocytes should prove useful as a model for study of immunopathologic mechanisms in human dengue.

Halstead, SB; O'Rourke, EJ; Allison, AC



Sildenafil prevents indomethacin-induced gastropathy in rats: role of leukocyte adherence and gastric blood flow  

PubMed Central

Nitric oxide (NO) is an important mediator of gastric mucosal defense. Sildenafil (SILD), a cyclic GMP-specific phosphodiesterase inhibitor, promotes an increase in cGMP concentrations in the gastrointestinal tract. cGMP mediates many of the biological actions of NO.We tested the hypothesis that SILD could increase mucosal defense against indomethacin-induced gastropathy in rats.SILD (1, 4 or 10?mg?kg?1, p.o.) pretreatment significantly reduced (P<0.01) the gastric damage and the increase in gastric myeloperoxidase (MPO) activity elicited by indomethacin (20?mg?kg?1 p.o.), with the maximal effect at the dose of 10?mg?kg?1.L-NAME (3, 10 or 20?mg?kg?1, i.p.) dose dependently reversed the protective effects of SILD, an effect not seen when L-arginine (L-ARG) (200?mg?kg?1, i.p.) was co-administered with L-NAME.Indomethacin-induced leukocyte adhesion, assessed by intravital microscopy, was decreased (P<0.01) by SILD, and this effect was reversed by L-NAME cotreatment.Indomethacin elicited a decrease in gastric blood flow and in gastric PGE2 levels. SILD was able to prevent the decrease in gastric blood flow (P<0.01), without diminishing the inhibitory effect of indomethacin on prostaglandin synthesis.These results indicate that SILD, acting via NO-dependent mechanisms, prevents indomethacin-induced gastropathy, possibly through a reduction of leukocyte adhesion and maintenance of gastric blood flow.

Santos, Camila L; Souza, Marcellus H L P; Gomes, Antoniella S; Lemos, Henrique P; Santos, Armenio A; Cunha, Fernando Q; Wallace, John L



Comparison of in vitro effects of flunixin and tolfenamic acid on human leukocyte and platelet functions  

Microsoft Academic Search

A study was made to compare the effects of two nonsteroidal antiinflammatory drugs (NSAIDs), flunixin and tolfenamic acid, on the leukotriene B4 (LTB4) production and migration of human polymorphonuclear leukocytes (PMNs) as well as on platelet aggregation and thromboxane B2 (TxB2) production during blood clotting. Tolfenamic acid inhibited LTB4 production in PMNs as well as FMLP- and LTB4-induced PMN migration

H. Kankaanranta; E. Moilanen; H. Vapaatalo



Sensitive microculture method for isolation of human immunodeficiency virus type 1 from blood leukocytes.  

PubMed Central

A study was conducted to compare our standard culture with a new microculture procedure for isolation of human immunodeficiency virus type 1 (HIV-1) from blood leukocytes. A total of 137 blood specimens from 102 HIV-1 antibody-positive individuals (52 were asymptomatic, 31 were symptomatic, and 19 had AIDS) were cultured in a microculture system in which 10(6) of the patients' peripheral blood mononuclear cells (PBMC) were cocultured with 10(6) phytohemagglutinin (PHA)-stimulated PBMC from an HIV-1 antibody-negative blood donor in 1.2 ml of culture medium. Results were compared with those of a historical control group of 139 standard HIV-1 cultures from 108 HIV-1 antibody-positive subjects (58 were asymptomatic, 36 were symptomatic, and 14 had AIDS). For standard cultures, 10 x 10(6) of the patients' PBMC were cocultured with 5 x 10(6) PHA-stimulated PBMC from an HIV-1 antibody-negative blood donor in 15 ml of culture medium. HIV-1 was isolated in 128 (93%) microcultures and 133 (96%) standard cultures. Both methods identified more than 75% of the positive cultures within 7 days and 100% of the positive cultures within 14 days. The isolation rates for HIV-1 in microcultures compared with standard cultures were 91 versus 93% (specimens from asymptomatic individuals), 93 versus 96% (specimens from symptomatic individuals), and 97 versus 100% (specimens from patients with AIDS). The median time to positivity for both culture methods was 7 days, and this correlated significantly with symptoms and CD4+ cell counts. The microculture method is a sensitive and less expensive system for isolation of HIV-1 from PBMC of HIV-1 antibody-positive individuals, and we recommend it as the culture method of choice, especially for children and patients with AIDS and severe anemia or leukopenia whose blood volume is an important consideration.

Erice, A; Sannerud, K J; Leske, V L; Aeppli, D; Balfour, H H



In vivo endotoxin synchronizes and suppresses clock gene expression in human peripheral blood leukocytes  

PubMed Central

Objectives The intravenous administration of a bolus dose of endotoxin to healthy human subjects triggers acute systemic inflammatory responses that include cytokine production and dynamic changes in gene expression in peripheral blood leukocytes (PBL). This study sought to determine the state of clock gene expression in human PBL, and leukocytes subpopulations, challenged with in vivo endotoxin at two circadian/diurnal phases of the clock. Design Clinical and laboratory investigation. Setting University-based research laboratory and clinical research center Subjects Human volunteers. Interventions Human subjects were administered a standard dose of endotoxin (2ng/kg) or saline at either 09:00 or 21:00 h. Blood samples were collected at selected time points pre- and post-infusion. Measurements and Mains results Clock gene expression was determined in human PBL, neutrophils, and monocytes, by quantitative real-time polymerase chain reaction. The fold change for each gene was determined using the 2(-??Ct) method. We show that endotoxin causes profound suppression of circadian clock gene expression, clearly manifested in human PBL, neutrophils, and monocytes. Clock, Cry1-2, Per3, CSNK1?, Rora and Rev-erb gene expression were all reduced by 80-90% with the nadir between 3 to 6 hours post-infusion. Per1 and Per2 reached an expression nadir between 13 and 17 hours post-infusion. The levels of plasma interleukin-6 and tumor necrosis factor peaked and then returned to baseline within 6 hours. In contrast, clock gene expression remained suppressed for up to 17 hours, irrespective of the phase of the clock at the time of the endotoxin challenge. Endotoxin did not perturb the melatonin secretory rhythm. Conclusions Circadian clock gene expression in PBL is dramatically altered, and possibly uncoupled from the activity of the central clock, during periods of acute systemic inflammation. The realignment of the central and peripheral clocks may constitute a previously unappreciated key factor affecting recovery from disease in humans.

Haimovich, Beatrice; Calvano, Jacqueline; Haimovich, Adrian D.; Calvano, Steve E.; Coyle, Susette M.; Lowry, Stephen F.



[Comparative transcriptome analysis of human aorta atherosclerotic lesions and peripheral blood leukocytes from essential hypertension patients].  


One of the major cardiovascular risk factor which predisposes to and accelerates atherosclerosis is arterial hypertension (AH). To determine the molecular basis of the crosslink between AH and atherosclerosis for the development of new treatment strategies large-scale transcriptome analysis of the cells implicated in atherogenesis is needed. We used cDNA microarray technique for simultaneous analysis of gene expression in human abdominal aorta normal sites and atherosclerotic lesions of different histological types, as well as in peripheral blood leukocytes from patients with essential hypertension (EH) and donors. The microarray data were verified by quantitative RT-PCR (reverse transcription coupled with polymerase chain reaction) and immunohistochemical analysis. Differential expression of 40 genes has been found, among which twenty two genes demonstrated up-regulation and 18 genes demonstrated down-regulation in atherosclerotic aorta compared with normal vessel. New gene-candidates, implicated in atherogenesis, have been identified - FPRL2, CD37, CD53, RGS1, LCP1, SPI1, CTSA, EPAS1, FHL1, GEM, RHOB, SPARCL1, ITGA8, PLN, and COL14A1. These genes participate in cell migration and adhesion, phenotypic changes of smooth muscle cells, immune and inflammatory reactions, oxidative processes and extracellular matrix remodeling. We have found increased expression levels of CD53, SPI1, FPRL2, SPP1, CTSD, ACP5, LCP1, CTSA and LIPA genes in peripheral blood leukocytes from EH patients and in atherosclerotic lesions of human aorta. The majority of these genes significantly (p<0.005) positively (r>0.5) correlated with AH stage as well as with histological grading of atherosclerotic lesions. PMID:19772500

Timofeeva, A V; Goriunova, L E; Khaspekov, G L; Il'inskaia, O P; Sirotkin, V N; Andreeva, E R; Tararak, E M; Bulkina, O S; Buza, V V; Britareva, V V; Karpov, Iu A; Bibilashvili, R Sh



Cytokine Generation in Whole Blood, Leukocyte–Depleted and Temporarily Warmed Red Blood Cell Concentrates  

Microsoft Academic Search

Background and Objectives: It has been suggested that inflammatory cytokines such as Interleukin (IL)–1?, IL–6, tumor necrosis factor–? (TNF–?) and IL–8 might be responsible for a large number of non–antibody–mediated adverse reactions to the transfusion of blood components, especially of platelet concentrates (PCs). The aim of this study was to compare the levels of proinflammatory cytokines in different blood components

V. Weisbach; C. Wanke; J. Zingsem; R. Zimmermann; R. Eckstein



Chronic inflammation and hemodialysis reduce immune competence of peripheral blood leukocytes in end-stage renal failure patients  

Microsoft Academic Search

Immunoincompetence is a profound problem in end-stage renal failure patients undergoing hemodialysis, and chronic inflammation with altered serum levels of inflammation markers has been reported. Gene expression patterns have had little relevance for leukocyte research so far because of limitations in transcript levels and stability. Using a new stimulation system we induced the expression of immune-relevant transcripts in whole blood

Kerstin Knerr; Reiner Füth; Patrick Hemsen; Walburga Mohné; Antonia Heinig; Werner Kleophas; Werner A. Scherbaum; Stephan Martin



Cryptic sialic acid binding lectins on human blood leukocytes can be unmasked by sialidase treatment or cellular activation  

Microsoft Academic Search

We recently reported that the sialic acid-specific binding sites of CD22 molecules on B cells are masked by endog- enous ligands, and can be unmasked by sialidase treatment or cellular activation. Here, we show that many other human blood leukocyte types have endogenous sialic acid binding sites that can be unmasked by sialidase treatment. Truncation of sialic acid side chains

Nahid Razi; Ajit Varki



Gender dimorphism in differential peripheral blood leukocyte counts in mice using cardiac, tail, foot, and saphenous vein puncture methods  

Microsoft Academic Search

BACKGROUND: In many animal models that investigate the pathology of various diseases, there is a need to monitor leukocyte counts and differentials. However, various researchers use a range of different techniques in male and female laboratory animals to collect such blood variable information. These studies are then compared to one another without consideration of the possibility that different bleeding sites

Diana C Doeing; Jessica L Borowicz; Elahé T Crockett



Influence of blood transfusion on bactericidal activity of human leukocytes and sera against Yersinia enterocolitica and Salmonella typhimurium  

Microsoft Academic Search

Patients undergoing joint surgery and blood transfusion were studied. Serum and leukocyte bactericidal tests in vitro against Salmonella typhimurium and Yersinia enterocolitica were carried out preoperatively as well as on the 1st, 3rd and 7th days after the operation. The serum complement (C3 and C4) concentrations were determined at the same intervals. It was found that after blood transfusion the

Nadya Markova; T Radoucheva; V Kussovski; K Dilova; M Shtarbova; I Paskaleva



Forces on a Wall-Bound Leukocyte in a Small Vessel Due to Red Cells in the Blood Stream  

PubMed Central

As part of the inflammation response, white blood cells (leukocytes) are well known to bind nearly statically to the vessel walls, where they must resist the force exerted by the flowing blood. This force is particularly difficult to estimate due to the particulate character of blood, especially in small vessels where the red blood cells must substantially deform to pass an adhered leukocyte. An efficient simulation tool with realistically flexible red blood cells is used to estimate these forces. At these length scales, it is found that the red cells significantly augment the streamwise forces that must be resisted by the binding. However, interactions with the red cells are also found to cause an average wall-directed force, which can be anticipated to enhance binding. These forces increase significantly as hematocrit values approach 25% and decrease significantly as the leukocyte is made flatter on the wall. For a tube hematocrit of 25% and a spherical protrusion with a diameter three-quarters that of the vessel, the average forces are increased by ?40% and the local forces are more than double those estimated with an effective-viscosity-homogenized blood. Both the enhanced streamwise and wall-ward forces and their unsteady character are potentially important in regard to binding mechanisms.

Isfahani, Amir H.G.; Freund, Jonathan B.



Methylation markers for small cell lung cancer in peripheral blood leukocyte DNA  

PubMed Central

Introduction Small-cell lung cancer (SCLC) is the most aggressive form of lung malignancy. Methods To identify and validate potential DNA methylation markers for risk assessment and disease detection, we examined peripheral blood leukocyte DNA specimens for methylation differences between SCLC cases and controls. We tested 1,505 CpG sites using the Illumina Beadchip assay and validated 9 CpG sites using pyrosequencing technology. Results In 44 matched SCLC case-control pairs, we identified significant differences at 62 CpG sites (false discovery rate ? 0.05) in 52 independent genes. Of those, we further determined 43 sites in 36 genes with a mean methylation level difference greater than 0.03 between the cases and controls. We then selected and validated 9 CpG sites for methylation differences in an independent set of 138 matched case-control pairs. The 9 validated CpG sites predicted a higher risk for cases than controls in 85.8% of all pairs of cases and controls, and two (in genes CSF3R and ERCC1) jointly contributed most of the discriminating ability. Conclusions Our replicated results demonstrated feasibility of applying large-scale methylation arrays for biomarker discovery and subsequent validation in peripheral blood DNA. The CpG sites identified in this study may potentially assist in risk prediction and diagnosis of SCLC.

Wang, Liang; Aakre, Jeremiah A.; Jiang, Ruoxiang; Marks, Randolph S.; Wu, Yanhong; Thibodeau, Stephen N.; Pankratz, V. Shane; Yang, Ping



Polymorphonuclear function in naturally occurring renal failure in dogs  

Microsoft Academic Search

Chronic renal failure causes immunosuppression in people and is thought to be one of the causes of non-infectious secondary immunosuppression in dogs. The purpose of this study was to evaluate changes in counts and activity of polymorphonuclears in dogs with chronic renal failure in various stages. Haematological, biochemical examinations and examination of non-specific immune response cells (total and differential leukocyte

S. Kralova; L. Leva; M. Toman


Oxidative Stress by Peripheral Blood Mononuclear Cells Is Increased in Hypertensives with an Extreme-Dipper Pattern and\\/or Morning Surge in Blood Pressure  

Microsoft Academic Search

Because oxidative stress and inflammation are known to play important roles in the pathogenesis of cardiovascular events that occur most frequently in the morning, we studied the association between reactive oxygen species (ROS) formation by polymorphonuclear leukocytes (PMNs) or mononuclear cells (MNCs) and morning blood pressure (BP) rhythm. A total of 31 hypertensives in whom ambulatory BP monitoring was performed

Kensaku Maeda; Kenichi Yasunari; Takanori Watanabe; Munehiro Nakamura



Blood leukocyte DNA hypomethylation and gastric cancer risk in a high-risk Polish population  

PubMed Central

Global hypomethylation has been shown to increase genome instability potentially leading to increased cancer risk. We determined whether global methylation in blood leukocyte DNA was associated with gastric cancer in a population-based study on 302 gastric cancer cases and 421 age- and sex-matched controls in Warsaw, Poland, between 1994 and 1996. Using PCR-pyrosequencing, we analyzed methylation levels of Alu and LINE-1, 2 CG-rich repetitive elements, to measure global methylation levels. Gastric cancer risk was highest among those with lowest level of methylation in either Alu (OR = 1.3, 95% CI = 0.9–1.9) or LINE-1 (OR = 1.4, 95% CI = 0.9–2.0) relative to those with the highest levels, although the trends were not statistically significant. For Alu, the association was stronger among those aged 70 or older (OR = 2.6, 95% CI = 1.3–5.5, p for interaction = 0.02). We did not observe meaningful differences in the associations by other risk factors and polymorphisms examined. For LINE-1, the association tended to be stronger among individuals with a family history of cancer (OR = 3.1, 95% CI = 1.4–7.0, p for interaction = 0.01), current alcohol drinkers (OR = 1.9, 95% CI = 1.0–3.6, p for interaction = 0.05), current smokers (OR = 2.3, 95% CI = 1.1–4.6, p for interaction = 0.02), those who rarely or never consumed fruit (OR = 3.1, 95% CI = 1.2–8.1, p for interaction = 0.03), CC carriers for the MTRR Ex5+123C>T polymorphism (OR = 2.3, 95% CI = 1.2–4.4, p for interaction = 0.01) and TT carriers for the MTRR Ex15+572T>C polymorphism (OR = 1.7, 95% CI = 1.0–2.8, p for interaction = 0.06). The association was not different by sex, Helicobacter pylori infection, intake of folate, vitamin B6 and total protein and the remaining polymorphisms examined. Our results indicate that interactions between blood leukocyte DNA hypomethylation and host characteristics may determine gastric cancer risk.

Hou, Lifang; Wang, Hao; Sartori, Samantha; Gawron, Andrew; Lissowska, Jolanta; Bollati, Valentina; Tarantini, Letizia; Zhang, Fang Fang; Zatonski, Witold; Chow, Wong-Ho; Baccarelli, Andrea



The Activation Pattern of Blood Leukocytes in Head and Neck Squamous Cell Carcinoma Is Correlated to Survival  

PubMed Central

Head and neck squamous cell carcinoma (HNSCC) is known to cause substantial immunosuppression. The present study was designed to characterize blood leukocyte activation in HNSCC and to investigate if the individual activation pattern could be related to tumor progress and survival. The leukocyte activation profile of HNSCC patients and healthy controls was assessed with flow cytometry. HNSCC patients displayed increased numbers of monocytes, neutrophils and total leukocytes as well as an enhanced neutrophil/lymphocyte ratio. In addition, patients had a higher percentage of CD69+, CD71+ and CD98+ T cell subsets and NK cells, and a reduced expression of L-selectin in CD14highCD16+ monocytes and neutrophils, when compared to controls. These changes could be correlated to both tumor burden and spread to lymph nodes. Among the cancer patients an increased neutrophil/lymphocyte ratio, a low neutrophil and CD14high CD16+ monocyte activation state and an elevated CD4/CD8 ratio were related to poor survival. In contrast, a high percentage of CD98+ Th cells appeared to be associated with a better outcome. Taken together, the present data indicate that HNSCC causes activation of blood leukocytes and that the individual activation pattern can be linked to prognosis.

Millrud, Camilla Rydberg; Mansson Kvarnhammar, Anne; Uddman, Rolf; Bjornsson, Sven; Riesbeck, Kristian; Cardell, Lars Olaf



Cytokine Gene Expression by Peripheral Blood Leukocytes in Horses Experimentally Infected with Anaplasma phagocytophila  

PubMed Central

Human granulocytic ehrlichiosis (HGE), a tick-borne zoonosis, is caused by an obligatory intragranulocytic bacterium, the HGE agent, a strain of Anaplasma phagocytophila. The equine model of HGE is considered valuable in understanding pathogenic and immune mechanisms of HGE. In the present study, cytokine mRNA expression by peripheral blood leukocytes (PBLs) in horses was examined during the course of infection by intravenous inoculation of A. phagocytophila or by allowing feeding by infected ticks. The p44 genes encoding the major outer membrane protein P44s of A. phagocytophila were detected by PCR in PBLs of all four horses from 4 to 20 days postexposure. During the 20-day infection period, interleukin-1? (IL-1?) and tumor necrosis factor alpha (TNF-?) mRNA expression was upregulated in PBLs of all four horses, and IL-8 mRNA expression was upregulated in three horses. Gamma interferon, IL-10, and IL-12 p35 mRNAs were weakly expressed in only one horse each. IL-2, IL-4, IL-6, and IL-12 p40 mRNA expression , however, could not be detected in the PBLs of any of the four horses. These results suggest that IL-1?, TNF-?, and IL-8 generation during A. phagocytophila infection has a primary role in HGE pathogenesis and immunomodulation.

Kim, Hyung-Yong; Mott, Jason; Zhi, Ning; Tajima, Tomoko; Rikihisa, Yasuko



Stress-associated modulation of proto-oncogene expression in human peripheral blood leukocytes.  


Changes in the cellular immune response associated with psychological stress were studied by using an academic stress model with medical students. The authors examined the expression of 2 proto-oncogenes, c-myc and c-myb, in peripheral blood leukocytes (PBLs) obtained from medical students at the time of examinations and at a baseline period approximately 1 month prior to the examinations. The level of messenger ribonucleic acid (mRNA) expression of both protooncogenes was significantly lower in PBLs obtained during examinations than in those from the baseline period. In addition, a significant decrease in the level of mRNA to the glucocorticoid receptor and gamma interferon was also found in the same preparations. The decrease in mRNA content of c-myc, c-myb, the glucocorticoid receptor, and gamma interferon in PBLs obtained from subjects during examinations is consistent with data from previous studies using the same model that have demonstrated a down-regulation of T-lymphocyte activation and proliferation in response to mitogens. PMID:8329139

Glaser, R; Lafuse, W P; Bonneau, R H; Atkinson, C; Kiecolt-Glaser, J K



Effects of resistance exercise and protein ingestion on blood leukocytes and platelets in young and older men  

Microsoft Academic Search

This study investigated, in a multi-experiment design, the acute effects of milk protein ingestion, aging [50 young (~26 years)\\u000a vs. 45 older (~61 years) men] and training state for the blood leukocyte and platelet responses acutely after a single bout\\u000a of resistance exercise (RE). Moreover, basal effects of 21 weeks of resistance training (RT) were examined. The single bout\\u000a of RE rapidly increased

Juha J. Hulmi; T. Myllymäki; M. Tenhumäki; N. Mutanen; R. Puurtinen; G. Paulsen; A. A. Mero



Leukocyte orchestration in blood and tumour tissue following interleukin-2 based immunotherapy in metastatic renal cell carcinoma  

Microsoft Academic Search

With the objective of evaluating leukocyte orchestration in situ, serial blood samples and tumour tissue core needle biopsies were obtained at baseline and repeated after 1 month of therapy, among 49 consecutive single-institution patients with metastatic renal cell carcinoma (mRCC). Patients were treated with outpatient low-dose subcutaneous interleukin 2 (IL-2) and interferon a (IFN-a) alone ( n=23) or in combination with

Frede Donskov; Karen Marie Bennedsgaard; Marianne Hokland; Niels Marcussen; Rune Fisker; Hans Henrik Torp Madsen; Kirsten Fode; Hans von der Maase



Prevalence of BK virus and JC virus in peripheral blood leukocytes and normal arterial walls in healthy individuals in China.  


Several studies have demonstrated that BK virus (BKV) and JC virus (JCV) establish latent infection in peripheral blood leukocytes (PBLs) of healthy individuals; however, the main populations studied are European. In this study, the prevalence of BKV and JCV DNA in PBLs from healthy adult individuals and umbilical cord blood from newborn children in China was detected by semi-nested polymerase chain reaction (snPCR) followed by restriction enzyme analysis. The results suggest that the healthy adult Chinese population harbors BKV and JCV DNA in peripheral leukocytes. Overall, the prevalence of BKV and JCV DNA in PBLs of healthy adult individuals was 42.1% and 7.8%, respectively. The overall prevalence of BKV DNA was significantly higher than that of JCV DNA. None of the umbilical cord blood samples from newborn children were positive for BKV and JCV DNA. To understand further the target tissues involved in establishment of BKV and JCV latency in healthy individuals, the presence of DNA from both viruses was detected in normal arterial wall samples from 20 young trauma victims by the same method used for leukocyte DNA. BKV DNA was detected alone in 20% of samples tested; JCV DNA was not detected alone in any of the samples. DNA from both viruses was found in 5% of samples. This is the first report to show that normal arterial walls of healthy individuals may be another target site of latency for BKV and JCV. PMID:12794723

Gu, Zhi-Yuan; Li, Qi; Si, Yi-Ling; Li, Xue; Hao, Hao-Jie; Song, Hai-Jing



Detection of cytomegalovirus from blood leukocytes separated by sepracell-MN and Ficoll-Paque/Macrodex methods.  

PubMed Central

Two density gradient separation techniques for separation of blood leukocytes were compared for the laboratory diagnosis of cytomegalovirus (CMV) viremia. Of 510 blood specimens processed by both methods, 76 (14.9%) yielded CMV. Of the 76 positive specimens, 66 (87%) and 65 (86%) were processed by the Ficoll-Paque/Macrodex (F-P/M; Macrodex is dextran 70 in normal saline; Pharmacia, Pisataway, N.J.) and Sepracell-MN methods, respectively. Of the 76 CMV-positive blood specimens, 72 (95%) were detected in shell vial cell cultures, whereas only 42 (55%) were detected in conventional tube cell cultures. The time for recognition of specific cytopathic effects due to CMV in tube cell cultures (8.0 versus 7.1 days), the number of fluorescent foci in each positive shell vial culture (19.3 versus 20.1), and the costs of the reagents ($3.50 versus $2.80) were similar and independent of the leukocyte separation method (F-P/M versus Sepracell-MN). Recovery of CMV from heparinized blood (F-P/M method) was similar to that from EDTA-anticoagulated blood (Sepracell-MN method). The Sepracell-MN method is a rapid and sensitive method for detection of CMV from blood specimens and is recommended as a replacement for the more tedious and time-consuming F-P/M procedure.

Paya, C V; Wold, A D; Smith, T F



Differential expression of intracellular and extracellular CB(2) cannabinoid receptor protein by human peripheral blood leukocytes.  


mRNA encoding for the CB(2) cannabinoid receptor is expressed by many subsets of human peripheral blood leukocytes (PBL), but little is known about the resulting protein expression and function. Employing clones from the A549 and 293T cell lines that were constructed to express both full-length human CB(2) and GFP, we developed a flow cytometry assay for characterizing CB(2) protein expression. A monoclonal antibody directed against human CB(2) selectively stained the surface of transduced but not parental cell lines. When cells were fixed and permeabilized, imaging flow cytometry identified large stores of intracellular protein. Total cellular staining for CB(2) corresponded closely with the level of GFP expression. When exposed to ?(9)-tetrahydrocannabinol, CB(2)-expressing cells internalized cell surface CB(2) receptors in a time- and dose-dependent manner. Applying these approaches to human PBL, CB(2) protein was identified on the surface of human B cells but not on T cells or monocytes. In contrast, when PBL were fixed and permeabilized, intracellular CB(2) expression was readily detected in all three subsets by both conventional and imaging flow cytometry. Similar to the protein expression pattern observed in fixed and permeabilized PBL, purified B cells, T cells, and monocytes expressed relatively equal levels of CB(2) mRNA by quantitative real-time RT-PCR. Our findings confirm that human PBL express CB(2) protein but that its distribution is predominantly intracellular with only B cells expressing CB(2) protein at the extracellular membrane. The differential role of intracellular and extracellular CB(2) receptors in mediating ligand signaling and immune function remains to be determined. PMID:23299999

Castaneda, Julie T; Harui, Airi; Kiertscher, Sylvia M; Roth, Jeffrey D; Roth, Michael D



Production of Interferon by Human and Animal Macrophages and Leukocytes.  

National Technical Information Service (NTIS)

Study of interferon production by human and animal phagocytic cells revealed that the process is more intense in cultures of macrophages than in cultures of polymorphonuclear leukocytes. The amount of interferon produced was found to be directly to the nu...

V. I. Rudenko A. A. Smorodintsev



Leukocyte, red blood cell and morphological adaptation to moderate physical training in rats undernourished in the neonatal period  

PubMed Central

Objective To analyze the impact of moderate physical exercise on the total and differential leukocyte counts and red blood cell count of 36 sixty-day-old adult male Wistar rats subjected to early malnourishment. Methods The rats were divided in nourished (N - casein 17%) and malnourished groups (M - casein 8%) and thesegroups were then subdivided in trained (T) untrained (U) creating four groups NT, NU, MT and MU. The NT and MTgroups were submitted to moderate physical exercise using a treadmill (60 min/day, 5 days/week for 8 weeks). Onthe 1st day, before the training started T0 and 24 hours after the last training day of the week (T1 until T8), a 1 mLaliquot of blood was collected from the animals' tails for analysis. The total leukocyte count was evaluated in a cellcounter with an electronic microscope. The cyanmethemoglobin technique was used to measure the hemoglobin level. The hematocrit values were determined as a percentage using the micro-hematocrit technique with a microcapillaryreader and a cell counter was used to determine the red blood cell count. The t-test was used for statistical analysis and a p-value < 0.05 was considered significant. Data are expressed as means ± standard deviation. Results There was a significant difference in the total leukocyte count between the NT (9.1 ± 0.1) and MT groups (8.0 ± 0.1) from T1 and in neutrophils between the NT (22.1 ± 0.6) and MT groups (24.6 ± 1.8) from T7 (p < 0.05). There was no statistical significance in the hemoglobin, hematocrit and red blood cell count from T1. Conclusions According to the results of this study, moderate physical exercise seems to have induced physiologic adaptation in adult rats from T1.

Viana, Marcelo Tavares; Perez, Manuella Cavalcanti; Ribas, Valdenilson Ribeiro; Martins, Gilberto de Freire; de Castro, Celia Maria Machado Barbosa



Comparison of metabolic, hematological, and peripheral blood leukocyte cytokine profiles of dairy cows and heifers during the periparturient period.  


The periparturient period presents major physiological challenges for the dairy cow. It is a period that is affected by metabolic stressors, major changes in endocrine status, and altered immune function, which together result in an increased risk of disease. Immunological, hematological, and metabolic profiles from the periparturient period of heifers (primipara) were compared with those of cows (pluripara) to test the hypothesis that at the time of calving they have qualitatively different peripheral blood profiles. Blood samples were collected from 22 Holstein-Friesian animals on 3 occasions: approximately 2 wk before calving, within 24h after calving, and approximately 2 wk after calving. Quantitative PCR was used to measure the expression of a selected set of cytokines and receptors by peripheral blood leukocytes. Additional analyses included hemoglobin concentration, red cell, platelet and white cell counts (total and differentiated), and clinical diagnostic biochemical profiles. Total leukocyte counts, neutrophils, and lymphocytes were higher in heifers than cows before calving and within 24h after calving. Alkaline phosphatase was consistently higher in heifers than cows and several significant differences were observed between the 2 groups with regards to cytokine and cytokine-receptor mRNA expression. The results warrant further investigation from the perspective of identifying risk factors for metabolic and parturient disease in dairy cattle. PMID:23462170

Jonsson, N N; Fortes, M R S; Piper, E K; Vankan, D M; de Cisneros, J Prada J; Wittek, T



Amifostine (WR2721) Confers DNA Protection to In Vivo Cisplatin-Treated Murine Peripheral Blood Leukocytes  

PubMed Central

Amifostine [S-2-3-aminopropil amino ethyl phosphorotioic acid], a modulator agent for antineoplastic drugs involved in free radicals generation has given controversial results in cisplatin treated leukocytes in vitro. We have evaluated the amifostine protection over leukocytes in vivo, using comet assay. Groups of five OF1 male mice were given one of three doses of amifostine (56, 105 and 200 mg/Kg) after a cisplatin single injection (10 mg/Kg). Serum malonyldialdehide levels, catalase and superoxide dismutase activity were also evaluated. Amifostine showed significant DNA protection (p< 0.01) at the two lower doses evaluated. Malonyldildehide decreased in all amifostine treatments with respect to cisplatin while antioxidant enzyme activities remained unchanged. However, DNA migration increased with the highest amifostine dose; in fact highest dose of amifostine did no protect damage caused by cisplatin this result have implications on amifostine treatment schedules in clinical practice.

Prieto Gonzalez, E. A.; Fuchs, A. G.; Sanchez, Gonzalez S.



Analysis of the thiol status of peripheral blood leukocytes in rheumatoid arthritis patients  

Microsoft Academic Search

Although the exact etiology of rheuma- toid arthritis (RA) remains unknown, there is in- creasing evidence that reactive oxygen species and a pro-oxidant\\/antioxidant imbalance are an impor- tant part of the pathogenesis of joint tissue injury. Flow cytometry was used to evaluate the thiol sta- tus (surface-thiols and intracellular glutathione (iGSH)) of leukocytes from RA patients and con- trols. Levels

Robert B. Zurier; David A. Lawrence



Kinetics of peripheral blood leukocyte alterations in Thai children with dengue hemorrhagic fever.  

PubMed Central

Peripheral leukocytes from 16 Thai children with dengue hemorrhagic fever were examined to determine the leukocyte composition on the day of presentation and on convalescent days 15 and 30. Mononuclear cells were isolated each time, and the concentrations of T, B, Fc receptor-bearing, and "null" cells were determined. On the day of hospitalization, in comparison to convalescent values, there was a significant increase in total lymphocytes, primarily due to concentrations of atypical lymphocytes. There was a significant loss of T cells with an increase in non-T, non-B, non-Fc receptor-bearing null cells. There were no changes in the concentrations of monocytes, B cells, or Fc receptor-bearing cells when acute and convalescent values were compared. During the convalescent period, a progressive increase in eosinophils was noted. Also, on day 15 but not on day 30 of the convalescent period, an increase was observed in the total leukocyte number due to an increase in granulocytes. There results indicate that in Thai children with dengue hemorrhagic fever, there are major shifts within several component cell subpopulations of the immune system.

Wells, R A; Scott, R M; Pavanand, K; Sathitsathein, V; Cheamudon, U; Macdermott, R P



Inhibition of peripheral blood neutrophil oxidative burst in periodontitis patients with a homeopathic medication Traumeel S  

PubMed Central

Summary Background The anti-inflammatory effects of a homeopathic remedy, Traumeel S, have been observed in experimental and clinical studies; however, its antioxidant properties have not been elucidated. The aim of the present study was to evaluate the antioxidant effects of Traumeel S on peripheral blood neutrophils in patients with periodontitis. Material/Methods The study was performed using venous blood of 22 individuals with chronic periodontitis and 21 healthy subjects. The antioxidant effects of Traumeel S on the production of reactive oxygen species by unstimulated and stimulated with unopsonized E. coli neutrophils were investigated using luminol- and lucigenin-dependent chemiluminescence (CL). Results Polymorphonuclear leukocytes of periodontitis patients produced higher levels (p<0.01) of light output of lucigenin-dependent chemiluminescence and significantly reduced (p<0.01) light output of luminol-dependent chemiluminescence than analogous cells of healthy subjects. Highly diluted (10?4 of the stem solution) Traumeel S significantly (by approximately 50%) reduced superoxide-induced oxidation of lucigenin by unstimulated and stimulated with unopsonized E. coli polymorphonuclear leukocytes of periodontitis patients and had a tendency to intensify luminol-dependent chemiluminescence. Preincubation of the unstimulated and stimulated with unopsonized E. coli polymorphonuclear leukocytes of healthy subjects with Traumeel S exerts no inhibitory action on the luminol- and lucigenin-dependent chemiluminescence of the above-mentioned cells. Conclusions This study indicates that Traumeel S may significantly reduce production of superoxide anion by unstimulated and stimulated peripheral blood polymorphonuclear neutrophils of periodontitis patients.

zilinskas, Juozas; zekonis, Jonas; zekonis, Gediminas; Sadzeviciene, Renata; Sapragoniene, Marija; Navickaite, Justina; Barzdziukaite, Ingrida



Development of methods to examine the effects of atmospheric particulate matter (PM) on human peripheral blood leukocytes  

NASA Astrophysics Data System (ADS)

In vitro methods to study the effect of atmospheric particulate matter (PM) on leukocyte function using human peripheral blood were developed. These methods were demonstrated using the blood of 1-5 individuals and National Institute of Standards and Technology (NIST) urban PM #1648, diesel PM #1650, silica PM, and a locally collected PM sample (New Jersey PM10). For the blood samples analyzed in this study NIST urban PM and New Jersey PM10 treatment mediated the release of granule contents from peripheral blood leukocytes and induced structural changes associated with degranulation. Flow cytometry revealed PM-induced changes in phagocytosis and cell structure associated with degranulation. Transmission electron microscopy confirmed NIST urban PM-induced cell structure changes were associated with PM internalization. Colorametric and electrophoretic methods showed no PM-induced release of primary granules and a slight PM-induced release of secondary granules associated with only NIST urban PM. Enzyme Immunosorbent Assays detected increased histamine release from basophils treated with NIST urban PM, a locally collected PM, and the soluble and insoluble components of these particles. NIST urban PM was found to be a potent inducer of histamine release in 4 out of 6 individuals tested. Fractionation studies revealed that soluble (aqueous) and insoluble fractions of NIST urban PM contain histamine-releasing activity. This was also demonstrated for the New Jersey PM10 sample for which the soluble fraction exhibited the most activity. Complementary studies with inhibitors of IgE-mediated histamine release conducted on one test subject suggest that PM-induced histamine release was partially mediated by IgE. A new hypothesis has been formed, suggesting that particle toxicity is related to PM-induced histamine release. Due to the bioactive nature of histamine and its association with many cardiopulmonary responses, the PM- mediated release of histamine should be investigated further.

Zussman, Lisa Ann


[The importance of studying the acid phosphatase of the blood serum and bone marrow lymphoblasts and polymorphonuclear neutrophils in the prognosis of the course of acute lymphoblastic leukemia].  


The activity of serum acid phosphatase (AP), bone marrow lymphoblasts and polymorphonuclear neutrophils was studied in 45 ALL patients. Cytochemical coefficients (CCC) and the percentage of positively reacting bone marrow cells were determined. All the patients received programmed polychemotherapy. They were investigated before the start of therapy, during recurrence and at different time of remission (from 1 to 60 mos) during each reinduction cycle. At the climax of ALL the activity of serum AP was increased 2.8-fold, a CCC value for lymphoblastic AP--10-fold, for polymorphonuclear neutrophils--3-fold as compared with normal values. A tendency toward the reduction of indices was noted at different time of remission, the approximation to normal values was noted on the 40th-46th months of remission only. In recurrence development the level of the serum and cellular enzyme as well as the percentage of positively reacting cells significantly exceeded normal values and were close to indices at the climax of disease. The above tendency permitted the use of these tests to evaluate the completeness of remission and to predict recurrences during a follow-up of ALL patients. PMID:3175926

Vaiuta, N P; Kha?fets, L M; Mendeleev, I M



Comparison of sample fixation and the use of LDS-751 or anti-CD45 for leukocyte identification in mouse whole blood for flow cytometry.  


Flow cytometry methods used to measure leukocyte function often entail sample preparation procedures that cause artifactual cell activation. To avoid leukocyte activation by isolation techniques, some preparation methods use fluorescent markers to discriminate leukocytes from erythrocytes in whole blood. One of these markers, laser dye styryl-751(LDS-751), has been used to distinguish leukocytes by staining nucleic acid, but has been found to stain other blood cells and dead cells indiscriminately. Thus, LDS-751 may not be an appropriate reagent for leukocyte identification in whole blood. Fixing samples with formaldehydes increases cell permeability and causes surface protein cross-linking that may alter staining of both intra- and extracellular markers. The degree of this sample alteration by formaldehyde fixation, however, remains in question. In addition, little is known about flow cytometry and sample preparation methods in mouse whole blood. The purpose of this study was to determine if labeling leukocytes with a monoclonal antibody specific to leukocyte common antigen (CD45) was superior to labeling with LDS-751 and to determine the effect of sample fixation on a mouse whole blood preparation for flow cytometry. Samples were incubated with CD16/CD32 Fc receptor blocker, and either 10 microg/ml LDS-751 or phosphate buffered saline (PBS). The samples were then fixed with paraformaldehyde or diluted with PBS followed by incubation with 5 microg/ml PerCP-conjugated anti-CD45, 5 microg/ml FITC-conjugated anti-CD11b, or 80 microM dichlorofluorescein diacetate. We found that samples labeled with LDS-751 demonstrated decreased fluorescence intensity for granulocyte CD11b expression and ROS production compared to samples labeled with anti-CD45. In addition, sample fixation decreased mean fluorescence intensity in samples labeled with either LDS-751 or anti-CD45. We conclude that labeling leukocytes with monoclonal antibody CD45 in a mouse whole blood preparation is preferable, as it provides improved measurement of leukocyte indices compared to LDS-751. Also, while sample fixation prior to antibody staining caused a decrease in overall fluorescence; it can be used to successfully identify extra-cellular markers. PMID:17187818

Maes, Melissa L; Davidson, Lisa B; McDonagh, Paul F; Ritter, Leslie S



Fasudil, a Rho-kinase inhibitor, inhibits leukocyte adhesion in inflamed large blood vessels in vivo  

Microsoft Academic Search

.\\u000a Objective and Design  Emerging data suggest that Rho-kinase signaling may regulate numerous aspects of inflammatory reactions. Herein, we investigated\\u000a the role of Rho-kinase in inflammatory interactions between leukocytes and the endothelium in femoral arteries and veins in vivo.\\u000a \\u000a \\u000a \\u000a Material and methods  Mice were injected with lipopolysaccharide (LPS) and Rho-kinase was inhibited by pre-treatment with fasudil, which is a highly\\u000a selective inhibitor

J. E. Slotta; O. Ö. Braun; M. D. Menger; H. Thorlacius



Influence of the red blood cell preparation method on the efficacy of a leukocyte reduction filter.  


The performance of a leukocyte reduction bedside filter with different types of RBC concentrates was analyzed. Three types of RBCs were prepared: buffycoat-depleted RBCs suspended in saline-adenine-glucose-mannitol (SAGM)-additive solution (BC-RBCs; n = 20), RBCs suspended in SAGM-additive solution without buffy coat removal (SAGM-RBCs; n = 20), and RBCs drawn in CPDA-I conservative solution and processed for component preparation by the platelet-rich plasma method (CPDA-RBCs; n = 20). The units were filtered within 8 h of collection. One filter was used for every 2 units. High numbers of residual WBCs were found even in the units filtered first. Filtration of CPDA-RBCs resulted in a higher residual WBC content than SAGM-RBCs or BC-RBCs (p = 0.0032 and p = 0.0002, respectively). The filter performance strikingly decreased when the WBC load per filter exceeded 4 x 10(9) or the platelet load was less than 100 x 10(9). We conclude that filter performance varies with the WBC and platelet content of the RBC concentrates. Under the experimental conditions assayed in this study CPDA-RBCs are the least appropriate ones to be used for bedside leukocyte reduction. PMID:8873416

Alcorta, I; Pereira, A; Sanz, C; Terol, M J; Ordinas, A



Immunomodulatory effect of the homoeopathic drug Engystol-N on some activities of isolated human leukocytes and in whole blood.  


Engystol-N at the doses of 10(-4) and 10(-8) in isolated human leukocytes stimulates the superoxide anion generation by neutrophils and the cytokine(s) production by T lymphocytes. In whole blood the same concentrations of the drug produce the decrease of the superoxide anion generation of neutrophils, this inhibiting activity appears 6 h after the administration of the drug and persists only in presence of lymphocytes. Culture media of T lymphocytes treated with Engystol-N show the same inhibiting effect on superoxide anion generation by neutrophils. From these data it is possible to conclude that the drug stimulates the secretion of lymphokine(s) with inhibiting action on superoxide anion generation of neutrophils that prevail over the direct stimulating effect, confirming and extending the immunomodulatory ability of the drug. PMID:10737260

Fimiani, V; Cavallaro, A; Ainis, O; Bottari, C



Leukocyte Filtration Does Not Affect Lymphocyte Subpopulations and NK Cell Function in Recipients of Blood Transfusions  

Microsoft Academic Search

Background and objectives: The possible immunosuppressive action of blood transfusion has aroused great interest recently, particularly with respect to its effects on tumor growth and recurrence rate of malignant disease. Materials and methods: The effect of blood transfusion on lymphocyte subpopulations and NK cell function preoperatively and 6 months postoperatively was studied in 129 patients treated with elective surgery for

O. Mathiesen; L. Lund; U. Brodthagen; P. Gandrup; N. Grunnet; I. Balslev; C. Jersild



Increased soluble human leukocyte antigen-G levels in peripheral blood from climbers on Mount Everest.  


Soluble human leukocyte antigen-G (HLA-G) is involved in maternal-fetal tolerance, transplant acceptance, and tumor escape from immunosurveillance, operating by inhibiting activity of T, antigen presenting cells (APC), and natural killer (NK) cells. HLA-G gene expression is modulated in vitro after hypoxic conditions, a situation evidenced during pregnancy and tumor progression. In extreme altitude, mountaineers are in hypoxic conditions that generate physiologic adaptative responses, some of them giving rise to pathologic signs. We performed measurements of plasma soluble HLA-G in six climbers before departure of the expedition and during their ascent to and descent from summit of Mount Everest, and in 3 Sherpas at 5300-6400 m. We found that HLA-G levels are upregulated during the ascent with a unique pattern in comparison with angiogenic/lymphangiogenic factors. Our data suggest that HLA-G has to be taken into account in the mechanisms participating in adaptation to high altitudes and reinforce hypoxia as an important factor in the regulation of HLA-G expression. PMID:20732367

Bourguignon, Michel; Yaghi, Layale; Flajollet, Sébastien; Radanne-Krawice, Irène; Rouas-Freiss, Nathalie; Lugrin, Didier; Richalet, Jean-Paul; Carosella, Edgardo D; Moreau, Philippe



Increased anticoagulant osmolality improves separation of leukocytes from red blood cells (RBC)  

Microsoft Academic Search

Background: The bottom-and-top (BAT) procedure separates the buffy coat (BC) from plasma and red blood cells (RBC). The contents of mononuclear cells (MNC) remaining in the RBC are about 1×106 cells\\/unit, whereas the granulocytes are removed less effectively, 500–800×106 or more remaining in the RBC unit. The aim was to improve the separation efficacy by collecting the blood in an

F Knutson; H Lööf; C. F Högman



Cell-type specific gene expression profiles of leukocytes in human peripheral blood  

Microsoft Academic Search

BACKGROUND: Blood is a complex tissue comprising numerous cell types with distinct functions and corresponding gene expression profiles. We attempted to define the cell type specific gene expression patterns for the major constituent cells of blood, including B-cells, CD4+ T-cells, CD8+ T-cells, lymphocytes and granulocytes. We did this by comparing the global gene expression profiles of purified B-cells, CD4+ T-cells,

Chana Palmer; Maximilian Diehn; Ash A Alizadeh; Patrick O Brown



Red Blood Cell Size Is Inversely Associated with Leukocyte Telomere Length in a Large Multi-Ethnic Population  

PubMed Central

Although mutations in the genes encoding either the protein or RNA component of telomerase have been found in patients with various blood disorders, the impact of telomere length on hematopoiesis is less well understood for subjects from the general population. Here we have measured telomere lengths of genomic DNA isolated from circulating leukocytes of 3157 subjects, ranging from 18 to 85 years of age, enrolled in a large multiethnic population based study, the Dallas Heart Study 2. Shorter telomere lengths are marginally associated with lower red blood cell counts in this cohort, but are significantly associated with larger mean red blood cell size (as measured by the MCV), increased red blood cell distribution width (RDW), higher hemoglobin levels and lower platelet counts, even after correction for age, gender and ethnicity (p-values of <0.0001, <0.0001, 0.0009 and 0.0016, respectively). In a multiple regression model we find that telomere length is a significant covariate of MCV (p?=?7.6×10?8), independent of age, ethnicity, BMI, current smoking, alcohol consumption, iron or homocysteine levels. The effect of telomere length on MCV variation is comparable to the effect of smoking or alcohol consumption and is more significant in older individuals (p?=?9.2×10?7 for >50 years vs. p?=?0.0006 for <50 years of age). To our knowledge, this is the first report of an association between telomere length and red cell size in a large urban US population and suggests a biologic mechanism for macrocytosis of aging.

Kozlitina, Julia; Garcia, Christine Kim



Effects of immunoglobulin binding on signal transduction in bovine polymorphonuclear neutrophils  

Technology Transfer Automated Retrieval System (TEKTRAN)

Immunoglobulins are major molecules that mediate humoral immune responses. Their functional effects on leukocytes are mediated by the cell surface receptors for the Fc domain of immunoglobulins (FcR). Ligation of FcR on human polymorphonuclear neutrophils (PMN) is capable of triggering a wide rang...


Microfluidic lysis of human blood for leukocyte analysis using single cell impedance cytometry.  


This paper demonstrates an integrated microfluidic system that performs a full blood count using impedance analysis. A microfluidic network design for red blood cell (RBC) lysis is presented, and the diffusive mixing processes are analyzed using experimental and simulated results. Healthy and clinical bloods analyzed with this system, and the data shows good correlation against data obtained from commercial hematology machines. The data from the microfluidic system was compared against hospital data for 18 clinical samples, giving R(2) (coefficient of determination) values of 0.99 for lymphocytes, 0.89 for monocytes, and 0.99 for granulocytes in terms of relative counts and 0.94 for lymphocytes, 0.91 for monocytes, and 0.95 for granulocytes in terms of absolute counts. This demonstrates the potential clinical utility of this new system for a point-of-care purpose. PMID:22148390

Han, Xiaojun; van Berkel, Cees; Gwyer, James; Capretto, Lorenzo; Morgan, Hywel




Microsoft Academic Search

When rabbit peritoneal exudates (97 % polymorphonuclear (PMNJ leukocytes, 2% mono- nuclear cells) were fractionated by zonal sedimentation or isopycnic centrifugation, four fractions (A, B, C, and D) were obtained, as reported earlier. \\




Impact of pre-storage and bedside filtered leukocyte-depleted blood transfusions on infective morbidity after colorectal resection: a single-center analysis of 437 patients.  


Abstract Background: Leukocyte-depleted blood transfusions were introduced to reduce transfusion-associated immunomodulation, but the clinical effects of different types of leukocyte depletion have been analyzed rarely. The aim of this survey was to analyze the clinical impact of pre-storage leukocyte-depleted blood transfusions (considered as pre-storage or bedside-filtered) on post-operative complications in patients undergoing elective or urgent colorectal resection. Methods: Data were collected retrospectively from the medical records of 437 consecutive patients who underwent colorectal resection from 2005 to 2010. All patients requiring transfusion received pre-storage or bedside-filtered leukocyte-depleted red blood cell concentrates according to availability at the blood bank. The outcomes were measured by the analysis of post-operative morbidity in patients receiving the different types of transfusions or having other potentially predictive risk factors. Results: The overall morbidity rate, infective morbidity rate, and non-infective morbidity rate were, respectively, 35.6%, 28.1%, and 21.0%. Two hundred five patients (46.9%) received peri-operative transfusions. On multivariable analysis, leukocyte-depleted transfusion (odds ratio [OR] 3.33; 95% confidence interval [CI] 2.14-5.20; p<0.001) and both pre-storage (OR 2.82; 95% CI 1.73-4.59; p<0.001) and bedside-filtered (OR 4.69; 95% CI 2.54-8.67; p<0.001) transfusions were independent factors for post-operative morbidity. Prolonged operation (p=0.035), American Society of Anesthesiologists score?3 points (p=0.023), diagnosis of cancer rather than benign disease (p=0.022), and urgent operation (p=0.020) were other independent predictors of post-operative complications. Patients transfused with bedside-filtered blood showed significantly higher rates of infective complications (51.4% vs. 31.8%; p=0.006), but not non-infectious complications (35.7% vs. 32.6; p=0.654) than patients who received pre-storage transfusions. Conclusions: Leukocyte-depleted blood transfusions and, in particular, bedside-filtered blood have a significant negative effect on infectious complications after colorectal resection. PMID:23859683

Garancini, Mattia; Degrate, Luca; Carpinelli, Maria Rosaria; Maternini, Matteo; Uggeri, Fabio; Giordano, Laura; Uggeri, Franco; Romano, Fabrizio



Leukocyte count affects expression of reference genes in canine whole blood samples  

PubMed Central

Background The dog is frequently used as a model for hematologic human diseases. In this study the suitability of nine potential reference genes for quantitative RT-PCR studies in canine whole blood was investigated. Findings The expression of these genes was measured in whole blood samples of 263 individual dogs, representing 73 different breeds and a group of 40 mixed breed dogs, categorized into healthy dogs and dogs with internal and hematological diseases, and dogs that underwent a surgical procedure. GeNorm analysis revealed that a combination of 5 to 6 of the most stably expressed genes constituted a stable normalizing factor. Evaluation of the expression revealed different ranking of reference genes in Normfinder and GeNorm. The disease category and the white blood cell count significantly affected reference gene expression. Conclusions The discrepancy between the ranking of reference genes in this study by Normfinder and Genorm can be explained by differences between the experimental groups such as "disease category" and "WBC count". This stresses the importance of assessing the expression stability of potential reference genes for gene experiments in canine whole blood anew for each specific experimental condition.



Peripheral Blood Leukocyte Production of BDNF following Mitogen Stimulation in Early Onset and Regressive Autism  

Microsoft Academic Search

Brain-derived neurotrophic factor (BDNF) is critical for neuronal differentiation and synaptic development. BDNF is also implicated in the development of psychological disorders including depression, bipolar disorder and schizophrenia. Previously, elevated BDNF levels were observed in neonatal blood samples from infants who were later diagnosed with autism when compared with children who developed normally, suggesting that BDNF may be involved in

Amanda Enstrom; Charity Onore; Angela Tarver; Irva Hertz-Picciotto; Robin Hansen; Lisa Croen



Systemic suppression of human peripheral blood phagocytic leukocytes after whole-body UVB irradiation  

Microsoft Academic Search

We examined systemic effects of whole- body UVB irradiation on human peripheral blood phagocytes. We found that 24 h after a single erythemal dose of UVB radiation two phagocyte functions, adhesion and phagocytosis, were re- duced by 50%. This functional suppression was accompanied by a significant decrease in the expres- sion of complement receptors (CR1 and CR3) and IgG Fc

Lasse Leino; Kustaa Saarinen; Kaisa Kivisto; Leena Koulu; Christer T. Jansen; Kari Punnonen


Analysis of leukocyte differentiation antigens in blood and bone marrow from preleukemia (refractory anemia) patients using monoclonal antibodies.  


Peripheral blood and bone marrow mononuclear cells from patients with refractory anemia (RA) or RA with sideroblasts (defined according to the revised French-American-British classification with less than 5% blast cells in the bone marrow) were analyzed using a panel of monoclonal antibodies directed against leukocyte antigens on B lymphocytes, T lymphocytes, monocytes, and myeloid cells. In the peripheral blood an increased proportion of T lymphocytes (and correspondingly a decreased proportion of B cells) could be demonstrated. However, when expressed in terms of absolute numbers, the T cell component was depressed because of severely decreased numbers of T4+ helper cells. In contrast, the absolute numbers of T8+ suppressor cells were either normal or increased in the majority of the patients. This resulted in markedly decreased ratios of T4+/T8+ cells, which were closely correlated to the number of transfusions given to the patients because of their refractory anemia. Finally, nearly all of the patients exhibited decreased numbers of cells reactive with the N901 natural killer (NK) antibody, thus explaining our earlier finding of decreased NK activity in these patients. In the bone marrow increased proportions of myeloid cells reactive with monoclonal antibodies present on immature myeloid cells (My7 and My9) were found, suggesting the presence of malignant clones. Indeed, when the numbers of My7+ cells and the morphologic evaluations of bone marrow smears at the time of diagnosis were compared to the progression of the disease, a group of patients with high numbers of My7+ cells and normal morphology could be identified that had a high probability of progression to refractory anemia with an excess of blasts or to overt acute myeloid leukemia. Thus, the use of antibodies defining leukocyte differentiation antigens might be of significant value in the diagnosis and prognostication of the myelodysplastic syndromes. These findings are discussed in relation to the pathogenesis of this potentially premalignant condition with special emphasis on possible defects in the immunologic defense mechanisms against early neoplasias. PMID:3082390

Hokland, P; Kerndrup, G; Griffin, J D; Ellegaard, J



Flow Cytometric Analysis of Porcine Peripheral Blood Leukocytes Infected With Pseudorabies Virus  

Microsoft Academic Search

The susceptIbilIty of fractionated porcine peripheral blood ieukocytes (PBL) to pseudo- rabies virus (PRV) was studied by flow cytometry and defined by viral antigen expres- sion. Viral antigens on the surface of Infected cells and cell viability were evaluated by forward angie light scatter (FALS), 90-degree light scatter (9OLS), green fiuorescence (FITC-antl-PRV), and red fluorescence (propidium Iodide). Approximately 10% of

Fun-In Wang; Victor F. Pang; Edwin C. Hahn


[Influence of vibration-induced stress on functional-metabolic status of blood leukocytes].  


The chronic stress in albino rats caused by exposure to the whole-body vibration induced the significant changes in the functional-metabolic status of the blood cells. It involved the phagocytosis level and the lysosomal cationic proteins in the neutrophils, oxidative and hydrolytic processes in the neutrophils and lymphocytes. All the determined intracellular parameters revealed the differentiated response to stress as well as to the additive combined administration of the antioxidants (glycine and alpha-tocopherol acetate). PMID:23650727

Dolgushin, M V; Davydova, N S



Microsoft Academic Search

Emerging evidence indicates that chemokine receptor expression patterns are critical in determining the spectrum of action of the chemokines. We have analysed the expression patterns of 17 chemokine receptors and two orphan chemokine receptor-like genes in various freshly prepared human peripheral blood leucocyte populations, including neutrophils, lymphocytes, and na??ve and differentiated monocytes using real-time quantitative polymerase chain reaction (TaqMan®). This

L. Patel; S. J. Charlton; J. K. Chambers; C. H. Macphee



An extended convection diffusion model for red blood cell-enhanced transport of thrombocytes and leukocytes  

NASA Astrophysics Data System (ADS)

Transport phenomena of platelets and white blood cells (WBCs) are fundamental to the processes of vascular disease and thrombosis. Unfortunately, the dilute volume occupied by these cells is not amenable to fluid-continuum modeling, and yet the cell count is large enough that modeling each individual cell is impractical for most applications. The most feasible option is to treat them as dilute species governed by convection and diffusion; however, this is further complicated by the role of the red blood cell (RBC) phase on the transport of these cells. We therefore propose an extended convection-diffusion (ECD) model based on the diffusive balance of a fictitious field potential, ?, that accounts for the gradients of both the dilute phase and the local hematocrit. The ECD model was applied to the flow of blood in a tube and between parallel plates in which a profile for the RBC concentration field was imposed and the resulting platelet concentration field predicted. Compared to prevailing enhanced-diffusion models that dispersed the platelet concentration field, the ECD model was able to simulate a near-wall platelet excess, as observed experimentally. The extension of the ECD model depends only on the ability to prescribe the hematocrit distribution, and therefore may be applied to a wide variety of geometries to investigate platelet-mediated vascular disease and device-related thrombosis.

Hund, S. J.; Antaki, J. F.



To?pa Torf Preparation (TTP) induces interferon and tumor necrosis factor production in human peripheral blood leukocytes.  


To?pa Torf Preparation (TTP) is a natural immunomodulating drug registered in Poland for use in humans. TTP is a biologically active low molecular weight fraction of an extract from peat containing organic substances, primary bound sugars, amino-acids, uronic and huminic acids and mineral salts. The toxicity of TTP is remarkably low, eg. cytotoxicity (CD50) for human peripheral blood leukocytes (PBL) is 1-9 mg/ml. We have discovered that TTP is an interferon (IFN) and tumor necrosis factor (TNF) inducer in human PBL. The IFN and TNF response of the PBL cultures was dose dependent. The optimal concentration of TTP for IFN or TNF response was 10-100 micrograms/ml. The cytokines stimulated by TTP were IFN-gamma, IFN-alpha and TNF-alpha. Ten commercial batches of TTP have been found to be active as cytokines inducers although variations in their activities were observed. On the other hand, 8 batches of TTP rejected by the producer because of the inadequate immunostimulating activity determined in mice, were found to be significantly less active than the commercial preparations. Over 115 buffy coats from the individual blood donors were used to prepare PBL cultures for this study. Approximately 20% of the PBL cultures were unresponsive to TTP. The IFN and TNF response of PBL to other inducers: phytohemagglutinin (PHA) or lipopolysaccharides (LPS) also varied. Whereas only 7% of PBL could not be stimulated by PHA, as much as 20-50% of PBL failed to produce IFN or TNF when treated with LPS. We suggest that TTP may have clinically useful activities connected with the capacity of stimulation of IFNs and TNF production. PMID:7694559

Inglot, A D; Zieli?ska-Jenczylik, J; Piasecki, E



In Vitro Study of Interactions between Silicon-Containing Nanoparticles and Human Peripheral Blood Leukocytes.  


The effects of silicon dioxide-based nanoparticles on the viability and proliferative activity of human peripheral blood cultured lymphocytes were studied. All nanoparticles in a concentration of 100 ?g/ml produced a significant cytotoxic effect, its intensity depending on particles' structure: SiO2 nanoparticles were least toxic, while Ce3(+)-intercaled montmorillonite nanoparticles were most toxic. The cells died mainly by apoptosis and postapoptotic necrosis. Incubation with nanoparticles in a concentration of 100 ?g/ml for 72 h caused death of all phytohemagglutinin-activated lymphocytes, while in concentrations of 1 and 10 ?g/ml the nanoparticles had no effect of proliferative activity of cells. The results suggest that the effects of nanoparticles on cells are determined by the nanoparticle concentration and size, as well as by their structure. PMID:24137611

Andreeva, E R; Rudimov, E G; Gornostaeva, A N; Beklemyshev, V I; Makhonin, I I; Maugeri, U O G; Buravkova, L B



Full blood count and haemozoin-containing leukocytes in children with malaria: diagnostic value and association with disease severity  

PubMed Central

Background Diligent and correct laboratory diagnosis and up-front identification of risk factors for progression to severe disease are the basis for optimal management of malaria. Methods Febrile children presenting to the Medical Research Unit at the Albert Schweitzer Hospital (HAS) in Lambaréné, Gabon, were assessed for malaria. Giemsa-stained thick films for qualitative and quantitative diagnosis and enumeration of malaria pigment, or haemozoin (Hz)-containing leukocytes (PCL) were performed, and full blood counts (FBC) were generated with a Cell Dyn 3000® instrument. Results Compared to standard light microscopy of Giemsa-stained thick films, diagnosis by platelet count only, by malaria pigment-containing monocytes (PCM) only, or by pigment-containing granulocytes (PCN) only yielded sensitivities/specificities of 92%/93%; 96%/96%; and 85%/96%, respectively. The platelet count was significantly lower in children with malaria compared to those without (p < 0.001), and values showed little overlap between groups. Compared to microscopy, scatter flow cytometry as applied in the Cell-Dyn 3000® instrument detected significantly more patients with PCL (p < 0.01). Both PCM and PCN numbers were higher in severe versus non-severe malaria yet reached statistical significance only for PCN (p < 0.0001; PCM: p = 0.14). Of note was the presence of another, so far ill-defined pigment-containing group of phagocytic cells, identified by laser-flow cytometry as lymphocyte-like gated events, and predominantly found in children with malaria-associated anaemia. Conclusion In the age group examined in the Lambaréné area, platelets are an excellent adjuvant tool to diagnose malaria. Pigment-containing leukocytes (PCL) are more readily detected by automated scatter flow cytometry than by microscopy. Automated Hz detection by an instrument as used here is a reliable diagnostic tool and correlates with disease severity. However, clinical usefulness as a prognostic tool is limited due to an overlap of PCL numbers recorded in severe versus non-severe malaria. However, this is possibly because of the instrument detection algorithm was not geared towards this task, and data lost during processing; and thus adjusting the instrument's algorithm may allow to establish a meaningful cut-off value.

Hanscheid, Thomas; Langin, Matthias; Lell, Bertrand; Potschke, Marc; Oyakhirome, Sunny; Kremsner, Peter G; Grobusch, Martin P



Coincident Activation of Th2 T Cells with Onset of the Disease and Differential Expression of GRO-Gamma in Peripheral Blood Leukocytes in Minimal Change Disease  

Microsoft Academic Search

Background: Involvement of Th2 T cells\\/NF?B in minimal change disease (MCD) has been postulated. A promising but unconfirmed glomerular permeability factor (GPF) from MCD T cells has been described. We explored whether GPF was the consequence of Th2 cell activation. Methods: Peripheral blood leukocytes (PBL) from 16 MCD patients and 7 normal controls were analyzed and the results were statistically

Horacio E. Adrogue; Jason Borillo; Lisa Torres; Arundhati Kale; Cindy Zhou; Daniel Feig; Justin Merszei; Richard Johnson; Ya-Huan Lou



Is increased mortality associated with post-operative infections after leukocytes containing red blood cell transfusions in cardiac surgery? An extended analysis  

Microsoft Academic Search

In two randomized trials in cardiac surgery we observed that leukoreduced allogeneic red blood cell (RBC) transfusions (LR) compared with standard buffy-coat-depleted RBC transfusions (BCD) resulted in lower rates of post-operative infections and mortality. To unravel whether this comprises two independent side effects or could be related complications of allogeneic leukocytes, we performed a re-analysis on the patients of these

Y. M. Bilgin; L. M. G. van de Watering; L. Eijsman; M. I. M. Versteegh; M. H. J. van Oers; A. Brand



Leukocyte-depletion of blood components does not significantly reduce the risk of infectious complications. Results of a double-blinded, randomized study.  


Allogeneic blood transfusions are claimed to be an independent risk factor for postoperative infections in open colorectal surgery due to immunomodulation. Leukocyte-depletion of erythrocyte suspensions has been shown in some open randomized studies to reduce the rate of postoperative infection to levels observed in nontransfused patients. Using a double-blinded, randomized design, we studied the postoperative infection rate in patients undergoing open colorectal surgery transfused with either leukocyte-depleted erythrocyte suspensions (LD-SAGM) or non-leukocyte-depleted erythrocyte suspensions (SAGM). Unselected patients (n 279) were allocated to receive LD-SAGM (n 139) or SAGM (n 140) if transfusion was indicated. Forty-five percent were transfused, yielding 48 patients in the LD-SAGM group and 64 in the SAGM group. Thirteen patients were excluded because they received one type of transfusion in spite of randomization to the other type. No significant differences in the rates of postoperative infections (P=0.5250) or postoperative complications (P=0.1779) were seen between the two transfused groups. Infection rates were 45% and 38% in the transfused groups and 21% and 23% in the nontransfused groups. No significant difference between the transfused groups was seen on any single infectious event, mortality rate, or duration of hospitalization. Leukocyte-depletion of erythrocyte suspensions transfused to patients undergoing open colorectal surgery does not reduce postoperative infection rates. PMID:11459288

Titlestad, I L; Ebbesen, L S; Ainsworth, A P; Lillevang, S T; Qvist, N; Georgsen, J



Selective activation of cannabinoid receptor 2 in leukocytes suppresses their engagement of the brain endothelium and protects the blood-brain barrier.  


Cannabinoid receptor 2 (CB2) is highly expressed in immune cells and stimulation decreases inflammatory responses. We tested the idea that selective CB2 activation in human monocytes suppresses their ability to engage the brain endothelium and migrate across the blood-brain barrier (BBB), preventing consequent injury. Intravital videomicroscopy was used to quantify adhesion of leukocytes to cortical vessels in lipopolysaccharide-induced neuroinflammation, after injection of ex vivo CB2-activated leukocytes into mice; CB2 agonists markedly decreased adhesion of ex vivo labeled cells in vivo. In an in vitro BBB model, CB2 activation in monocytes largely attenuated adhesion to and migration across monolayers of primary human brain microvascular endothelial cells and diminished BBB damage. CB2 stimulation in monocytes down-regulated active forms of integrins, lymphocyte function-associated antigen 1 (LFA-1), and very late antigen 4 (VLA-4). Cells treated with CB2 agonists exhibited increased phosphorylation levels of inhibitory sites of the actin-binding proteins cofilin and VASP, which are upstream regulators of conformational integrin changes. Up-regulated by relevant stimuli, Rac1 and RhoA were suppressed by CB2 agonists in monocytes. CB2 stimulation decreased formation of lamellipodia, which play a key role in monocyte migration. These results indicate that selective CB2 activation in leukocytes decreases key steps in monocyte-BBB engagement, thus suppressing inflammatory leukocyte responses and preventing neuroinflammation. PMID:24055259

Rom, Slava; Zuluaga-Ramirez, Viviana; Dykstra, Holly; Reichenbach, Nancy L; Pacher, Pal; Persidsky, Yuri



Exposure to Metal-Rich Particulate Matter Modifies the Expression of Candidate MicroRNAs in Peripheral Blood Leukocytes  

PubMed Central

Background Altered patterns of gene expression mediate the effects of particulate matter (PM) on human health, but mechanisms through which PM modifies gene expression are largely undetermined. MicroRNAs (miRNAs) are highly conserved, noncoding small RNAs that regulate the expression of broad gene networks at the posttranscriptional level. Objectives We evaluated the effects of exposure to PM and PM metal components on candidate miRNAs (miR-222, miR-21, and miR-146a) related with oxidative stress and inflammatory processes in 63 workers at an electric-furnace steel plant. Methods We measured miR-222, miR-21, and miR-146a expression in blood leukocyte RNA on the first day of a workweek (baseline) and after 3 days of work (postexposure). Relative expression of miRNAs was measured by real-time polymerase chain reaction. We measured blood oxidative stress (8-hydroxyguanine) and estimated individual exposures to PM1 (< 1 ?m in aerodynamic diameter), PM10 (< 10 ?m in aerodynamic diameter), coarse PM (PM10 minus PM1), and PM metal components (chromium, lead, cadmium, arsenic, nickel, manganese) between the baseline and postexposure measurements. Results Expression of miR-222 and miR-21 (using the 2???CT method) was significantly increased in postexposure samples (miR-222: baseline = 0.68 ± 3.41, postexposure = 2.16 ± 2.25, p = 0.002; miR-21: baseline = 4.10 ± 3.04, postexposure = 4.66 ± 2.63, p = 0.05). In postexposure samples, miR-222 expression was positively correlated with lead exposure (? = 0.41, p = 0.02), whereas miR-21 expression was associated with blood 8-hydroxyguanine (? = 0.11, p = 0.03) but not with individual PM size fractions or metal components. Postexposure expression of miR-146a was not significantly different from baseline (baseline = 0.61 ± 2.42, postexposure = 1.90 ± 3.94, p = 0.19) but was negatively correlated with exposure to lead (? = ?0.51, p = 0.011) and cadmium (? = ?0.42, p = 0.04). Conclusions Changes in miRNA expression may represent a novel mechanism mediating responses to PM and its metal components.

Bollati, Valentina; Marinelli, Barbara; Apostoli, Pietro; Bonzini, Matteo; Nordio, Francesco; Hoxha, Mirjam; Pegoraro, Valeria; Motta, Valeria; Tarantini, Letizia; Cantone, Laura; Schwartz, Joel; Bertazzi, Pier Alberto; Baccarelli, Andrea



Effect of antiarrhythmic drugs on In-111-labeled leukocytes: chemotaxis and adherence to nylon wool  

SciTech Connect

The influence of lidocaine (L) and procainamide (P) on the chemotactic ability and adherence to nylon wool of In-111-labeled human polymorphonuclear leukocytes (PMNs) was investigated. At the normal therapeutic levels of L (0.022 mM whole blood) or P (0.03 mM whole blood) no change in PMN function was observed. However, at and above five times the aforementioned blood levels of L, significant reduction in the chemotactic ability of PMNs was noted (P <0.005). The adverse effects of In-111 radiation appeared insignificant at all L or P concentrations during the 3-hr observation period. The labeled PMNs were resistant to the toxic effects of a higher concentration of P than that of L, and the reduction in PMN chemotaxis and adherence to nylon wool was not apparent until the P concentration reached 1.5 mM.

Thakur, M.L.; Walsh, L.J.; Zaret, B.L.; Gottschalk, A.



Effect of antiarrhythmic drugs on In-111-labeled leukocytes: chemotaxis and adherence to nylon wool  

SciTech Connect

The influence of lidocaine (L) and procainamide (P) on the chemotactic ability and adherence to nylon wool of In-111-labeled human polymorphonuclear leukocytes (PMNs) was investigated. At the normal therapeutic levels of L (0.022 mM whole blood) or P (0.03 mM whole blood) no change in PMN function was observed. However, at and above five times the aforementioned blood levels of L, significant reduction in the chemotactic ability of PMNs was noted (p less than 0.005). The adverse effects of In-111 radiation appeared insignificant at all L or P concentrations during the 3-hr observation period. The labeled PMNs were resistant to the toxic effects of a higher concentration of P than that of L, and the reduction in PMN chemotaxis and adherence to nylon wool was not apparent until the P concentration reached 1.5 mM.

Thakur, M.L.; Walsh, L.J.; Zaret, B.L.; Gottschalk, A.



Interactions between Shiga toxins and human polymorphonuclear leukocytes  

Microsoft Academic Search

Human intestinal infections by Shiga toxin (Stx)-producing Escherichia coli cause hem- orrhagic colitis and hemolytic uremic syndrome (HUS), which represents the main cause of acute renal failure in early childhood. In HUS, Stx re- leased in the gut enter the bloodstream and are targeted to renal endothelium. The mechanism of toxin delivery is still a matter of debate, although the

Maurizio Brigotti; Domenica Carnicelli; Elisa Ravanelli; Stefania Barbieri; Francesca Ricci; Andrea Bontadini; Alberto E. Tozzi; Gaia Scavia; Alfredo Caprioli; Pier Luigi Tazzari



Intracellular localization of Rickettsia tsutsugamushi in polymorphonuclear leukocytes  

Microsoft Academic Search

Because rickettsiae proliferate exclusively in the cytoplasm of the host cells, rickett- sial invasion into the host cell cytoplasm is an essential step in rickettsial infections. Although this process is incompletely understood, two possible mechanisms that have been suggested are: the phagocytosis of the rickettsiae by the host cells followed by their escape from the phagosomes (1) or the direct

Y. Rikihisa; S. ITO



Damage of human polymorphonuclear leukocytes by Junin virus  

Microsoft Academic Search

One of the most constant manifestations of Argentine Haemorrhagic Fever (AHF) is a marked leukopenia, which is also observed in experimental infection of guinea pigs [6] and monkeys [9] with pathogenic strains of Junin virus (JV), the etiologic agent of AHF. The pathogenic mechanism leading to leukopenia is at present poorly understood. Although destruction of lymphatic tissue, characteristic of lethal

R. P. Laguens; P. H. Gonzalez; C. Ponzinibbio; J. Chambo



Immunomodulatory effects upon in vitro exposure of California sea lion and southern sea otter peripheral blood leukocytes to domoic acid.  


During red tide bloom events, the marine diatom Pseudo-nitzschia produces the toxin domoic acid (DA), which has been associated with stranding and mortality events involving California sea lions (Zalophus californianus) and southern sea otters (Enhydra lutris). In addition to these well-documented DA-induced neurotoxic events, there is increasing concern that DA may exert chronic effects, such as immunomodulation, which may potentially increase an individual's susceptibility to a number of opportunistic infections following nonlethal exposure. We investigated the effects of DA on innate (phagocytosis and respiratory burst) and adaptive (mitogen-induced lymphocyte proliferation) immune functions with the use of peripheral blood leukocytes collected from healthy California sea lions and southern sea otters upon in vitro exposure to 0 (unexposed control), 0.0001, 0.001, 0.01, 0.1, 1.0, 10, and 100 microM DA. Domoic acid did not significantly modulate phagocytosis or respiratory burst in either species. For California sea lions, DA significantly increased ConA-induced T-lymphocyte proliferation upon exposure to DA concentrations ranging from 0.0001 to 10 microM, resulting in a nonlinear dose-response curve. There was no effect on lymphocyte proliferation at the highest concentration of DA tested. No effects on lymphocyte proliferation were observed in southern sea otters. Importantly, the in vitro DA concentrations affecting T-cell proliferation were within or below the range of DA in serum measured in free-ranging California sea lions following natural exposure, suggesting a risk for immunomodulation in free-ranging animals. Understanding the risk for immunomodulation upon DA exposure will contribute in the health assessment and management of California sea lions and southern sea otters, as well as guide veterinarians and wildlife rehabilitators in caring for and treating afflicted animals. PMID:20688647

Levin, Milton; Joshi, Dhanashree; Draghi, Andrew; Gulland, Frances M; Jessup, David; De Guise, Sylvain



Immunological characterization of peripheral blood leukocytes using vaccine for mycoplasmal pneumonia of swine (MPS) in swine line selected for resistance to MPS.  


This study was conducted to evaluate immunological changes in peripheral blood leukocytes in pigs that were genetically selected for their improved resistance to mycoplasmal pneumonia of swine (MPS), using MPS vaccine as an antigen. Twelve castrated MPS-selected Landrace pigs were compared with the same number of pigs from a nonselected line by using a time-course analysis at the hematological level. After the second sensitization with MPS vaccine, the percentages of B cells, CD4(+) T cells, and natural killer (NK) cells in total leukocytes were lower in the selected line than in the nonselected line, whereas the percentage of granulocytes in total leukocytes increased in the MPS-selected line. We also assessed the proliferative ability of peripheral blood mononuclear cells (PBMCs) stimulated with Mycoplasma hyopneumoniae, lipopolysaccharide or concanavalin A, and found that although the proliferative ability of the PBMC was not different between the two lines at a steady state, the nonselected line showed a significantly higher proliferative ability after sensitization with MPS vaccine than the selected line regardless of antigens used. These results thus indicate that the selection of pigs on the basis of MPS resistance changes their immunophenotype, and would give us beneficial information for the prevention of MPS infection. PMID:23607374

Shimazu, Tomoyuki; Borjigin, Liushiqi; Katayama, Yuki; Li, Meihua; Satoh, Takumi; Watanabe, Kouichi; Kitazawa, Haruki; Roh, Sang-Gun; Aso, Hisashi; Katoh, Kazuo; Suda, Yoshihito; Sakuma, Akiko; Nakajo, Mituru; Suzuki, Keiichi



Temperature-induced transcription of inflammatory mediators and the influence of Hsp70 following LPS stimulation of southern bluefin tuna peripheral blood leukocytes and kidney homogenates.  


Temperature is known to influence inflammatory signalling in mammals, but far less understood in fish. The aim of the present study was to explore the potential effects of temperature on innate immune signalling in head kidney and leukocyte populations of the economically important southern bluefin tuna through the identification and utilization of gene expression targets in vitro. Here, we identified the mRNA sequences of five potential inflammatory mediators - TNF? (1 and 2), IL-1?, IL-8, and Cox2 - and demonstrate induction of four - TNF? (2), IL-1?, IL-8, and Cox2 - following LPS stimulation of both peripheral blood leukocytes and head kidney homogenates in vitro by real-time quantitative PCR. Comparison of transcriptional expression in cultures held at 18 and 25 °C (both within the presumed natural temperature range of this heterothermic species) showed accelerated transcription of cytokines TNF?, IL-1? and IL-8 following LPS stimulation at 25 °C in both tissue types. Peak induction reached comparable levels for each transcript at both temperatures during the 24 h test period with only limited (if any) protraction in expression resulting from cold temperature (18 °C) incubation. Partial mRNA sequences were also identified for both the constitutively expressed and heat inducible chaperone proteins Hsc70 and Hsp70, and 24 h incubation at 25 °C was sufficient to induce Hsp70 transcription in leukocyte but not in head kidney cell populations. Taken together these findings suggest that temperature exerts influence in the timing but not the degree of an innate inflammatory response in bluefin tuna and that different cell populations have differential responsiveness to heat shock in this heterothermic species. Further, LPS stimulation failed to induce Hsp70 at either incubation temperature in leukocytes; whereas 25 °C incubation caused Hsp70 up-regulation in leukocytes with or without the presence of LPS. This suggests that Hsp70 does not play a direct role in immune responsiveness for this species and that an environmental temperature of 25 °C in excess of 24 h initiates a cellular stress response in blood cells of this organism. Lastly, a strong correlation between Hsp70 and IL-8 transcriptional expression was observed following LPS/heat shock stimulation of leukocytes and five potential heat shock response elements were subsequently identified on the gene promoter region of IL-8 indicating that heat shock co-activation of this chemokine previously identified in mammals is also likely present in fish. PMID:23439399

Polinski, Mark; Bridle, Andrew; Nowak, Barbara



Endothelial Cell Activation by Leukocyte Microparticles1  

Microsoft Academic Search

The ability of polymorphonuclear leukocytes (PMNs) to modulate endothelial cell (EC) activation was investigated. Adding PMNs to cultured HUVECs resulted in a release of IL-6 (888 6 71 pg\\/ml, a 35-fold increase over release by the two cell types alone) and IL-8 (45.2 6 14.5 ng\\/ml, a 6.4-fold over PMN release alone and a 173-fold increase over EC release alone).

Mehdi Mesri; Dario C. Altieri



Levels of DNA damage in blood leukocyte samples from non-diabetic and diabetic female rats and their fetuses exposed to air or cigarette smoke.  


The objective of the present study was to evaluate DNA damage level in blood leukocytes from diabetic and non-diabetic female Wistar rats exposed to air or to cigarette smoke, and to correlate the findings with levels of DNA damage detected in blood leukocyte samples from their fetuses. A total of 20 rats were distributed into four experimental groups: non-diabetic (control; G1) and diabetic exposed to filtered air (G2); non-diabetic (G3) and diabetic (G4) exposed to cigarette smoke. Rats placed into whole-body exposure chambers were exposed for 30min to filtered air (control) or to tobacco smoke generated from 10 cigarettes, twice a day, for 2 months. Diabetes was induced by a pancreatic beta-cytotoxic agent, streptozotocin (40mg/kgb.w.). At day 21 of pregnancy, each rat was anesthetized and humanely killed to obtain maternal and fetal blood samples for genotoxicity analysis using the alkaline comet assay. G2, G3 and G4 dams presented higher DNA damage values in tail moment and tail length as compared to G1 group. There was a significant positive correlation between DNA damage levels in blood leukocyte samples from G2 and G3 groups (tail moment); G3 and G4 groups (tail length) and G3 group (tail intensity) and their fetuses. Thus, this study showed the association of severe diabetes and tobacco cigarette smoke exposure did not exacerbate levels of maternal and fetal DNA damages related with only diabetes or cigarette smoke exposure. Based on the results obtained and taking into account other published data, maternal diabetes requires rigid clinical control and public health and education campaigns should be increased to encourage individuals, especially pregnant women, to stop smoking. PMID:18455954

Lima, Paula Helena Ortiz; Damasceno, Débora Cristina; Sinzato, Yuri Karen; de Souza, Maricelma da Silva Soares; Salvadori, Daisy Maria Fávero; Calderon, Iracema de Mattos Paranhos; Rudge, Marilza Vieira Cunha




PubMed Central

Direct observations by phase microscopy have demonstrated that small numbers of pathogenic staphylococci survive prolonged periods of time within living human polymorphonuclear leukocytes. Non-pathogenic microorganisms are rapidly destroyed in similar preparations. Leukocytes in which staphylococci remained viable often appeared less vigorous after ingesting microorganisms, but intracellular survival could not be correlated with obvious leukocyte damage with any consistency. Both pathogenic and non-pathogenic cocci were seen to divide within living granulocytes during the first few minutes after ingestion. Occasionally pathogenic staphylococci multiplied in dying cells after long periods of intracellular residence. Phagocytosis of more than one pair of staphylococci by a single leukocyte appeared to act as a stimulus to bacterial destruction. Multiple ingestions of pathogenic staphylococci reduced the incidence of survival of the total microbiol population contained within the cell.

Melly, Marian Ann; Thomison, John B.; Rogers, David E.



Polymorphonuclear neutrophils function in splenectomized patients.  

PubMed Central

Some essential functions of polymorphonuclear neutrophils (PMN) were evaluated in 30 patients splenectomized because of rupture of the spleen. These cells revealed normal random migration, adherence, unstimulated O2- and H2O2 production. Phagocytosis of viable staphylococci was higher than in controls, whereas chemotaxis, bactericidal capacity, aggregation and stimulated O2- and H2O2 production were significantly impaired. PMN from splenectomized patients manifested also the decreased intracellular myeloperoxidase activity. The percentage of cells with receptor for Fc IgG in peripheral blood was markedly decreased. Plasma of these patients induced increased adherence of autologous as well as control neutrophils. The possible mechanisms leading to the observed events are discussed.

Wysocki, H; Wierusz-Wysocka, B; Karon, H; Dotka, J; Wykretowicz, A; Szczepanik, A; Klimas, R



Expression of tlr4, md2 and cd14 in equine blood leukocytes during endotoxin infusion and in intestinal tissues from healthy horses.  


The expression of tlr4, md2 and cd14 was studied in equine blood leukocytes and in intestinal samples using real time PCR. The stability of three commonly used reference genes, glyceraldehyde-3P-dehydrogenase (GAPDH), hypoxantine ribosyltransferase (HPRT) and succinate dehydrogenase complex subunit A (SDHA), was evaluated using qbase(PLUS). The equine peripheral blood mononuclear cells (eqPBMC) examined were either stimulated in vitro with Phorbol 12-myristate 13-acetate (PMA) and ionomycin or with the CpG oligodeoxynuclotide 2216 (CpG-ODN 2216) or obtained from horses before, during and after infusion of endotoxin. Intestinal tissue from healthy horses was sampled at ileum, right dorsal colon and rectum. Ranking of the three reference genes used for normalisation identified the combination HPRT/SDHA as most suitable both when determined ex vivo in leukocytes obtained from experimentally induced endotoxaemia and in eqPBMC activated in vitro while HPRT/GAPDH were most appropriate for the intestinal samples. The relative amounts of mRNA for TLR4 and MD-2 increased threefold during in vitro activation of the cells with CpG-ODN 2216 but was decreased in cultures stimulated with PMA/ionomycin. A transient elevation in the transcription of tlr4 and md2 was also evident for equine blood leukocytes following endotoxaemia. The levels of mRNA for CD14 on the other hand remained unaffected both during the induction of endotoxaemia and in the in vitro stimulated PBMCs. A low steady expression of TLR4, MD-2 and CD14 mRNA was demonstrated for the intestinal samples with no variation between the intestinal segments analysed. Thus, the foundation for real time PCR based levels of analysis of mRNA for all three components in the equine LPS receptor complex in different intestinal segments was set, making it possible to carry out future expression studies on clinical material. PMID:23036528

Fossum, C; Hjertner, B; Olofsson, K M; Lindberg, R; Ahooghalandari, P; Camargo, M M; Bröjer, J; Edner, A; Nostell, K



Evaluation of leukocyte stabilisation in TransFix®-treated blood samples by flow cytometry and transmission electron microscopy  

Microsoft Academic Search

In this report, we have evaluated the effects of a TransFix®-based stabilisation technique on leukocyte scatter characteristics, immunophenotyping, membrane permeability, absolute cell counting and morphology to extend previously reported flow cytometric data focused on the lymphocyte population. We show that scatter characteristics, immunophenotyping and absolute cell counting are well preserved, particularly in the lymphocyte population. Nevertheless, a general increase in

B. Canonico; L. Zamai; S. Burattini; V. Granger; F. Mannello; P. Gobbi; C. Felici; E. Falcieri; J. T. Reilly; D. Barnett; S. Papa



Morphometry of Human Leukocytes  

Microsoft Academic Search

In order to establish quantitative models of leukocyte functions. several morphometric parameters on individual white cells are needed. These include the diameter. volume. and membrane area of the cells and their nuclei in the undeformed state. A stereologic method was used to obtain these quantities from transmission electron microscopy of random sections through human white blood cells (neutrophils. lymphocytes. monocytes.

Geert W. Schmid-Sch; Yuan Y. Shih; Shu Chien



Hemodialysis-Induced Degranulation of Polymorphonuclear Cells: No Correlation between Membrane Markers and Degranulation Products  

Microsoft Academic Search

Background\\/Aims: Degranulation of polymorphonuclear leukocytes (PMN) during hemodialysis (HD) is usually assessed by measuring degranulation products. However, this process might also be estimated by the assessment of cell surface markers. In this study, the relationship between the expression of PMN degranulation markers (CD63 and CD66b) and the release of degranulation products [myeloperoxidase (MPO) and lactoferrin (LF)] was investigated during clinical

M. P. C. Grooteman; A. van Tellingen; A. J. van Houte; J. C. Bos; M. Schoorl; J. van Limbeek; M. J. Nubé



Experimental Closed Head Injury: Analysis of Neurological Outcome, Blood-Brain Barrier Dysfunction, Intracranial Neutrophil Infiltration, and Neuronal Cell Death in Mice Deficient in Genes for Pro-Inflammatory Cytokines  

Microsoft Academic Search

Cytokines are important mediators of intracranial inflammation following traumatic brain injury (TBI). In the present study, the neurological impairment and mortality, blood-brain barrier (BBB) function, intracranial polymorphonuclear leukocyte (PMN) accumulation, and posttraumatic neuronal cell death were monitored in mice lacking the genes for tumor necrosis factor (TNF)\\/lymphotoxin-? (LT-?) (TNF\\/LT-??\\/?) and interleukin-6 (IL-6) and in wild-type (WT) littermates subjected to experimental

Philip F. Stahel; Esther Shohami; Firas M. Younis; Karin Kariya; Viviane I. Otto; Philipp M. Lenzlinger; Maurice B. Grosjean; Hans-Pietro Eugster; Otmar Trentz; Thomas Kossmann; Maria C. Morganti-Kossmann



Whole blood fixation and permeabilization protocol with red blood cell lysis for flow cytometry of intracellular phosphorylated epitopes in leukocyte subpopulations  

Microsoft Academic Search

Background: Previous studies of intracellular expression of phospho-epitopes in human leukocytes using flow cyto- metry have used erythrocyte removal or lysis before fixation. Because many of the phospho-epitopes of interest are part of signaling networks that respond to the environment and turn over rapidly, the interval and manipulations used to eliminate erythrocytes from samples have the potential to introduce artifacts.

Sue Chow; David Hedley; Patricia Grom; Robert Magari; James W. Jacobberger; T. Vincent Shankey



Enhanced Chemotactic and Phagocytic Activities of Leukocytes in Psoriasis Vulgaris  

Microsoft Academic Search

Leukocytes derived from the peripheral blood of peripheral patients demonstrated an enhanced chemotactic response compared with leukocytes from healthy subjects. No significant difference was detected between the chemotactic response of leukocytes from patients with minimal or no skin involvement and those from patients with extensive lesions. Psoriatic leukocytes also had a significantly higher capacity to engulf 125I labeled Shigella flexneri

A. Wahba; H. A. Cohen; M. Bar-Eli; R. Gallily



Anandamide inhibits Theiler's virus induced VCAM-1 in brain endothelial cells and reduces leukocyte transmigration in a model of blood brain barrier by activation of CB1 receptors  

PubMed Central

Background VCAM-1 represents one of the most important adhesion molecule involved in the transmigration of blood leukocytes across the blood-brain barrier (BBB) that is an essential step in the pathogenesis of MS. Several evidences have suggested the potential therapeutic value of cannabinoids (CBs) in the treatment of MS and their experimental models. However, the effects of endocannabinoids on VCAM-1 regulation are poorly understood. In the present study we investigated the effects of anandamide (AEA) in the regulation of VCAM-1 expression induced by Theiler's virus (TMEV) infection of brain endothelial cells using in vitro and in vivo approaches. Methods i) in vitro: VCAM-1 was measured by ELISA in supernatants of brain endothelial cells infected with TMEV and subjected to AEA and/or cannabinoid receptors antagonist treatment. To evaluate the functional effect of VCAM-1 modulation we developed a blood brain barrier model based on a system of astrocytes and brain endothelial cells co-culture. ii) in vivo: CB1 receptor deficient mice (Cnr1-/-) infected with TMEV were treated with the AEA uptake inhibitor UCM-707 for three days. VCAM-1 expression and microglial reactivity were evaluated by immunohistochemistry. Results Anandamide-induced inhibition of VCAM-1 expression in brain endothelial cell cultures was mediated by activation of CB1 receptors. The study of leukocyte transmigration confirmed the functional relevance of VCAM-1 inhibition by AEA. In vivo approaches also showed that the inhibition of AEA uptake reduced the expression of brain VCAM-1 in response to TMEV infection. Although a decreased expression of VCAM-1 by UCM-707 was observed in both, wild type and CB1 receptor deficient mice (Cnr1-/-), the magnitude of VCAM-1 inhibition was significantly higher in the wild type mice. Interestingly, Cnr1-/- mice showed enhanced microglial reactivity and VCAM-1 expression following TMEV infection, indicating that the lack of CB1 receptor exacerbated neuroinflammation. Conclusions Our results suggest that CB1 receptor dependent VCAM-1 inhibition is a novel mechanism for AEA-reduced leukocyte transmigration and contribute to a better understanding of the mechanisms underlying the beneficial role of endocannabinoid system in the Theiler's virus model of MS.



N-Formylmethionyl Peptide Receptors on Equine Leukocytes Initiate Secretion but not Chemotaxis  

NASA Astrophysics Data System (ADS)

The chemotaxis of leukocytes appears to be initiated by the binding of chemotactic factors to the surface of these cells. N-Formylated peptides induce chemotaxis and lysosomal enzyme secretion of leukocytes; because these peptides are available in a purified radiolabeled form, they have been useful in the characterization of receptors for chemotactic factors. Equine polymorphonuclear leukocytes secrete lysosomal enzymes but do not exhibit chemotaxis in response to the N-formylated peptides, even though they have a high-affinity cell surface receptor for these agents. The specificity of the equine receptor resembles the specificity of the receptor on chemotactically responsive leukocytes from other species. Equine polymorphonuclear leukocytes may thus be an excellent model for the study of the events that lead to a biological response following receptor occupancy.

Snyderman, Ralph; Pike, Marilyn C.



Label-free in vivo imaging of human leukocytes using two-photon excited endogenous fluorescence.  


We demonstrate that two-photon excited endogenous fluorescence enables label-free morphological and functional imaging of various human blood cells. Specifically, we achieved distinctive morphological contrast to visualize morphology of important leukocytes, such as polymorphonuclear structure of granulocyte and mononuclear feature of agranulocyte, through the employment of the reduced nicotinamide adenine dinucleotide (NADH) fluorescence signals. In addition, NADH fluorescence images clearly reveal the morphological transformation process of neutrophils during disease-causing bacterial infection. Our findings also show that time-resolved NADH fluorescence can be potentially used for functional imaging of the phagocytosis of pathogens by leukocytes (neutrophils) in vivo. In particular, we found that free-to-bound NADH ratios measured in infected neutrophils increased significantly, which is consistent with a previous study that the energy consumed in the phagocytosis of neutrophils is mainly generated through the glycolysis pathway that leads to the accumulation of free NADH. Future work will focus on further developing and applying label-free imaging technology to investigate leukocyte-related diseases and disorders. PMID:23552632

Zeng, Yan; Yan, Bo; Sun, Qiqi; Teh, Seng Khoon; Zhang, Wei; Wen, Zilong; Qu, Jianan Y



Effect of infection with BHV-1 on peripheral blood leukocytes and lymphocyte subpopulations in calves with subclinical BVD.  


Bovine viral diarrhea virus (BVDV) and bovine herpesvirus-1 (BHV-1) are important cattle pathogens that induce a broad immunosuppression on cell-mediated immune response on its own participating in the bovine respiratory disease complex (BRDC). The aim of our study was to evaluate the quantitative changes in immunocompetent cells in healthy calves and calves with subclinical bovine viral diarrhea (BVD), both inoculated with BHV-1. Total leukocyte counts exhibited changes mainly in neutrophils and lymphocytes that can contribute to the BVDV immunosuppression, thus accounting for some of the intergroup differences. Monocytes did not display numerical changes in either group. Regarding lymphocyte subpopulations, even though CD4+ T lymphocytes and B cells were depleted around 4 dpi in both infected groups, the main difference observed between both groups was in CD8+ T cells which displayed an earlier depletion in BVDV inoculated calves that can promote a greater BHV-1 dissemination, thus aggravating the course of the disease. PMID:23541923

Molina, V; Risalde, M A; Sánchez-Cordón, P J; Pedrera, M; Romero-Palomo, F; Luzzago, C; Gómez-Villamandos, J C



Peripheral Blood Leukocyte Gene Expression Patterns and Metabolic Parameters in Habitually Snoring and Non-Snoring Children with Normal Polysomnographic Findings  

PubMed Central

Background: Children who snore but do not have gas exchange abnormalities or alterations of sleep architecture have primary snoring (PS). Since increasing evidence suggest that PS may be associated with morbidity, we hypothesized that assessing genome-wide gene expression in peripheral blood leukocytes (PBL) will identify a distinct signature in PS children. Methods: Children (aged 4–9 years) with and without habitual snoring and a normal PSG were designated as either PS or controls. Whole genome expression profiles of PBL and metabolic parameters in 30 children with PS and 30 age-, gender-, ethnicity-, and BMI-matched controls were compared. Pathway-focused gene network analysis of the PBL transcriptome was performed. Metabolic parameters were measured in an independent follow-up cohort of 98 children (64 PS and 34 controls) to evaluate the computationally derived findings. Results: PS was not associated with a distinct transcriptional signature in PBL. Exploratory functional network analysis of enriched gene sets identified a number of putative pathways—including those mapping to insulin signaling, adipocyte differentiation, and obesity—with significant alterations in glucose metabolism and insulin sensitivity emerging in the follow-up cohort of children with PS, but no differences in lipid profiles. Conclusions: PS children do not exhibit global perturbations in their PBL transcriptional response, suggesting that current normative PSG criteria are overall valid. However, subtle differences in functionally coherent pathways involved in glycemic homeostasis were detected and confirmed in a larger independent pediatric cohort indicating that PS may carry increased risk for end-organ morbidity in susceptible children. Citation: Khalyfa A; Gharib SA; Kim J; Capdevila OS; Kheirandish-Gozal L; Bhattacharjee R; Hegazi M; Gozal D. Peripheral blood leukocyte gene expression patterns and metabolic parameters in habitually snoring and non-snoring children with normal polysomnographic findings. SLEEP 2011;34(2):153-160.

Khalyfa, Abdelnaby; Gharib, Sina A.; Kim, Jinkwan; Capdevila, Oscar Sans; Kheirandish-Gozal, Leila; Bhattacharjee, Rakesh; Hegazi, Mohamed; Gozal, David



Biphasic control of polymorphonuclear cell migration by Kupffer cells. Effect of exposure to metabolic products of ethanol  

SciTech Connect

In order to investigate the role of the Kupffer cells in the regulation of the inflammatory reaction seen in alcoholic hepatitis, rat liver Kupffer cells were cultured and exposed to products of ethanol metabolism. The resultant supernatants were tested to study their ability to stimulate or inhibit polymorphonuclear cell chemotaxis. Kupffer cells produced increased chemokinetic activity for human polymorphonuclear leukocytes; when incubated with soluble products of microsomal peroxidation, the Kupffer cells engendered more chemokinetic activity than that produced by untreated Kupffer cells. When Kupffer cells were incubated with acetaldehyde, the chemokinetic activity that appeared in the supernatant did not differ from control. Chemotaxis of polymorphonuclear cells was not observed when the Kupffer cell supernatants were tested by checkerboard analysis.

Fainsilber, Z.; Feinman, L.; Shaw, S.; Lieber, C.S.



Inactivation of Viruses, Bacteria, Protozoa, and Leukocytes in Labile Blood Components by Using Nucleic Acid Targeted Methods  

Microsoft Academic Search

\\u000a Substantial increments in the safety of blood transfusion have been achieved through continued improvements in donor testing,\\u000a yet residual concern about the safety of blood components persists. To further reduce the risk of transfusion-associated infection,\\u000a additional measures, such as nucleic acid testing for selected pathogens, are being introduced. Transfusion of cellular components\\u000a has been implicated in transmission of viral, bacterial,

L. M. Corash


Comparative Study of the In Vitro Proliferative Responses of Blood and Synovial Fluid Leukocytes of Rheumatoid Arthritis Patients  

PubMed Central

Lymphocyte-rich suspensions from blood and synovial fluid (SF) of 20 patients with rheumatoid arthritis (RA) and from blood of 12 normal subjects, were cultured with heat-aggregated, aggregate-free, and native human gamma globulin (HGG), with autologous IgG separated from RA-SF by anion-exchange chromatography and with phytohemagglutinin (PHA). No significant differences were noted between the in vitro proliferative responses of blood lymphocytes of RA and normal controls to any of these preparations. Significant differences were noted between blood and SF lymphocytes of RA patients with respect to their responses to the aggregate-free HGG and to PHA. Incubation of RA-SF cells but not RA-blood cells with aggregate-free HGG before their culture with the aggregated HGG markedly suppressed the in vitro proliferative response to the latter. The observed differences between blood and SF lymphocytes and the suppression of blastogenic response of SF cells by exposure to the aggregate-free preparation raise the possibility of modulating the immune and/or the inflammatory responses in RA.

Reynolds, Michael D.; Abdou, Nabih I.



Disorders of sex development expose transcriptional autonomy of genetic sex and androgen-programmed hormonal sex in human blood leukocytes  

Microsoft Academic Search

BACKGROUND: Gender appears to be determined by independent programs controlled by the sex-chromosomes and by androgen-dependent programming during embryonic development. To enable experimental dissection of these components in the human, we performed genome-wide profiling of the transcriptomes of peripheral blood mononuclear cells (PBMC) in patients with rare defined \\

Paul-Martin Holterhus; Jan-Hendrik Bebermeier; Ralf Werner; Janos Demeter; Annette Richter-Unruh; Gunnar Cario; Mahesh Appari; Reiner Siebert; Felix Riepe; James D Brooks; Olaf Hiort



Leukocyte esterase  


... esterase is a urine test to look for white blood cells and other signs associated with infection. ... to detect a substance that suggests there are white blood cells in the urine, which may mean you have ...


Increased expression of TLR-2, COX-2, and SOD-2 genes in the peripheral blood leukocytes of opisthorchiasis patients induced by Opisthorchis viverrini antigen.  


Re-infection with liver fluke, Opisthorchis viverrini, increases proinflammatory molecules involved in inflammation-mediated disease and carcinogenesis in an animal model. To clarify whether these genes respond to parasite antigen in peripheral blood leukocytes (PBL) of opisthorchiasis patients, we examined the transcriptional level of oxidant-generating (toll-like receptor 2 (TLR-2), nuclear factor-kappa B (NF-KB), and cyclooxygenase 2 (COX-2)), anti-oxidant-generating (manganese superoxide dismutase 2 (SOD-2) and catalase (CAT)), proinflammatory cytokine (interleukin (IL)-1?), and anti-inflammatory cytokine (IL-10), in PBL exposed to parasite antigen in O. viverrini-infected patients compared with healthy individuals in an in vitro experiment. After O. viverrini antigen-treated PBL, quantitative RT-PCR analysis revealed that increased expression of cytokines and oxidant-generating genes in PBL was similar between O. viverrini-infected and healthy groups. Interestingly, compared with healthy subjects, increase of TLR-2, COX-2, and SOD-2 and decreased CAT mRNA expression levels were observed in O. viverrini-infected group. The results indicate that O. viverrini antigen induces upregulation of TLR-2, COX-2, and SOD-2 and downregulation of CAT genes in opisthorchiasis patients, suggesting that imbalance of oxidant/anti-oxidant transcripts during re-infection may be involved in the inflammatory-driven carcinogenesis. These molecules may be used as the chemopreventive target for intervention of opisthorchiasis patients in an endemic area. PMID:22160279

Yongvanit, Puangrat; Thanan, Raynoo; Pinlaor, Somchai; Sithithaworn, Paiboon; Loilome, Watcharin; Namwat, Nisana; Techasen, Anchalee; Dechakhamphu, Somkid



Whole blood transcriptional profiling reveals significant down-regulation of human leukocyte antigen class I and II genes in essential thrombocythemia, polycythemia vera and myelofibrosis.  


Abstract Gene expression profiling studies in the Philadelphia-negative chronic myeloproliferative neoplasms have revealed significant deregulation of several immune and inflammation genes that might be of importance for clonal evolution due to defective tumor immune surveillance. Other mechanisms might be down-regulation of major histocompatibility (MHC) class I and II genes, which are used by tumor cells to escape antitumor T-cell-mediated immune responses. We have performed whole blood transcriptional profiling of genes encoding human leukocyte antigen (HLA) class I and II molecules, ?2-microglobulin and members of the antigen processing machinery of HLA class I molecules (LMP2, LMP7, TAP1, TAP2 and tapasin). The findings of significant down-regulation of several of these genes may possibly be of major importance for defective tumor immune surveillance. Since up-regulation of HLA genes is recorded during treatment with epigenome modulating agents (DNA-hypomethylators and DNA-hyperacetylators [histone deacetylase inhibitors]) and interferon-?2, our findings call for prospective transcriptional studies of HLA genes during treatment with these agents. PMID:23302045

Skov, Vibe; Riley, Caroline Hasselbalch; Thomassen, Mads; Larsen, Thomas Stauffer; Jensen, Morten K; Bjerrum, Ole Weis; Kruse, Torben A; Hasselbalch, Hans Carl



Differential expression of leukocyte immunoglobulin-like receptors on cord-blood-derived human mast cell progenitors and mature mast cells.  


The leukocyte Ig-like receptors (LILRs) comprise a family of cell-surface immunoregulatory receptors with activating and inhibitory members. The inhibitory LILRs possess cytoplasmic ITIMs that down-regulate signaling by nonreceptor tyrosine kinase cascades. The activating members have a truncated cytoplasmic domain and signal through the FcR gamma chain. We examined the expression of LILRs on human mast cells during their development in vitro. Progenitor mast cells expressed cell surface inhibitory LILRB1, -B2, -B3, and -B4 and activating LILRA1. However, although mature cord blood-derived mast cells (hMCs) had detectable mRNA encoding multiple LILRs, none were expressed on the cell surface. Culture of progenitor mast cells or hMCs with various cytokine combinations failed to retain or induce cell surface expression of the LILRs. It is interesting that hMCs expressed LILRB5 in cytoplasmic granules and upon cross-linking of the high-affinity IgE receptor, released LILRB5 into the culture medium. Our results demonstrate that LILRs are developmentally regulated in human mast cells and that LILRB5 is expressed in mast cell granules and the release of soluble LILRB5 following IgE FcR-dependent stimulation, which has potential for amplification of mast cell-dependent, inflammatory responses. PMID:17998301

Tedla, Nicodemus; Lee, Chyh-Woei; Borges, Luis; Geczy, Carolyn L; Arm, Jonathan P



Effects of recombinant trout leptin in superoxide production and NF-?B/MAPK phosphorylation in blood leukocytes.  


Studies in mammals indicate that leptin is a multifunctional cytokine involved in regulation of energy metabolism and the modulation of the immune function. However, evidence for an immunomodulatory effect of leptin in fish is still missing. At least in part, this lack of knowledge is due to the absence of materials and models. In this study, we produced trout recombinant leptin (rt-lep) and tested its capacity to trigger cellular pathways, usually active in mammal immune system cells. STAT3, NF-?B, and the three major MAPK cascades (JNK, p38 and ERK), were activated by rt-lep in in vitro incubations with blood leucocytes of the rainbow trout Oncorhynchus mykiss. We also showed that rt-lep causes a decrease in superoxide anion production in trout blood leucocytes. Thus our data indicate that as in mammals also in teleosts leptin plays pleiotropic activities. Importantly, its actions in fishes do not always conform to the picture emerging for mammals. PMID:23932941

Mariano, Giovanna; Stilo, Romania; Terrazzano, Giuseppe; Coccia, Elena; Vito, Pasquale; Varricchio, Ettore; Paolucci, Marina




Microsoft Academic Search

In acute immunologic injury of tissues one of the common features noted is the accumulation of polymorphonuclear leukocytes (polymorphs) along with antigen, antibody, and host complement (C') at the site of damage. This has been noted in venules of the Arthus phenomenon, in arteries in serum sickness, and in glomeruli of acute nephritis both in experimental animals and in human



Technetium-99m-labeled white blood cells: a new method to define the local and systemic role of leukocytes in acute experimental pancreatitis.  

PubMed Central

OBJECTIVE: We developed a new method to quantitate leukocyte accumulation in tissues and used it to examine the time course and severity of acute experimental pancreatitis. BACKGROUND: Leukocyte activation and infiltration are believed to be critical steps in the progression from mild to severe pancreatitis and responsible for many of its systemic complications. METHODS: Pancreatitis of graded severity was induced in Sprague-Dawley rats with a combination of caerulein and controlled intraductal infusion. Technetium-99m (99mTc)-labeled leukocytes were quantified in pancreas, lung, liver, spleen, and kidney and compared with myeloperoxidase activity. The severity of pancreatitis was ascertained by wet/dry weight ratio, plasma amylase, and trypsinogen activation peptide in the pancreas. The time course of leukocyte accumulation was determined over 24 hours. RESULTS: Pancreatic leukocyte infiltration correlated well with tissue myeloperoxidase concentrations. In mild pancreatitis, leukocytes accumulated only in the pancreas. Moderate and severe pancreatitis were characterized by much greater leukocyte infiltration in the pancreas than in mild disease (p < 0.01), and increased 99mTc radioactivity was detectable in the lung as early as 3 hours. 99mTc radioactivity correlated directly with the three levels of pancreatitis. CONCLUSIONS: Mild pancreatitis is characterized by low-level leukocyte activation and accumulation in the pancreas without recruitment of other organs; marked leukocyte accumulation was found in the pancreas and in the lung in more severe grades of pancreatitis. These findings provide a basis for the pathophysiologic production of cytokines and oxygen free radicals, which potentiate organ injury in severe pancreatitis. This study validates a new tool to study local and systemic effects of leukocytes in pancreatitis as well as new therapeutic hypotheses.

Werner, J; Dragotakes, S C; Fernandez-del Castillo, C; Rivera, J A; Ou, J; Rattner, D W; Fischman, A J; Warshaw, A L



The effect of donor leukocyte infusion on refractory pure red blood cell aplasia after allogeneic stem cell transplantation in a patient with myelodysplastic syndrome developing from Kostmann syndrome.  


We describe the clinical course of a patient who experienced refractory pure red cell aplasia (PRCA) after undergoing HLA-matched allogeneic peripheral blood stem cell transplantation (allo-PBSCT) for refractory anemia with an excess of blasts in transformation that had evolved from Kostmann syndrome. The treatment for patients with myelodysplastic syndrome (MDS) or acute myeloid leukemia (AML) developing from Kostmann syndrome has not been standardized. We treated this patient with allo-PBSCT using a regimen combining high-dose cytosine arabinoside with granulocyte colony-stimulating factor, in addition to total body irradiation and cyclophosphamide without preceding intensive chemotherapy. The donor was ABO incompatible. Myeloid and platelet recoveries were achieved rapidly. Erythroid engraftment was not evident, however, and the patient was given a diagnosis of PRCA. Regimen-related toxicity and graft-versus-host disease (GVHD) were limited. The PRCA did not respond to various therapies, including the discontinuation of immunosuppressants for the induction of chronic GVHD, human recombinant erythropoietin, immunosuppressive treatment with steroids, cyclosporin A, and human anti-CD20 antibody (rituximab). The patient received transfusions 48 times until the resolution of his anemia by donor leukocyte infusion (DLI) at 25 months after PBSCT. He is now clinically well (performance status, 100%) with normal blood cell counts at 5 years after SCT. An in vitro study demonstrated that serum from the recipient blocked the differentiation of erythroid cells in the bone marrow. The results indicate that the conditioning regimen we describe seems safe and effective for those who have MDS/AML and that DLI might be a valuable approach for refractory PRCA after ABO-incompatible SCT. PMID:18192114

Ebihara, Yasuhiro; Manabe, Atsushi; Tsuruta, Toshihisa; Ishikawa, Kumiko; Hasegawa, Daisuke; Ohtsuka, Yoshitoshi; Kawasaki, Hirohide; Ogami, Kazuo; Wada, Yuka; Kanda, Tadayasu; Tsuji, Kohichiro



Selective Upregulation of microRNA Expression in Peripheral Blood Leukocytes in IL-10?/? Mice Precedes Expression in the Colon1  

PubMed Central

IL-10?/? mice, an animal model of Th1-mediated inflammatory bowel disease, were screened for the expression of 600 microRNAs (miRNAs) using colonic tissues and peripheral blood leukocytes (PBLs) from animals having either mild inflammation or severe intestinal inflammation. The development of colonic inflammation in IL-10?/? mice was accompanied by upregulation in the expression of ten miRNAs (miR-19a, miR-21, miR-31, miR-101, miR-223, miR-326, miR-142-3p, miR-142-5p, miR-146a, and miR-155). Notably, the expression of all of these miRNAs plus miR-375 was elevated in PBLs of IL-10?/? mice at a time when colonic inflammation was minimal, suggesting that changes in specific miRNAs in circulating leukocytes may be harbingers of ensuing colonic pathology. In vitro exposure of colonic intraepithelial lymphocytes to IL-10 resulted in down-regulation of miR-19a, miR-21, miR-31, miR-101, miR-223, and miR-155. Interestingly, unlike IL-10?/? mice, changes in miRNAs in PBL of dextran sulfate sodium-treated mice were minimal, but were selectively elevated in the colon after pathology was severe. We further show that miR-223 is a negative regulator of the Roquin ubiquitin ligase, that Roquin curtails IL-17A synthesis, and that the 3? UTR of Roquin is a target for miR-223, thus defining a molecular pathway by which IL-10 modulates IL-17-mediated inflammation. To identify additional miRNAs that may be involved in the regulation of Roquin, transcriptome analysis was done using cDNAs from HeLa cells transfected with 90 miRNA mimics. Twenty-six miRNAs were identified as potential negative regulators of Roquin, thus demonstrating functional complexity in gene expression regulation by miRNAs.

Schaefer, Jeremy S.; Montufar-Solis, Dina; Vigneswaran, Nadarajah; Klein, John R.




Microsoft Academic Search

The recruitment of leukocytes from the blood stream to extravascular tissue is a critical event in host defense against microbial invasion and in the repair of tissue damage. Studies by intravital microscopy have established a sequence of events involved in phagocyte emigration at sites of inflammation. In response to extravascular stimuli such as bacterial-derived chemoattractants or endogenous lipid and peptide

John M. Harlan


Cytotoxic Activity of Peripheral Blood Mononuclear Leukocytes, Activated by Interleukin-2/?-Cyclodextrin Nanocomposition against Androgen Receptor-Negative Prostate Cancers.  


Nanocomposition comprised of interleukin-2 in suboptimal noneffective concentration and ?-cyclodextrin was studied in vitro. This preparation as well as interleukin-2 in optimal concentration was shown to increase natural killer activity to K-562 cells and cytotoxicity of activated peripheral blood mononuclear cells (PBMCs) against PC-3 and DU 145 cells. At the same time ?-cyclodextrin or interleukin-2 in equimolar concentrations did not influence the spontaneous killer activity of PBMC. This combination of cyclodextrin + interleukin-2 led to the decrease of interleukin-2 effective concentration by an order. This phenomenon could be explained by cyclodextrins ability to promote the formation of nanoparticles with drugs, which results in enhancing their water solubility and bioavailability. Besides, interleukine-2/?-cyclodextrin nanocomposition as opposed to interleukin-2 alone led to increasing the number of not only lymphocytes, but also macrophages contained in activated PBMC population. Application of low concentration of interleukin-2 allowing for good clinical efficiency may significantly mitigate the side effects of the drug and enable to develop adoption of immunotherapy for patients with androgen-resistant prostate cancer. PMID:22084730

Anisimova, Natalia Yu; Sosnov, Andrey V; Ustyuzhanina, Nadezhda E; Baronzio, Gianfranco; Kiselevsky, Mikhail V



Disorders of sex development expose transcriptional autonomy of genetic sex and androgen-programmed hormonal sex in human blood leukocytes  

PubMed Central

Background Gender appears to be determined by independent programs controlled by the sex-chromosomes and by androgen-dependent programming during embryonic development. To enable experimental dissection of these components in the human, we performed genome-wide profiling of the transcriptomes of peripheral blood mononuclear cells (PBMC) in patients with rare defined "disorders of sex development" (DSD, e.g., 46, XY-females due to defective androgen biosynthesis) compared to normal 46, XY-males and 46, XX-females. Results A discrete set of transcripts was directly correlated with XY or XX genotypes in all individuals independent of male or female phenotype of the external genitalia. However, a significantly larger gene set in the PBMC only reflected the degree of external genital masculinization independent of the sex chromosomes and independent of concurrent post-natal sex steroid hormone levels. Consequently, the architecture of the transcriptional PBMC-"sexes" was either male, female or even "intersex" with a discordant alignment of the DSD individuals' genetic and hormonal sex signatures. Conclusion A significant fraction of gene expression differences between males and females in the human appears to have its roots in early embryogenesis and is not only caused by sex chromosomes but also by long-term sex-specific hormonal programming due to presence or absence of androgen during the time of external genital masculinization. Genetic sex and the androgen milieu during embryonic development might therefore independently modulate functional traits, phenotype and diseases associated with male or female gender as well as with DSD conditions.

Holterhus, Paul-Martin; Bebermeier, Jan-Hendrik; Werner, Ralf; Demeter, Janos; Richter-Unruh, Annette; Cario, Gunnar; Appari, Mahesh; Siebert, Reiner; Riepe, Felix; Brooks, James D; Hiort, Olaf



Polymorphonuclear leucocyte function in Behçet's disease  

Microsoft Academic Search

Polymorphonuclear leucocyte function was investigated in 19 patients with active Behçet's disease. Spontaneous free leucocyte migration was found to be significantly reduced, yet after stimulation the leucocyte's chemotactic activity was considerably increase (p less than 0-05) when compared to control leucocytes. Control leucocytes migrated more rapidly when incubated in serum taken from patients with Behçet's disease (p less than 0-005).

J D Sobel; S Haim; N Obedeanu; T Meshulam; D Merzbach



Cytoplasmatic Alkaline Phosphatase and Allergic Alteration of Blood Leukocytes after Sensitization during Sonne Keratoconjunctivitis in Animals and during Dysentery in Humans.  

National Technical Information Service (NTIS)

Working on the basis of the pathogenetic significance of bacterial sensitization during dysenteric infection, the authors investigated allergic alterations of leukocytes. They determined the nature of changes and distribution of the activity of cytoplasma...

D. M. Nedopryadko G. I. Fredman




PubMed Central

Lactoferrin, an iron-binding protein previously shown to occur in many external secretions, is identified as one of the major proteins present in human and guinea pig neutrophilic polymorphonuclear leukocytes. The identification of this protein in leukocyte extracts was based upon a comparison of its electrophoretic, antigenic, and iron-combining properties with the corresponding properties of the same protein isolated from human and guinea pig milk. Immunochemical quantitations showed that lactoferrin occurs in human neutrophilic leukocytes at the concentration of 3 µg per 106 cells. Tissue cultures from guinea pig bone marrow and spleen actively synthesized the protein, as shown both by net production of lactoferrin and incorporation of labeled amino acids into the protein. Immunohistochemical data indicate that lactoferrin first appears in myeloid cells at the stage of the promyelocyte.

Masson, P. L.; Heremans, J. F.; Schonne, E.



Efficiency and safety of leukocyte filtration during cardiopulmonary bypass for cardiac surgery  

Microsoft Academic Search

Background. Leukocyte filtration of systemic blood during cardiopulmonary bypass surgery to reduce post-operative morbidity has not yet been established because of the enormous leukocyte release from the third space. This study was designed to examine the efficiency and safety of leukocyte filtration by a new prototype large capacity leukocyte filter.Patients and methods. Patients undergoing cardiopulmonary bypass surgery were prospectively divided

J. J. J Smit; A. J de Vries; Y. J Gu; W van Oeveren



Two-Step Leukocyte Migration in Agarose Technique  

Microsoft Academic Search

A two-step modification of Clausen’s technique of leukocyte migration under agarose is described. Blood leukocytes from a subject are incubated for approximately 4 days with or without antigen. The cultures, after reconstitution with antigen followed by repeated washing of the cultured cells, are added to fresh, dextran-separated blood leukocytes, and migration tests are performed in the absence of the antigen.

Gun Agrup; Bengt Källén; Olle Nilsson



Substitution of Aspartate for glycine 1018 in the Type III procollagen (COL3AI) gene causes type IV Ehlers-Danlos Syndrome: The mutated allele is present in most blood leukocytes of the asymptomatic and mosaic mother  

SciTech Connect

A proband with arterial ruptures and skin changes characteristic of the type IV variant of Ehlers-Danlos syndrome was found to have a single-base mutation in the type III procollagen gene, which converted the codon for glycine at amino position 1018 to a codon for aspartate. (Amino acid positions are numbered by the standard convention in which the first glycine of the triple-helical domain of an [alpha] chain is number 1. The numbers of positions in the [alpha]1(III) chains can be converted to positions in the human pro[alpha](III) chain by adding 167.). Nucleotide sequencing of overlapping PCR products in which the two alleles were distinguished demonstrated that the mutation of glycine 1018 was the only mutation that changed the primary structure of type III procollagen. The glycine substitution markedly decreased the amount of type III procollagen secreted into the medium by cultured skin fibroblasts from the proband. It is surprising that the same mutation was found in about 94% of the peripheral blood leukocytes from the proband's asymptomatic 72-year-old mother. Other tissues from the mother contained the mutated allele; it was present in 0%-100% of different samples of hair cells and in about 40% of cells from the oral epithelium. Therefore, the mother was a mosaic for the mutation. Since the mutated allele was present in cells derived from all three germ layers, the results indicated that the mutation arose by the late blastocyst stage of development. The results also indicate that assays of blood leukocytes do not always reveal mosaicism or predict phenotypic involvement of tissues, such as blood vessels, that are derived from the same embryonic cells as are leukocytes. 66 refs., 6 figs., 1 tab.

Kontusaari, S.; Tromp, G.; Kuivaniemi, H.; Prockop, D.J. (Thomas Jefferson Univ., Philadelphia, PA (United States)); Stolle, C. (Robert Wood Johnson Medical School, Piscataway, NJ (United States)); Pope, F.M.



In utero infection with porcine reproductive and respiratory syndrome virus modulates leukocyte subpopulations in peripheral blood and bronchoalveolar fluid of surviving piglets  

Microsoft Academic Search

It is well known that piglets congenitally infected with porcine reproductive and respiratory syndrome virus (PRRSV) can be viremic at birth, and that preweaning mortality due to secondary infections often increases during acute outbreaks of PRRS. Therefore, an immunosuppressive effect of in utero infection has been suggested. The aim of the present study was to characterise the changes of leukocyte

J Nielsen; A Bøtner; J.-E Tingstedt; B Aasted; C. K Johnsen; U Riber; P Lind



Phagocytosis of neutrophil polymorphonuclears by macrophages in human bone marrow: importance in granulopoiesis.  

PubMed Central

The cytological and electron-microscopic appearance of aneutrophil phagocytosis by macrophages in normal human bone marrow is described. This feature can be observed in every normal bone marrow and is especially frequent in autoimmune disease. Bone marrrow phagocytosis of polymorphonuclear neutrophils seems to be a physiological process resulting from the random egress of neutrophils from bone marrow to blood. Images Fig 1, 2, and 3 Fig. 4

Dresch, C; Flandrin, G; Breton-Gorius, J



Correlation between reproductive status and steady-state messenger ribonucleic acid levels of the Myxovirus resistance gene, MX2, in peripheral blood leukocytes of dairy heifers.  


The objectives of the current study were to evaluate the correlation between reproductive status and steady-state levels of Myxovirus resistance gene (MX2) mRNA in peripheral blood leukocytes (PBL) of dairy heifers and the reliability of using change in MX2 messenger RNA (mRNA) for identification of nonpregnant heifers 18 to 19 d after AI. Holstein heifers (n = 266), 13 +/- 1 mo of age, were assigned randomly to be inseminated (BRED; n = 214) or not (NONBRED; n = 52). Estrous cycles of all heifers were synchronized with an intravaginal insert containing progesterone for 7 d. At insert removal, heifers received an injection of PGF2alpha. Heifers in the BRED group were inseminated on detection of estrus or at a fixed time, 72 h after insert removal concomitant with a GnRH treatment. Heifers in the NONBRED group received an injection of GnRH 48 h after insert removal. Blood samples collected on d 0 (d of AI or estrus) and 18 were used to determine steady-state levels of MX2 mRNA. Samples collected on d 0, 7, 14, and 21 were analyzed for progesterone concentration. Pregnancy was determined retrospectively by progesterone concentration on d 21 and was diagnosed at 30 +/- 1 and 60 +/- 3 d after AI. The fold change in levels of MX2 mRNA from d 0 to 18 was greater for heifers classified and diagnosed as pregnant on d 21 (P < 0.05) and 30 +/- 1 (P < 0.05) and 60 +/- 3 (P < 0.05) d after AI compared with nonpregnant (bred but not pregnant) and NONBRED heifers. Heifers that experienced pregnancy loss from 21 to 30 +/- 1 (P = 0.11) or 21 to 60 +/- 3 (P = 0.08) d of gestation tended to have smaller fold increases in MX2 mRNA expression than those that maintained pregnancy. The sensitivity (range 57.1 to 65.6%) and negative predictive values (range 47.9 to 57.1%) of determining reproductive status on d 18 according to the change in the level of MX2 mRNA expression in PBL were low, and the correlation between diagnosis of pregnancy by fold change in MX2 mRNA expression and other methods was small (r = 0.20 to 0.36). The current study indicates that increased expression of MX2 mRNA in PBL is related to pregnancy approximately 21, 30, and 60 d after AI in dairy heifers and that losses that occurred later in pregnancy were associated with lower fold increases in MX2 mRNA. However, using the change in MX2 mRNA expression was not a reliable method for diagnosis of pregnancy at 18 d after AI because of the low sensitivity and negative predictive value. PMID:17431047

Stevenson, J L; Dalton, J C; Ott, T L; Racicot, K E; Chebel, R C



Surface modification of polymeric materials and its effect on blood compatibility  

SciTech Connect

The surfaces of commercially available polymeric materials have been modified through the chemical infusion process and physical vapor deposition. The surfaces of poly(methylmethacrylate) (PMMA) have been modified through a chemical infusion process by treatment of the sample with a solution containing varying amounts of titanium(IV)isopropoxide and polyvinylpyrrolidone (PVP). The surfaces of silicone rubber samples have been coated with a thin coating of titanium dioxide with an ion beam sputtering technique. The treated samples were characterized by scanning electron microscopy, optical microscopy, and neutron activation analysis. The infused samples were evaluated for blood compatibility using two biological assays: an adherence assay in which the adherence of human polymorphonuclear leukocytes to the samples was determined, and a hemolysis assay using rat blood erythrocytes to determine the hemolytic activity of the samples. Based on the results of these assays, the PMMA samples treated with PVP alone resulted in an improvement in reactivity with the blood cells. 16 refs., 4 figs.

Wrobleski, D.A.; Cash, D.L.; Archuleta, T.; Barthell, B.L.; Kossowsky, R.; London, J.E.; Lehnert, B.E.; Duchane, D.V.



Junctional adhesion molecule-A-deficient polymorphonuclear cells show reduced diapedesis in peritonitis and heart ischemia-reperfusion injury  

PubMed Central

Junctional Adhesion Molecule-A (JAM-A) is a transmembrane adhesive protein expressed at endothelial junctions and in leukocytes. Here we report that JAM-A is required for the correct infiltration of polymorphonuclear leukocytes (PMN) into an inflamed peritoneum or in the heart upon ischemia-reperfusion injury. The defect was not observed in mice with an endothelium-restricted deficiency of the protein but was still detectable in mice transplanted with bone marrow from JAM-A-/- donors. Microscopic examination of mesenteric and heart microvasculature of JAM-A-/- mice showed high numbers of PMN adherent on the endothelium or entrapped between endothelial cells and the basement membrane. In vitro, in the absence of JAM-A, PMN adhered more efficiently to endothelial cells and basement membrane proteins, and their polarized movement was strongly reduced. This paper describes a nonredundant role of JAM-A in controlling PMN diapedesis through the vessel wall.

Corada, Monica; Chimenti, Stefano; Cera, Maria Rosaria; Vinci, Maria; Salio, Monica; Fiordaliso, Fabio; De Angelis, Noeleen; Villa, Antonello; Bossi, Mario; Staszewsky, Lidia I.; Vecchi, Annunciata; Parazzoli, Dario; Motoike, Toshiyuki; Latini, Roberto; Dejana, Elisabetta



Leukocyte adhesion deficiency II syndrome, a generalized defect in fucose metabolism  

Microsoft Academic Search

Leukocyte adhesion deficiency II has been described in only 2 patients; herein we report extensive investigation of another patient. The physical stigmata were detected during prenatal ultrasonographic investigation. Sialyl-Lewis X (sLex) was absent from the surface of polymorphonuclear neutrophils, and cell binding to E- and P-selectin was severely impaired, causing an immunodeficiency. The elevation of peripheral neutrophil counts occurred within

Thorsten Marquardt; Thomas Brune; Kerstin Lühn; Klaus-Peter Zimmer; Christian Körner; Larissa Fabritz; Natascha van der Werft; Josef Vormoor; Hudson H. Freeze; Frank Louwen; Bettina Biermann; Eric Harms; Kurt von Figura; Dietmar Vestweber; Hans Georg Koch



Proteoglycans synthesized by human polymorphonuclear leucocytes in vitro.  


Polymorphonuclear leucocytes (PMN) were assessed in vitro for their ability to synthesize and secrete proteoglycans. The PMN were isolated from human peripheral blood and were found to contain less than 5% mononuclear cells. Following 24 h incubation in the presence of (35S)-sulfate, significant quantities of 35S-labelled macromolecules were detected both within the culture medium and cells. Although the PMN preparations contained some platelets (approximately five platelets:one PMN), culture of platelets alone did not result in the detection of any 35S-labelled macromolecules in either the medium or platelets. 35S/3H-labelled macromolecules from the PMN cultures were identified as proteoglycans on the basis of their degradation by papain, alkaline sodium borohydride, chondroitinase ACII, chondroitinase ABC and nitrous acid. The labelled proteoglycans isolated from the medium and cells eluted from Sepharose CL-4B with a Kav of 0.63; this indicated a small size compared with many other proteoglycans. The glycosaminoglycans associated with the proteoglycans were identified as heparan sulfate, chondroitin sulfate and dermatan sulfate, with chondroitin sulfate being the principal component. The average molecular weight of the glycosaminoglycans was determined to be 16,000. Therefore, the data from this study demonstrate the ability of human PMN to synthesize and secrete proteoglycans in vitro which appear to differ from those synthesized by mesenchymal cells with respect to molecular size and glycosaminoglycan composition. PMID:2498202

Bartold, P M; Harkin, D G; Bignold, L P



A daunting task: manipulating leukocyte function with RNAi.  


RNA interference (RNAi) has advanced into clinical trials. In spite of the progress made in systemic RNAi delivery to the liver and solid tumors, delivery of RNAi to leukocytes remains challenging and less advanced. Manipulating leukocyte function with RNAi holds great promise for streamlining the drug discovery process by facilitating in vivo drug target validation and for facilitating the development of RNAi-based therapy platforms for leukocyte-implicated diseases, such as blood cancer, inflammation, and leukocyte-tropic viral infections. In this review, progress in delivery strategies of RNAi payloads to leukocytes, which are notoriously difficult cells to transduce with RNAi, is discussed with special emphasis on the challenges and potential opportunities for manipulating leukocyte function with RNAi. PMID:23550647

Peer, Dan




Technology Transfer Automated Retrieval System (TEKTRAN)

Bacterial invasion and growth within the cistern of the mammary gland is the main cause of bovine mastitis. Mastitis can affect essentially all lactating mammals, but is especially problematic for dairy cattle. Intramammary infections of dairy cows with Gram-positive bacteria such as Staphylococcu...


Serum opsonin, bacteria, and polymorphonuclear leukocyte interactions in subacute bacterial endocarditis  

PubMed Central

The effect of anti-?-globulin factors on 7S ?-globulin opsonins from patients with subacute bacterial endocarditis has been examined with a quantitative in vitro phagocytosis system. Human anti-?-globulin factors from patients with subacute bacterial endocarditis and rheumatoid arthritis inhibited the opsonic action of 7S ?-globulin specifically bound to bacteria. A similar antiopsonic effect was obtained with rabbit antiserum to human ?G globulin. The antiopsonic effect of anti-?-globulin factors did not correlate with their ability to potentiate agglutination of bacteria by 7S antibody. Competition was demonstrated between the antiopsonic effect of anti-?-globulin factors and the phagocytosis-promoting action of heat-labile serum factors containing hemolytically active complement.

Messner, Ronald P.; Laxdal, Throstur; Quie, Paul G.; Williams, Ralph C.



[Effect of erythromycin and phosphomycin on the ingestion and destruction capacity of the human polymorphonuclear leukocyte].  


The aim of the present study was to evaluate the uptake and intracellular destruction capacity of the human neutrophil. Human neutrophils from healthy individuals not receiving any therapy were incubated during 15, 90 and 180 minutes with the minimal, intermediate and maximal doses achieved in serum during treatment with therapeutic doses of these antibiotics (3, 6.45 and 9.9 micrograms/ml of erythromycin and 30, 45 and 60 micrograms/ml of phosphomycin). After this incubation, the uptake capacity and the intracellular destruction capacity of Candida guillermondii by neutrophils was assessed. Both erythromycin and phosphomycin maintained the baseline levels of both capacities in all the incubations evaluated. In addition, during the incubation with maximal dose erythromycin (15 min) a stimulatory tendency of the uptake and intracellular destruction capacity was observed. PMID:2490465

Amurrio, C; Nicolás, R; Larrauri, L; López, A; Cisterna, R


Complement Activation by Myeloperoxidase Products Released from Stimulated Human Polymorphonuclear Leukocytes  

Microsoft Academic Search

Purified human myeloperoxidase (MPO) converted human C5 to an activated form, i. e. the C5 protein adopted a configuration expressing a binding site for C6; the resulting C56 complex then reacted with C7, C8 and C9 forming a hemolytic C5-9 complex. For the activation by myeloperoxidase chloride and hydrogen peroxide were essential. This indicates that the peroxidase acted through the

Walther Vogt



Release of lactoferrin by polymorphonuclear leukocytes after aerosol challenge with Escherichia coli.  

PubMed Central

Mice with cyclophosphamide-induced granulocytopenia were challenged with aerosolized Escherichia coli, their lungs were lavaged at 1 and 4 h, and total cell counts, differential counts, and levels of lactoferrin, transferrin, and albumin were measured in the lung lavage fluid. Lung lavage fluid from cyclophosphamide-treated mice had few neutrophils and no increase in lactoferrin levels, whereas control mice had significant increases in both. Transferrin levels did not change in either group. Neutrophils are the source of increased lactoferrin levels in lung lavage fluid after aerosol challenge.

LaForce, F M; Boose, D S



Release of lactoferrin by polymorphonuclear leukocytes after aerosol challenge with Escherichia coli.  


Mice with cyclophosphamide-induced granulocytopenia were challenged with aerosolized Escherichia coli, their lungs were lavaged at 1 and 4 h, and total cell counts, differential counts, and levels of lactoferrin, transferrin, and albumin were measured in the lung lavage fluid. Lung lavage fluid from cyclophosphamide-treated mice had few neutrophils and no increase in lactoferrin levels, whereas control mice had significant increases in both. Transferrin levels did not change in either group. Neutrophils are the source of increased lactoferrin levels in lung lavage fluid after aerosol challenge. PMID:3305371

LaForce, F M; Boose, D S



Pharmacological control of neutrophil-mediated inflammation: Strategies targeting calcium handling by activated polymorphonuclear leukocytes  

PubMed Central

Unlike most other effector cells of the innate, as well as the adaptive immune systems, the neutrophil is a relatively undiscerning aggressor with scant regard for damage limitation. Although this highly combative, professional phagocyte has become increasingly implicated in the immunopathogenesis of many acute and chronic inflammatory disorders, of both infective and noninfective origin, effective pharmacological strategies to counter neutrophilaggression have remained elusive. Activation of neutrophils results in rapid mobilization of both stored and extracellular Ca2+, resulting in abrupt, usually transient increases in cytosolic Ca2+, which precede, and are a prerequisite for activation of the Ca2+-dependent pro-inflammatory activities of these cells. Mobilization of Ca2+ by, and restoration of Ca2+ homeostasis to activated neutrophils are multistep processes which present a number of potential targets, some well recognized and others noveland unconventional, for the pharmacological control of neutrophil-mediated inflammation. Uncovering these targets represents the primary focus of this review.

Tintinger, Gregory R; Steel, Helen C; Theron, Annette J; Anderson, Ronald



Ultrastructural alterations during ATP-induced secretion of lysosomal enzymes from rabbit polymorphonuclear leukocytes  

Microsoft Academic Search

Rabbit neutrophils incubated in low-ionic-strength media were stimulated by ATP to secrete lysosomal enzymes. This was greatly enhanced in the presence of cytochalasin B. ATP in these circumstances induced the cell to form large cytoplasmic extensions that were largely devoid of granules. In the presence of both ATP and cytochalasin B, however, the projections contained granules in close proximity to

P. M. Henson; J. E. Henson; E. L. Becker



Phospholipase C Activity in Human Polymorphonuclear Leukocytes: Partial Characterization and Effect of Indomethacin.  

National Technical Information Service (NTIS)

Several hormones act at the cellular level to increase diacylglycerol via increased catabolism of phosphatidylinositol by phospholipase C. Diacylglycerol stimulates protein kinase C, leading to protein phosphorylation adn hormone action. Since phospholipa...

K. M. Shakir C. O. Simpkins S. L. Gartner D. O. Sobel T. J. Williams



Phospholipase C Activity in Human Polymorphonuclear Leukocytes: Partial Characterization and Effect of Indomethacin.  

National Technical Information Service (NTIS)

Several hormones act at the cellular level to increase diacylglycerol via increased catabolism of phosphatidylinositol by phospholipase C (PLC). Diacyglycerol stimulates protein kinase C, leading to protein phosphorylation and hormone action. Since PLC ac...

C. O. Simpkins D. O. Sobel K. M. Shakir S. L. Gartner T. J. Williams



Fine structure and cytochemical study of the interaction between Fonsecaea pedrosoi and rat polymorphonuclear leukocyte.  


The induction of a granulomatous reaction is frequently observed in subcutaneous mycoses. Our previous studies demonstrated that Fonsecaea pedrosoi was able to survive and proliferate in tissue macrophages and that activated macrophages were fungistatic but not fungicidal. By contrast, our present studies revealed that neutrophils were able to kill F. pedrosoi cells in periods shorter than 20 min. Several phases of the interaction process were analysed by light and electron microscopy. The kinetic analysis demonstrated no significant difference during the first hour of F. pedrosoi-neutrophil interaction. Electron microscopy images showed that neutrophils readily associated with and killed extracellular fungi; however, few fungi were ingested. During this process the activation of respiratory burst took place as evaluated by light and electron microscopy. Cytochemical activity of acid and alkaline phosphatase was detected in low levels during the host cell parasite interaction. PMID:8912165

Rozental, S; Alviano, C S; de Souza, W


Micron-scale positioning of features influences the rate of polymorphonuclear leukocyte migration.  

PubMed Central

Microfabrication technology was used to create regular arrays of micron-size holes (2 microm x 2 microm x 210 nm) on fused quartz and photosensitive polyimide surfaces. The patterned surfaces, which possessed a basic structural element of a three-dimensional (3-D) network (i.e., spatially separated mechanical edges), were used as a model system for studying the effect of substrate microgeometry on neutrophil migration. The edge-to-edge spacing between features was systematically varied from 6 microm to 14 microm with an increment of 2 microm. In addition, collagen was used to coat the patterned quartz surfaces in an attempt to change the adhesive properties of the surfaces. A radial flow detachment assay revealed that cell adhesion was the strongest on the quartz surface (approximately 50% cell attached), whereas it was relatively weaker on polyimide and collagen-coated quartz (approximately 25% cell attached). Cell adhesion to each substrate was not affected either by the presence of holes or by the spacing between holes. A direct visualization assay showed that neutrophil migration on each patterned surface could be characterized as a persistent random walk; the dependence of the random motility coefficient (mu) as a function of spacing was biphasic with the optimal spacing at approximately 10 microm on each substrate. The presence of evenly distributed holes at the optimal spacing of 10 microm enhanced mu by a factor of 2 on polyimide, a factor of 2.5 on collagen-coated quartz, and a factor of 10 on uncoated quartz. The biphasic dependence on the mechanical edges of neutrophil migration on 2-D patterned substrate was strikingly similar to that previously observed during neutrophil migration within 3-D networks, suggesting that microfabricated materials provide relevant models of 3-D structures with precisely defined physical characteristics. In addition, our results demonstrate that the microgeometry of a substrate, when considered separately from adhesion, can play a significant role in cell migration.

Tan, J; Shen, H; Saltzman, W M



Fate of surface proteins of rabbit polymorphonuclear leukocytes during phagocytosis. I. Identification of surface proteins  

PubMed Central

To study the fate of external membrane proteins during phagocytosis, rabbit peritoneal neutrophils were labeled by enzymatic iodination. Iodine was incorporated into at least 13 proteins ranging in size from approximately 250,000 to 18,000 daltons as judged from autoradiography of gels after SDS-polyacrylamide gel electrophoresis of labeled cells. The major contractile proteins of neutrophils, actin and myosin, were not labeled when intact cells were iodinated but were labeled when homogenates of these cells were iodinated. Nine of the iodinated proteins were released by mild protease treatment of intact cells. A plasma membrane-rich fraction was isolated by density centrifugation. This fraction was enriched at least 10-fold for lactoperoxidase-labeled acid-insoluble proteins. It was enriched to the same extent for the presence of iodinated wheat germ agglutinin that had been bound to intact cells at 4 degrees C before homogenization. Analysis of SDS- polyacrylamide gel electrophoresis revealed that the proteins of this fraction were predominantly of high molecular weight. However, only 8 of the 13 proteins iodinated on intact cells were found in this fraction. The remaining five were enriched in a dense fraction containing nuclei, intact cells, and membranous vesicles, and may represent a specialized segment of the neutrophil cell surface.



Resistance of Capnocytophaga canimorsus to Killing by Human Complement and Polymorphonuclear Leukocytes  

Microsoft Academic Search

Capnocytophaga canimorsus is a bacterium of the canine oral flora known since 1976 to cause rare but severe septicemia and peripheral gangrene in patients that have been in contact with a dog. It was recently shown that these bacteria do not elicit an inflammatory response (H. Shin, M. Mally, M. Kuhn, C. Paroz, and G. R. Cornelis, J. Infect. Dis.

Hwain Shin; Manuela Mally; Salome Meyer; Chantal Fiechter; C. Paroz; U. Zaehringer; G. R. Cornelis



Novel post-translational incorporation of tyrosine in PMA-activated polymorphonuclear leukocytes (PMN)  

SciTech Connect

During studies undertaken to determine whether stimulation of tubulin tyrosinolation occurs in PMA-activated PMN, a distinctly different and novel post-translational incorporation of tyrosine into multiple PMN proteins was observed. The reaction also occurred in organelle-depleted neutrophil cytoplasts and was highly exaggerated in organelle-enriched karyogranuloplasts. The incorporation was specific for tyrosine, did not require extracellular Ca/sup 2 +/ and was inhibited in the presence of a variety of reducing agents, intracellular scavengers of oxygen radicals and inhibitors of peroxidase-mediated reactions. The PMA-induced incorporation of tyrosine was completely absent in PMN from patients with chronic granulomatous disease, but occurred normally in PMN of a patient with myeloperoxidase deficiency. Moreover, the incorporation of tyrosine was blocked by N-acetyl-L-tyrosine but not by phenylalanine suggesting a requirement for the phenolic group. A two-fold increase in stable protein carbonyl derivatives was demonstrated suggesting an increased oxidative modification of the proteins. SDS urea PAGE and reversed phase HPLC did not reveal any detectable changes in the extent of protein cross-linking. The PMN tyrosine pool was approximately 900 and yet only 1 tyrosine was added in these experiments. The functional significance of this reaction is not yet clear.

Nath, J.; Oliver, C.; Ohno, Y.; Gallin, J.I.



The inactivation of the polymorphonuclear leukocyte by non-steroidal anti-inflammatory drugs  

Microsoft Academic Search

When human neutrophils (PMNs) are activated by appropriate stimuli, they aggregate, generate Superoxide anion (O2-) and secrete lysosomal enzymes. Pre-incubation of PMNs in vitro with the cyclo-oxygenase (COx) inhibitor piroxicam (50µM) before stimulation with the chemotactic peptide f-met-leu-phe (FMLP, 10-7 M) inhibited all of these responses. The COx inhibitor ibuprofen inhibited FMLP-induced aggregation and lysozyme secretion, leaving O2- generation unaffected.

S. Abramson; H. Edelson; H. Kaplan; W. Given; G. Weissmann



Chemokines and leukocyte traffic  

Microsoft Academic Search

Twenty years after the discovery of chemokines is an appropriate time to review leukocyte traffic and to assess the knowledge and opportunities that have arisen from countless studies of the large and tight-knit family of chemotactic proteins.

Marco Baggiolini; Federica Sallusto



Getting Leukocytes to the Site of Inflammation  

PubMed Central

There is no “response” in either the innate or adaptive immune response unless leukocytes cross blood vessels. They do this through the process of diapedesis, in which the leukocyte moves in ameboid fashion through tightly apposed endothelial borders (paracellular transmigration) and in some cases through the endothelial cell itself (transcellular migration). This review summarizes the steps leading up to diapedesis, then focuses on the molecules and mechanisms responsible for transendothelial migration. Surprisingly, many of the same molecules and mechanisms that regulate paracellular migration also control transcellular migration, including a major role for membrane from the recently described lateral border recycling compartment. A hypothesis that integrates the various known mechanisms of transmigration is proposed.

Muller, W. A.



Comparison of leukocyte count and function in smoking and nonsmoking young men.  

PubMed Central

Leukocyte function and other hematological measurements were tested in 14 smoking and 13 nonsmoking young men free of intercurrent or chronic diseases. Leukocyte chemotaxis was depressed in smoking subjects when compared to the same subjects abstaining from cigarettes or to the nonsmokers. Smoking did not affect the whole blood bactericidal capacity of leukocytes and serum for Staphyloccus aureus or Klebsiella pneumoniae. Total leukocyte counts, hematocritis, and monocyte counts were higher in the smoking subjects when compared to the nonsmokers.

Noble, R C; Penny, B B



Coupled flow-structure-biochemistry simulations of dynamic systems of blood cells using an adaptive surface tracking method  

NASA Astrophysics Data System (ADS)

A method for the computation of low-Reynolds number dynamic blood cell systems is presented. The specific system of interest here is interaction between cancer cells and white blood cells in an experimental flow system. Fluid dynamics, structural mechanics, six-degree-of-freedom motion control, and surface biochemistry analysis components are coupled in the context of adaptive octree-based grid generation. Analytical and numerical verification of the quasi-steady assumption for the fluid mechanics is presented. The capabilities of the technique are demonstrated by presenting several three-dimensional cell system simulations, including the collision/interaction between a cancer cell and an endothelium adherent polymorphonuclear leukocyte (PMN) cell in a shear flow.

Hoskins, M. H.; Kunz, R. F.; Bistline, J. E.; Dong, C.



Effects of Physiological Concentrations of Heavy Metals Both Individually and in Mixtures on the Viability and Function of Peripheral Blood Human Leukocytes In Vitro  

Microsoft Academic Search

Among environmental contaminants recognized for their toxicity and global distribution, heavy metals are elements known to exert serious ecological consequences. Published experiments on the immunotoxic effects of metals such as methylmercury (MeHg), cadmium (Cd), and lead (Pb) were often conducted at concentrations higher than those present in the environment or those in human blood. In the present study the in

M. Fortier; F. Omara; J. Bernier; P. Brousseau; M. Fournier





... solid part of your blood contains red blood cells, white blood cells, and platelets. Red blood cells deliver oxygen from your lungs to your tissues and organs. White blood cells fight infection and are part of your body's ...


Connective tissue-degrading enzymes of human leukocytes.  


Human leukocytes, when exposed to aggregated human gamma-globulin (AHGG) or immune complexes (isolated from RA synovial fluid) fixed to a cartilagenous surface, release neutral proteases that degrade the extracellular matrix of cartilage. The chondromucoprotein destruction by these proteases is suppressed by a variety of synovial fluids but is not susceptible to inhibition by trypsin, chymotrypsin, elastase inhibitors, or a combination of these agents. The inhibitory effect of synovial fluid can be reversed in the presence of increasing enzyme concentrations. Intact viable human polymorphonuclear leukocytes in the presence of AHGG also release a collagenase precursor that can be activated by limited proteolysis with trypsin or RA synovial fluids. Enzyme release (neutral proteases) by phagocytosing cells is inhibited by the antiinflammatory agents phenylbutazone and colchicine; these agents do not affect release of the collagenase precursor. However, the latent collagenase release is susceptible to inhibition when leukocytes are preincubated (prior to exposure to AHGG) with inhibitors of protein synthesis. Under these conditions, neutral protease release is unaffected. PMID:51602

Oronsky, A L; Perper, R J




Technology Transfer Automated Retrieval System (TEKTRAN)

Polymorphonuclear neutrophil leukocytes (PMN) form the first line of cellular defense against invading pathogens. The PMN is characterized by a polymorphic segmented nucleus, numerous cytoplasmic granules that provide constituents for killing bacteria, large stores of glycogen for energy and a high...



Technology Transfer Automated Retrieval System (TEKTRAN)

Aberrations in adhesive mechanisms account for several clinical syndromes that involve an increase in susceptibility to bacterial infection. Leukocyte adhesion deficiency I (LAD I) (MIM 116920) was the first to be defined at a molecular level; it results from mutations in CD18, the beta subunit of b...


Sulfatides inhibit leukotriene synthesis in human polymorphonuclear granulocytes by a mechanism involving lipid rearrangement in intracellular membranes.  


Sulfatides - sulfated derivatives of galactocerebroside - are endogenous ligands for P- and L-selectins and are able to induce intracellular signaling in neutrophils through a L-selectin dependent pathway. Sulfatides are implicated in a variety of physiological functions and have been found to suppress the synthesis of 5-lipoxygenase (5-LO) metabolites and impede 5-LO translocation to the nuclear envelope in adherent human polymorphonuclear leukocytes (PMNs) [Sud'ina, G. F., Brock, T. G., Pushkareva, M. A., Galkina, S. I., Turutin, D. V., Peters-Golden, M., et al. (2001). Sulphatides trigger polymorphonuclear granulocyte spreading on collagen-coated surfaces and inhibit subsequent activation of 5-lipoxygenase. The Biochemical Journal, 359, 621-629]. In this study we investigated the mechanism of the leukotriene (LT) synthesis inhibition by sulfatides. Sulfatides neither attenuated the ionophore-induced rise in [Ca(2+)](i) nor promoted PKA activation. We demonstrated that sulfatides directly inhibited 5-LO enzyme activity in a cell-free assay. BODIPY-labeled sulfatides were able to rapidly penetrate into the cells. Sulfatides induced rearrangement and redistribution of cytoskeletal components in adherent PMNs. The lipid incorporation as well as sulfatide-induced inhibition of LT synthesis were abolished by cytochalasin D, an inhibitor of actin polymerization and endocytosis. Importantly, sulfatides caused a prominent intracellular cholesterol redistribution, increasing its abundance at the uropod region. On the basis of these data, we suggest that increased cholesterol accumulation in cell compartments represents a novel mechanism by which sulfatides abrogate 5-LO translocation and activation. PMID:17822942

Grishina, Zoryana V; Pushkareva, Marina A; Pletjushkina, Olga Yu; Reiser, Georg; Peters-Golden, Marc; Sud'ina, Galina F



Chemotaxis and chemiluminescence responses of synovial fluid polymorphonuclear leucocytes during acute reactive arthritis.  

PubMed Central

The chemotaxis and chemiluminescence responses of polymorphonuclear leucocytes (PMN) of synovial fluid and peripheral blood from patients with acute reactive arthritis were studied. Rates of chemotactic and chemokinetic migration of synovial fluid PMN were significantly decreased. In addition, chemiluminescence responses tended to be depressed, suggesting that the cells were deactivated for both chemotaxis and production of oxygen derived free radicals. Such deactivation has been described previously as a characteristic of synovial fluid PMN in rheumatoid arthritis. Compared with those with a mild disease, patients with severe acute reactive arthritis had higher chemiluminescence responses of synovial fluid PMN to phorbol myristate acetate during acute disease and developed increased migration of peripheral blood PMN towards zymosan treated serum after recovery from the disease. This supports the view that hyperreactive PMN contribute to the development of severe inflammatory symptoms in acute reactive arthritis.

Leirisalo-Repo, M; Lauhio, A; Repo, H



The effect of leukocyte hydrolases on bacteria  

Microsoft Academic Search

Small amounts of human leukocyte extracts (ENZ), inflammatory exudates, lysozyme (LYZ), and a variety of neutral proteases are capable of releasing (solubilizing) lipopolysaccharides (LPS) fromSalmonella typhi. LPS activity was determined by its capacity to sensitize red blood cells (RBC) passively to agglutination by antisalmonella serum. While the LPS-releasing capacity of ENZ, inflammatory exudates, and trypsin can be inhibited by phenyl-methyl-sulfonyl

Mina Ferne; Zvia Duchan; Sonia Rabinowitz-Begner; Michael N. Sela; Isaac Ginsburg



The effect of leukocyte hydrolases on bacteria  

Microsoft Academic Search

Lysis of14C-labeledStaph. aureus by human blood leukocyte lysates, by extracts of rabbit small intestines and pancreas, and by the “cocktail” of enzymes (containing trypsin, lysolecithin, and lysozyme) is strongly inhibited by anionic polyelectrolytes (e.g., heparin, chondroitin sulfate, liquoid (polyanethole sulfonic acid), and DNA). Most of the lytic agents employed were inhibited by cationic polyelectrolytes (e.g., histone, protamin sulfate and polylysin),

Michael N. Sela; Meir Lahav; Nurit Ne'eman; Zvia Duchan; Isaac Ginsburg



Peripheral blood stem cell transplantation from human leukocyte antigen-matched sibling donors and unrelated donors in acute myeloid leukemia patients.  


There have been rare comparative studies of hematopoietic stem cell transplantation from matched sibling donors (MSDs) and unrelated donors (URDs) with regard to peripheral blood stem cell transplantation (PBSCT). We performed a retrospective study of 104 consecutive acute myeloid leukemia (AML) patients who had received an allogeneic PBSCT from an MSD or a URD in order to compare transplant outcomes and posttransplant complications between the 2 groups of patients. The cumulative incidence of grade 2-4 acute graft-versus-host disease (aGVHD) at 100 days (22.6% with MSD vs. 35.3% with URD; p = 0.107) and that of chronic GVHD (cGVHD) at 2 years (72.9% with MSD vs. 56.1% with URD; p = 0.153) was not significantly different between the 2 groups. Multivariate analysis also indicated that a URD was not an independent predictor of grade 2-4 aGVHD or cGVHD. No statistically significant differences were observed in terms of relapse incidence (p = 0.371), nonrelapse mortality (p = 0.473), disease-free survival (p = 0.925) or overall survival (p = 0.534) at 2 years. URDs are comparable with MSDs as a donor type for PBSCT in AML patients if risk-stratified GVHD prophylaxis is adopted. PMID:23816761

Kim, Hee-Je; Kim, Sung-Yong; Lee, Mark Hong; Min, Woo-Sung



Sequential Chemotactic and Phagocytic Activation of Human Polymorphonuclear Neutrophils  

Microsoft Academic Search

Received 14 March 2007\\/Returned for modification 27 April 2007\\/Accepted 17 May 2007 Human polymorphonuclear neutrophils (PMN) chemotax to a foreign entity. When the chemoattractants' origins are reached, specific receptors bind to the invader's surface, initiating phagocytosis, phagosome formation, and fusion with granule membranes, generating the bactericidal oxidative burst, and releasing lytic enzymes, specific peptides, and proteins. We explored the initial

Jens Martin Herrmann; John Bernardo; Heidi J. Long; Kurt Seetoo; Mary E. McMenamin; Eraldo L. Batista; T. E. Van Dyke; E. R. Simons



Alterations in Blood-Brain Barrier Permeability to Large and Small Molecules and Leukocyte Accumulation after Traumatic Brain Injury: Effects of Post-Traumatic Hypothermia  

PubMed Central

Abstract We investigated the temporal and regional profile of blood-brain barrier (BBB) permeability to both large and small molecules after moderate fluid percussion (FP) brain injury in rats and determined the effects of post-traumatic modest hypothermia (33°C/4?h) on these vascular perturbations. The visible tracers biotin-dextrin-amine 3000 (BDA-3K, 3?kDa) and horseradish peroxidase (HRP, 44?kDa) were injected intravenously at 4?h or 3 or 7 days post-TBI. At 30?min after the tracer infusion, both small and large molecular weight tracers were detected in the contusion area as well as remote regions adjacent to the injury epicenter in both cortical and hippocampal structures. In areas adjacent to the contusion site, increased permeability to the small molecular weight tracer (BDA-3K) was evident at 4?h post-TBI and remained visible after 7 days survival. In contrast, the larger tracer molecule (HRP) appeared in these remote areas at acute permeable sites but was not detected at later post-traumatic time periods. A regionally specific relationship was documented at 3 days between the late-occurring permeability changes observed with BDA-3K and the accumulation of CD68-positive macrophages. Mild hypothermia initiated 30?min after TBI reduced permeability to both large and small tracers and the infiltration of CD68-positive cells. These results indicate that moderate brain injury produces temperature-sensitive acute, as well as more long-lasting vascular perturbations associated with secondary injury mechanisms.

Lotocki, George; de Rivero Vaccari, Juan Pablo; Perez, Enrique R.; Sanchez-Molano, Juliana; Furones-Alonso, Ofelia; Bramlett, Helen M.



Ontogenetic regulation of leukocyte recruitment in mouse yolk sac vessels.  


In adult mammals, leukocyte recruitment follows a well-defined cascade of adhesion events enabling leukocytes to leave the circulatory system and transmigrate into tissue. Currently, it is unclear whether leukocyte recruitment proceeds in a similar fashion during fetal development. Considering the fact that the incidence of neonatal sepsis increases dramatically with decreasing gestational age in humans, we hypothesized that leukocyte recruitment may be acquired only late during fetal ontogeny. To test this, we developed a fetal intravital microscopy model in pregnant mice and, using LysEGFP (neutrophil reporter) mice, investigated leukocyte recruitment during fetal development. We show that fetal blood neutrophils acquire the ability to roll and adhere on inflamed yolk sac vessels during late fetal development, whereas at earlier embryonic stages (before day E15), rolling and adhesion were essentially absent. Accordingly, flow chamber experiments showed that fetal EGFP(+) blood cells underwent efficient adhesion only when they were harvested on or after E15. Fluorescence-activated cell sorter analysis on EGFP(+) fetal blood cells revealed that surface expression of CXCR2 and less pronounced P-selectin glycoprotein ligand-1 (PSGL-1) begin to increase only late in fetal life. Taken together, our findings demonstrate that inflammation-induced leukocyte recruitment is ontogenetically regulated and enables efficient neutrophil trafficking only during late fetal life. PMID:23525796

Sperandio, Markus; Quackenbush, Elizabeth J; Sushkova, Natalia; Altstätter, Johannes; Nussbaum, Claudia; Schmid, Stephan; Pruenster, Monika; Kurz, Angela; Margraf, Andreas; Steppner, Alina; Schweiger, Natalie; Borsig, Lubor; Boros, Ildiko; Krajewski, Nele; Genzel-Boroviczeny, Orsolya; Jeschke, Udo; Frommhold, David; von Andrian, Ulrich H



Towards a computational model of leukocyte adhesion cascade: Leukocyte rolling  

NASA Astrophysics Data System (ADS)

Recruitment of leukocytes into sites of acute and chronic inflammation is a vital component of the innate immune response in humans and plays an important role in cardiovascular diseases, such as ischemia-reperfusion injury and atherosclerosis. Leukocytes extravasate into the inflamed tissue through a multi-step process called "leukocyte adhesion cascade", which involves initial contact of a leukocyte with activated endothelium (tethering), leukocyte rolling, firm adhesion, and transendothelial migration. Recently we developed a fully three-dimensional CFD model of receptor-mediated leukocyte adhesion to endothelium in a parallel-plate flow chamber. The model treats the leukocyte as a viscoelastic cell with the nucleus located in the intracellular space and cylindrical microvilli distributed over the cell membrane. Leukocyte-endothelial adhesion is assumed to be mediated by adhesion molecules expressed on the tips of cell microvilli and on endothelium. We show that the model can predict both shape changes and velocities of rolling leukocytes under physiological flow conditions. Results of this study also indicate that viscosity of the cytoplasm is a critical parameter of leukocyte adhesion, affecting the cell's ability to roll on endothelium. This work is supported by NIH Grant HL- 57446 and NCSA Grant BCS040006 and utilized the NCSA IBM p690.

Khismatullin, Damir



phagocytosis of apoptotic leukocytes  

Microsoft Academic Search

Various types of phagocytes mediate the clearance of apoptotic cells. We previously reported that human and murine high endothelial venule (HEV) cells ingest apoptotic cells. In this report, we examined endothelial cell fucoidin re- ceptor-mediated phagocytosis using a murine en- dothelial cell model mHEV. mHEV cell recognition of apoptotic leukocytes was blocked by fucoidin but not by other phagocytic receptor

Jacob D. Johnson; Krista L. Hess; Joan M. Cook-Mills


Characterization of uterine leukocyte infiltration in gilts after artificial insemination.  


The objective of this study was to characterize the uterine leukocyte influx after artificial insemination (AI). After detection of oestrus with a boar at intervals of 1.5 h, seventy-two gilts were randomly assigned to a 2 x 3 x 4 factorial arrangement. AI was performed with 100 ml extended semen containing 5 x 10(9) spermatozoa (semen; n = 36) or 100 ml VSP semen extender (extender; n = 36) at one of three times after detection of oestrus: 12, 24 or 36 h (n = 24/time). The uterus was lavaged at 6, 12, 18 or 24 h (n = 18/time) after AI to determine the total number of uterine leukocytes. In addition, uterine lavage was performed on nine untreated gilts immediately after the detection of oestrus to establish a baseline number of leukocytes. The leukocyte response in all samples consisted predominately (92-99%) of polymorphonuclear neutrophilic granulocytes (PMNs). The mean number of PMNs recovered from the uteri of gilts treated with semen was greater than in gilts treated with extender and in untreated gilts (P < 0.01). The greatest number of PMNs in semen-treated gilts was found 12 h after AI (P < 0.01), and this number was sustained for 24 h. In contrast, the number of uterine PMNs recovered from extender-treated gilts reached a peak at 6 h and had declined by 12 h after AI (P < 0.05). It was concluded that an extensive influx of PMNs into the uterus is a normal sequence to AI. The consequences and importance of semen-induced uterine leukocytosis needs further investigation. PMID:10070347

Rozeboom, K J; Troedsson, M H; Crabo, B G



Effect of cryoprotectants on the viability and function of unfrozen human polymorphonuclear cells.  


High concentrations of membrane permeable cryoprotectants are necessary to protect human polymorphonuclear leukocytes from osmotic stress injury during freezing, but there are reports that some cryoprotectants are chemically toxic. Cells were exposed to various concentrations of glycerol, dimethyl sulfoxide, or ethylene glycol for 5 min to 2 hr at 37, 22 or 0 degree C, adding or removing the cryoprotectant either slowly or rapidly. Assays included cell number recovery, membrane integrity, phagocytosis, microbicidal ability, and chemotaxis. We conclude that (1) 1 and 2 M concentrations generally are not toxic if they are added and removed slowly at 22 degrees C; (2) addition and removal of glycerol at 0 degree C was injurious even at 1 M; (3) slow addition and removal allowed better recovery than rapid addition or removal; (4) salt concentration in cryoprotectant solutions should be adjusted to isotonic on the basis of moles per liter of solution, rather than moles per kilogram of water; (5) the toxicity reported by other investigators can be largely explained by osmotic stress or dilution shock rather than chemical toxicity; and (6) ethylene glycol is the easiest cryoprotectant to add to and remove from these cells. PMID:4028780

Takahashi, T; Bross, J B; Shaber, R E; Williams, R J



MMP-8 promotes polymorphonuclear cell migration through collagen barriers in obliterative bronchiolitis  

PubMed Central

Increased levels of MMP-8 (neutrophil collagenase) have been reported in OB, but the biological role of MMP-8 in OB is not known. MMP-8 is an interstitial collagenase highly expressed by polymorphonuclear leukocytes, which are prominent in early OB. Here, we show that MMP-8 promotes migration of PMNs through the collagen-rich matrix in a mouse heterotopic airway transplant model of OB. Overall, MMP-8?/? mice had significantly fewer PMNs in the airway lumen 2 and 14 days post-transplantation, and the percentage of PMNs traversing the matrix to the lumen was decreased markedly in the MMP-8?/? compared with WT mice at 14 days. There were significantly more PMNs outside of the lumen in the ECM in the MMP-8?/? mice compared with WT mice. In vitro, significantly fewer MMP-8?/? PMNs migrated through 3D cross-linked collagen gels than WT PMNs. MMP inhibitor GM6001 was also able to impede migration of WT PMNs through collagen gels. The decreased migration was likely a result of pericollagenase activity of MMP-8, as WT PMNs expressing MMP-8 were not able to migrate effectively through collagen that was resistant to the collagenase. Protection from OB was seen in the MMP-8?/? mice, as the airway lumen had significantly less obliteration and collagen deposition, suggesting that MMP-8 plays an important role in the pathogenesis of OB.

Khatwa, Umakanth A.; Kleibrink, Bjoern E.; Shapiro, Steven D.; Subramaniam, Meera



Preparation of leukocyte-poor platelet concentrates from buffy coats. III. Effect of leukocyte contamination on storage conditions.  


To study the influence of contaminating leukocytes on the storage conditions of platelet concentrates (PC), various amounts of leukocytes were added to identical PC. From 12 blood donations, 12 leukocyte-poor PC were prepared and pooled. Subsequently, the pool was divided into 12 identical PC. The plasma volume of the PC was 58.6 +/- 0.6 ml, the platelet concentration was 1.01 +/- 0.04 x 10(9)/ml (mean +/- SD) and the red cell contamination did not exceed 10(7) per PC. To 4 groups of 3 PC, pooled leukocytes were added from the same 12 blood donations. The leukocyte contamination for each group of 3 PC was 0.14 +/- 0.05, 1.96 +/- 0.09, 5.53 +/- 0.98 and 13.0 +/- 0.93 x 10(6)/ml (mean +/- SD) for groups I-IV, respectively. The PC were stored for 7 days at 22 degrees C in normal polyvinylchloride bags. A significant correlation was found between increasing concentrations of leukocytes in the PC and the drop in pH (r = -0.93), glucose consumption (r = -0.91), lactic acid production (r = 0.93) and release of lactate dehydrogenase (r = 0.92) during storage of the PC. The excretion of beta-thromboglobulin, depletion of platelet adenine nucleotides, decreased ability to incorporate 3H-adenosine into metabolic nucleotides and poor morphology of the platelets were also significantly correlated with an increased number of leukocytes in the PC. These data show that high concentrations of leukocytes in PC have a significant detrimental effect on the viability of platelets during storage at 22 degrees C. We conclude that for good storage conditions of PC, the upper limit of leukocytes per PC should not exceed 10(7). PMID:2971295

Pietersz, R N; de Korte, D; Reesink, H W; van den Ende, A; Dekker, W J; Roos, D



First characterization of a teleost Epstein-Barr virus-induced gene 3 (EBI3) reveals a regulatory effect of EBI3 on the innate immune response of peripheral blood leukocytes.  


Epstein-Barr virus-induced gene 3 (EBI3) encodes a protein that in mammals is known to be a subunit of interleukin (IL)-27 and IL-35, both which regulate cytokine production and inflammatory response. To date, no studies on fish EBI3 have been documented. In this work, we report the identification of an EBI3 homologue, CsEBI3, from tongue sole (Cynoglossus semilaevis) and analysis of its expression and biological effect. CsEBI3 is composed of 245 amino acid residues and possesses a Fibronectin type 3 (FN3) domain that is preserved in lower and higher vertebrates. Expression of CsEBI3 was detected in a wide range of tissues, in particular those of immune relevant organs, and upregulated in a time-dependent manner by experimental challenge with bacterial and viral pathogens. Bacterial infection of peripheral blood leukocytes (PBL) enhanced CsEBI3 expression and caused extracellular secretion of CsEBI3. Purified recombinant CsEBI3 (rCsEBI3) stimulated the respiratory burst activity of PBL and upregulated the expression of IL-1?, IL-8, Myd88, interferon-induced gene 15, CD28, and chemokines. In contrast, rCsEBI3M, a mutant CsEBI3 that lacks the FN3 domain failed to activate PBL and induced much weaker expression of the immune genes. Treatment of PBL with rCsEBI3, but not with the mutant rCsEBI3M, enhanced cellular resistance against bacterial invasion, whereas antibody blocking of CsEBI3 on PBL significantly reduced cellular resistance against bacterial infection. Taken together, these results indicate for the first time that a teleost EBI3 possesses immunoregulatory property in a manner that is dependent on the conserved FN3 domain, and that CsEBI3 is involved in the innate immune defense of PBL against microbial pathogens. PMID:23932982

Li, Mo-Fei; Sun, Bo-Guang; Xiao, Zhi-Zhong; Sun, Li



Analysis of serotonin receptor 2A gene (HTR2A): association study with autism spectrum disorder in the Indian population and investigation of the gene expression in peripheral blood leukocytes.  


Several studies suggest involvement of serotoninergic system in the pathophysiology of Autism Spectrum Disorder (ASD). The 5-HT receptor binding studies using (3)H-lysergic acid diethylamide ((3)H-LSD) and linkage analysis provided evidences to consider HTR2A as a potential candidate gene for ASD. The three SNPs, -1438A/G (rs6311), 102T/C (rs6313) and 1354C/T (rs6314) of HTR2A have been well studied in the etiology of various neuropsychiatric disorders. But studies on association of this gene with ASD are limited to two reports from American and Korean populations. Additionally there are reports, which demonstrated paternal imprinting of HTR2A with expression from only one allele. So far no reports are available on HTR2A and its association with any neuropsychiatric disorders from Indian population. Therefore, the present study investigates association of the above mentioned three markers of HTR2A with ASD in Indian population using population and family-based approaches. The study also deals with allelic expression pattern of HTR2A in Peripheral Blood Leukocytes (PBLs) to understand the parental imprinting status. The genotyping analyses were carried out for probands, parents and controls. The subsequent association analyses did not show association of these markers with ASD. So, HTR2A is unlikely to be a genetic marker for ASD in Indian population. The expression analyses showed absence of monoallelic expression, suggesting lack of parental imprinting of HTR2A gene. However, we noticed methylation of the CpG sites at -1438A/G and 102T/C loci of HTR2A gene. Further bioinformatics analysis revealed absence of CpG islands in the promoter of the gene supporting biallelic expression pattern of HTR2A in PBLs. PMID:19647026

Guhathakurta, Subhrangshu; Singh, Asem Surindro; Sinha, Swagata; Chatterjee, Anindita; Ahmed, Shabina; Ghosh, Saurabh; Usha, Rajamma



Immunoglobulin-like transcript 7 (ILT7) but not bone marrow stromal cell antigen 2 (BST2, aka HM1.24, tetherin or CD317) modulates plasmacytoid dendritic cell function (pDC) in primary human blood leukocytes  

PubMed Central

The immunoglobulin-like transcript (ILT) 7 is a surface molecule selectively expressed by human plasmacytoid dendritic cells (pDCs). ILT7 cross-linking suppresses pDC activation and type I interferon (IFN-I) secretion following Toll-like receptors (TLR)7/9 engagement. The bone marrow stromal cell antigen 2 (BST2, aka HM1.24, tetherin or CD317) is expressed by different cell types upon exposure to IFN-I and is a natural ligand for ILT7. Here we show that ILT7 expression decreased spontaneously in pDCs upon in vitro culture, which correlates with pDC differentiation measured as increased side scatter properties and CCR7 expression. TLR7/9 Ligands , as well as HIV, induced BST2 upregulation on all tested cell types except T cells, which required TCR stimulation to respond to TLR9L-induced IFN-I. IFN-?, IL-4, IL-10 and TNF-? had only marginal effects on BST2 expression in blood leukocytes compared to TLR9L. Pre-incubation with ILT7-crosslinking Ab inhibited IFN-I production in PBMCs treated with TLR7/9L or HIV, whereas BST2 blockade did not affect IFN-I responses even when BST2 upregulation was further boosted with TCR agonists or immunoregulatory cytokines. Our data indicate that BST2-mediated ILT7 cross-linking may act as a homeostatic regulatory mechanism on immature circulating pDC, rather than a negative feedback for activated mature pDCs which have downregulated ILT7.

Tavano, Barbara; Galao, Rui Pedro; Graham, David R; Neil, Stuart JD; Aquino, Veronica N; Fuchs, Dietmar; Boasso, Adriano



Leukocyte labeling with technetium-99m tin colloids  

SciTech Connect

Triple density gradients of metrizamide in plasma (MP) were used to characterize label distribution in human leukocyte preparations incubated with /sup 99m/Tc tin colloids. Less than 50% of the cell-associated radioactivity was specifically bound to leukocytes when heparinized blood was rotated with stannous fluoride colloid ((Tc)SFC). Labeling efficiency in leukocyte rich plasma (LRP) averaged 44%, of which greater than 90% was specifically bound to leukocytes. MP-gradient analysis also revealed that leukocyte labeling did not occur with stannous chloride colloid, nor when citrate was present during rotation with (Tc)SFC. When citrate was added after labeling to solubilize unbound (Tc)SFC, radiocolloid was removed from the leukocytes, indicating that the mechanism of (Tc)SFC labeling is adherence rather than phagocytosis. Technetium-labeled neutrophils exhibited normal in vitro chemotaxis and no lung uptake in vivo. Technetium-labeled mononuclear leukocytes, on the other hand, exhibited prolonged lung transit in vivo. Neither (Tc)SFC cell preparation showed signs of in vivo reoxidation to pertechnetate.

Mock, B.H.; English, D.



B lymphocyte stimulator (BLyS) isoforms in systemic lupus erythematosus: disease activity correlates better with blood leukocyte BLyS mRNA levels than with plasma BLyS protein levels  

PubMed Central

Considerable evidence points to a role for B lymphocyte stimulator (BLyS) overproduction in murine and human systemic lupus erythematosus (SLE). Nevertheless, the correlation between circulating levels of BLyS protein and disease activity in human SLE is modest at best. This may be due to an inadequacy of the former to reflect endogenous BLyS overproduction faithfully, in that steady-state protein levels are affected not just by production rates but also by rates of peripheral utilization and excretion. Increased levels of BLyS mRNA may better reflect increased in vivo BLyS production, and therefore they may correlate better with biologic and clinical sequelae of BLyS overexpression than do circulating levels of BLyS protein. Accordingly, we assessed peripheral blood leukocyte levels of BLyS mRNA isoforms (full-length BLyS and ?BLyS) and plasma BLyS protein levels in patients with SLE, and correlated these levels with laboratory and clinical features. BLyS protein, full-length BLyS mRNA, and ?BLyS mRNA levels were greater in SLE patients (n = 60) than in rheumatoid arthritis patients (n = 60) or normal control individuals (n = 30). Although full-length BLyS and ?BLyS mRNA levels correlated significantly with BLyS protein levels in the SLE cohort, BLyS mRNA levels were more closely associated with serum immunoglobulin levels and SLE Disease Activity Index scores than were BLyS protein levels. Moreover, changes in SLE Disease Activity Index scores were more closely associated with changes in BLyS mRNA levels than with changes in BLyS protein levels among the 37 SLE patients from whom repeat blood samples were obtained. Thus, full-length BLyS and ?BLyS mRNA levels are elevated in SLE and are more closely associated with disease activity than are BLyS protein levels. BLyS mRNA levels may be a helpful biomarker in the clinical monitoring of SLE patients.

Collins, Christopher E; Gavin, Amanda L; Migone, Thi-Sau; Hilbert, David M; Nemazee, David; Stohl, William



Pulsatility of Parafoveal Capillary Leukocytes  

PubMed Central

The use of adaptive optics (AO) in a confocal scanning laser ophthalmoscope (AOSLO) allows for long-term imaging of parafoveal capillary leukocyte movement and measurement of leukocyte velocity without contrast dyes. We applied the AOSLO to investigate the possible role of the cardiac cycle on capillary leukocyte velocity by directly measuring capillary leukocyte pulsatility. The parafoveal regions of 8 eight normal healthy subjects with clear ocular media were imaged with an AOSLO. All subjects were dilated and cyclopleged. The AOSLO field of view was either 1.4 × 1.5 degrees or 2.35 × 2.5 degrees, the imaging wavelength was 532 nm and the frame rate was 30 fps. A photoplethysmograph was used to record the subject’s pulse synchronously with each AOSLO video. Parafoveal capillary leukocyte velocities and pulsatility were determined for two or three capillaries per subject. Leukocyte velocity and pulsatility were determined for all eight subjects. The mean parafoveal capillary leukocyte velocity for all subjects was Vmean = 1.30 mm/sec (SD = +/? 0.40 mm/sec). There was a statistically significant difference between leukocyte velocities, Vmax and Vmin, over the pulse cycle for each subject (p<0.05). The mean pulsatility was Pmean= 0.45 (+/? 0.09). Parafoveal capillary leukocyte pulsatility can be directly and non-invasively measured without the use of contrast dyes using an AOSLO. A substantial amount of the variation found in leukocyte velocity is due to the pulsatility that is induced by the cardiac cycle. By controlling for the variation in leukocyte velocity caused by the cardiac cycle, we can better detect other changes in retinal leukocyte velocity induced by disease or pharmaceutical agents.

Martin, Joy A.; Roorda, Austin



Broad spectrum antimicrobial activity of leukocyte extracts from the American alligator ( Alligator mississippiensis)  

Microsoft Academic Search

Leukocytes were isolated from whole blood of wild alligators by differential sedimentation. The leukocytes were disrupted in 5% AcOH and the crude extracts processed by ultrafiltration. The extracts were subjected to solvent exchange (0.1% AcOH) and the fraction that contained macromolecules between 1 and 10kDa were subjected to further analyses. The acid extracts of the alligator leukocytes exhibited substantial antimycotic

Mark E. Merchant; Noelle Leger; Erin Jerkins; Kaili Mills; Melanie B. Pallansch; Robin L. Paulman; Roger G. Ptak



Radiographic evaluation of destructive periodontal disease in blue mink in relation to age and blood morphology  

PubMed Central

Abstract In this study, blood samples and jaws were collected from 2 genotypes of blue mink (n = 289) in order to examine phenotypic expression of specific characteristics of Chediak-Higashi Syndrome (C-HS). Blood samples were subjected to differential counts to assess the proportion of abnormal polymorphonuclear leukocytes characteristic for CH-S (C-HS-leukocytes). Abnormal leukocytes with characteristic signs of C-HS were found in blood smears from all mink included in this study. Four teeth in one half of the mandible (P3, P4, M1, M2) were subjected to quantitative radiographic evaluation of alveolar bone loss and tooth loss. There was a high prevalence of destructive periodontal disease among blue mink included in this study. Mild to moderate periodontal disease (defined by less than 50% alveolar bone loss related to 1 or more teeth) affected 73.7% of young mink (age = 7 mo) and 67.9% of older animals (age ? 19 mo). Severe periodontal disease (defined by more than 50% bone loss related to one or more teeth) was not detected in mink aged 7 mo, but affected 15.3% of mink aged 19 mo and 39.6% of mink aged 31 mo. The positive relationship between age and periodontal disease was statistically significant (P < 0.01). The prevalence of tooth loss was found to be high among blue mink aged >19 mo (21.6%) and was also significantly related to age (P < 0.01). A significant positive interaction between alveolar bone loss and tooth loss (P < 0.01), implies that the highly prevalent tooth loss in the mink was related to and possibly caused by destructive periodontal disease. There was no significant difference in the prevalence of periodontal disease between the 2 genotypes and age was found to be the only statistical predictor of poor production results (P < 0.01) in blue mink.



Polymorphonuclear neutrophil chemotaxis modulated by Bacteroides fragilis peptidoglycan.  

PubMed Central

Peptidoglycan was isolated from Bacteroides fragilis with boiling sodium dodecyl sulfate, and some was treated with pronase to eliminate contaminating protein. This peptidoglycan was chemotactic for rabbit polymorphonuclear neutrophils and had even greater chemotactic activity along with some chemokinetic activity after it was partially hydrolyzed with lysozyme. Significant chemotaxis-inhibitory activity was observed for an acid-precipitable component of the lysozyme-treated crude peptidoglycan of B. fragilis.

Sperry, J F; Burns, J M



The polymorphonuclear leucocyte count in childhood haemolytic uraemic syndrome  

Microsoft Academic Search

Review of data from 79 children with the haemolytic uraemic syndrome (HUS) showed that the polymorphonuclear leucocyte (PMN) count at presentation in childhood HUS predicts outcome. Logistic regression analysis of several features at presentation identified only the PMN count and the presence of a diarrhoeal prodrome as having a significant effect on the outcome (PPt-test on log-transformed data,PPP<0.001). Multiple regression

Martin D. S. Walters; I. Ute Matthei; Richard Kay; Michael J. Dillon; T. Martin Barratt



Effects of Eugenol on Polymorphonuclear Cell Migration and Chemiluminescence  

Microsoft Academic Search

In this study, the effects of eugenol on human polymorphonuclear (PMN) cell migration and chemiluminescence were examined in vitro. Utilizing zymosan-activated serum or crude Bacteroides sonicate fractions as chemotractants, we found that eugenol inhibits PMN migration at 6.6 x 10-2to 6.6 x 10-5mol\\/L (P<0.05). Also, similar effects were observed in PMNs pre-incubated in eugenol. Regardless of concentration, eugenol was not

P. G. Fotos; C. J. Woolverton; K. Van Dyke; R. L. Powell



Protein synthesis rates of human PBMC and PMN can be determined simultaneously in vivo by using small blood samples.  


Immune cell functions can be evaluated in vivo by measuring their specific protein fractional synthesis rates (FSR). Using stable isotope dilution techniques, we describe a new method allowing simultaneous in vivo assessment of FSR in two leukocyte populations in healthy human subjects, using small blood samples. Peripheral blood mononuclear cell (PBMC) and polymorphonuclear neutrophil (PMN) FSR were measured during primed continuous intravenous infusion of l-[1-(13)C]leucine. Immune cells from 6 ml of whole blood were isolated by density gradient centrifugation. In a first study, we calculated the FSR using plasma [(13)C]leucine or alpha-[(13)C]ketoisocaproate (KIC) enrichments as precursor pools. In a second study, we compared protein FSR in leukocytes, using enrichments of either intracellular or plasma free [(13)C]leucine as immediate precursor pools. The present approach showed a steady-state enrichment of plasma and circulating immune cell free [(13)C]leucine precursor pools. The linearity of labeled amino acid incorporation rate within mixed PBMC and PMN proteins also was verified. Postabsorptive protein FSR was 4.09 +/- 0.39%/day in PBMC and 1.44 +/- 0.08%/day in PMN when plasma [(13)C]KIC was the precursor pool. The difference between PBMC and PMN FSR was statistically significant, whatever the precursor pool used, suggesting large differences in their synthetic activities and functions. Use of the plasma [(13)C]KIC pool led to an underestimation of leukocyte FSR compared with the intracellular pool (PBMC: 6.04 +/- 0.94%/day; PMN: 2.98 +/- 0.30%/day). Hence, the intracellular free amino acid pool must be used as precursor to obtain reliable results. In conclusion, it is possible to assess immune cell metabolism in vivo in humans by using small blood samples to directly indicate their metabolic activity in various clinical situations and in response to regulating factors. PMID:14749219

Walrand, Stéphane; Guillet, Christelle; Gachon, Pierre; Rousset, Paulette; Giraudet, Christophe; Vasson, Marie-Paule; Boirie, Yves



Gene Expression Profile of Endotoxin-stimulated Leukocytes of the Term New Born: Control of Cytokine Gene Expression by Interleukin-10  

PubMed Central

Introduction Increasing evidence now supports the association between the fetal inflammatory response syndrome (FIRS) with the pathogenesis of preterm labor, intraventricular hemorrhage and bronchopulmonary dysplasia. Polymorphonuclear leukocyte (PMNs) and mononuclear cell (MONOs) infiltration of the placenta is associated with these disorders. The aim of this study was to reveal cell-specific differences in gene expression and cytokine release in response to endotoxin that would elucidate inflammatory control mechanisms in the newly born. Methods PMNs and MONOs were separately isolated from the same cord blood sample. A genome-wide microarray screened for gene expression and related pathways at 4 h of LPS stimulation (n?=?5). RT-qPCR and ELISA were performed for selected cytokines at 4 h and 18 h of LPS stimulation. Results Compared to PMNs, MONOs had a greater diversity and more robust gene expression that included pro-inflammatory (PI) cytokines, chemokines and growth factors at 4 h. Only MONOs had genes changing expression (all up regulated including interleukin-10) that were clustered in the JAK/STAT pathway. Pre-incubation with IL-10 antibody, for LPS-stimulated MONOs, led to up regulated PI and IL-10 gene expression and release of PI cytokines after 4 h. Discussion The present study suggests a dominant role of MONO gene expression in control of the fetal inflammatory response syndrome at 4 hrs of LPS stimulation. LPS-stimulated MONOs but not PMNs of the newborn have the ability to inhibit PI cytokine gene expression by latent IL-10 release.

Davidson, Dennis; Zaytseva, Alla; Miskolci, Veronika; Castro-Alcaraz, Susana; Vancurova, Ivana; Patel, Hardik



Elevated secretion of reactive nitrogen and oxygen intermediates by inflammatory leukocytes in hyperacute experimental autoimmune encephalomyelitis: enhancement by the soluble products of encephalitogenic T cells  

PubMed Central

Perivascular lesions within the central nervous system (CNS) of rats with hyperacute experimental autoimmune encephalomyelitis (HEAE) contained large numbers of peripheral blood mononuclear cells (PBMC) and polymorphonuclear leukocytes (PMN), cells enzymatically capable of producing reactive nitrogen and oxygen intermediates (RNI and ROI), which, in excess, are mediators of tissue damage. PBMC and PMN isolated from the CNS and periphery of HEAE-affected rats secreted significantly (p less than 0.01-0.0001) elevated levels of ROI and RNI compared with that of similar cell populations from pertussis- and saline-treated control animals. Coincubation of systemically derived PBMC and PMN with antigen-stimulated myelin basic protein-specific T cell lines led to further increases in ROI and RNI output of between 15.3 and 83.1%, an effect that could be largely attributed to heat-labile, soluble products released by these T cell lines. Our studies suggest a putative neuropathological role for ROI and RNI in HEAE, which may be mediated via cytokines emanating from autoreactive T lymphocytes.