Chemiluminescence Response Of Polymorphonuclear Leukocytes Research Articles

The effect of low levels of dietary cobalt on the chemiluminescence response of polymorphonuclear leukocytes of goats

Twenty ten-week-old newly weaned male Batinah goats were randomly assigned to a control ( n = 10) and a treated ( n = 10) group and were fed a diet containing 0.1 mg/kg DM cobalt (Co). Goats in the treated group received bi-monthly subcutaneous injections of 2000 μg of hydroxycobalamin. The phagocytic function of the polymorphonuclear leukocytes (PMN) were tested using a luminol-dependent chemiluminescence assay with opsonized zymosan as the phagocytic target. One month after the onset of the experiment PMN from the control group exhibited a significantly ( p < 0.05) lower CL response, which continued for the second month. The results of the present study demonstrated that low levels of dietary cobalt leads to an early impairment of phagocytic function. This may at least in part, be an explanation as to why at the field level in Oman young goats fed diets containing low levels of Co appear to be more susceptible to infections.

Antioxidant properties of alpha-crystallin.

alpha-Crystallin is a major chaperone lens protein to which has been ascribed antioxidant functions. In the present work we have evaluated the antioxidant and free radical scavenging properties of bovine alpha-crystallin in a series of in vitro models: zimosan-induced, luminol-enhanced chemiluminescence response of polymorphonuclear leukocytes, the autoxidation of brain homogenate, bleaching of 2,2'-azinobis(3-ethylbenzothiazoline-6-sulfonic acid)-derived radical cations, trapping of peroxyl radicals, and reactivity toward hypochloric acid. In all these systems, the reactivity of alpha-crystallin is higher than or similar to that of bovine serum albumin. It is concluded that, given the high concentrations of ol-crystallin in the lenses, its capacity to interact with free radicals and to remove hypochlorous acid could contribute to the maintenance of the lens functionality.

Immunoglobulin enhances the bioactive-glass-induced chemiluminescence response of human polymorphonuclear leukocytes.

Bioactive glasses are bone substitutes that chemically bind to bone. Implanted materials always elicit a response from surrounding tissues and thereby can activate inflammatory cells, with subsequent release of biomaterial and tissue-damaging agents. Bioactive glasses can activate polymorphonuclear leukocytes (PMNL) and induce a release of reactive oxygen metabolites (ROM). Adsorption of proteins on the surface of the implanted material may influence the subsequent inflammatory cell response. The effect of Sandoglobulin(R) (SG) and albumin on the ROM release by PMNLs induced by a bioactive glass was studied by a chemiluminescence (CL) assay. An enhanced effect for SG and inhibitory effect for albumin on the CL response of PMNLs was observable. The CL response of the PMNLs was dependent on the incubation time of the glass in solution.

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

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.

Characterization of the chemiluminescence response of polymorphonuclear leukocytes stimulated with urate crystals.

We studied the luminol-amplified chemiluminescence (CL) response of human polymorphonuclear leukocytes (PMNLs) stimulated with synthetic monosodium urate monohydrate (MSUM) crystals. Stimulation of PMNLs prepared from healthy controls with MSUM resulted in a bimodal CL response with two peaks after 1 min and 4-6 min, respectively. MSUM caused neutrophils to produce reactive oxygen species (ROS), which are released from the cells as well as ROS with an intracellular localization. Proportionally more of the CL activity generated during the first peak was of extracellular origin. The MSUM-induced light-generating mechanism was azide-sensitive, required superoxide anion and was strongly suppressed by horseradish peroxidase.

Characterization of the Chemiluminescence Response of Polymorphonuclear Leukocytes Stimulated with Urate Crystals

We studied the luminol-amplified chemiluminescence (CL) response of human polymorphonuclear leukocytes (PMNLs) stimulated with synthetic monosodium urate monohydrate (MSUM) crystals. Stimulation of PMNLs prepared from healthy controls with MSUM resulted in a bimodal CL response with two peaks after 1 min and 4–6 min, respectively. MSUM caused neutrophils to produce reactive oxygen species (ROS), which are released from the cells as well as ROS with an intracellular localization. Proportionally more of the CL activity generated during the first peak was of extracellular origin. The MSUM-induced light-generating mechanism was azide-sensitive, required superoxide anion and was strongly suppressed by horseradish peroxidase.

Construction and characterization of arginine-specific cysteine proteinase (Arg-gingipain)-deficient mutants of Porphyromonas gingivalis. Evidence for significant contribution of Arg-gingipain to virulence.

Arginine-specific cysteine proteinase (Arg-gingipain; formerly, argingipain) is one of the major extracellular proteinases produced by the oral anaerobic bacterium Porphyromonas gingivalis. To determine whether Arg-gingipain is important for periodontopathogenicity of the organism, Arg-gingipain-deficient mutants were constructed via gene disruption by use of suicide plasmid systems. First, Southern hybridization analyses suggested that two separate Arg-gingipain-encoding genes designated rgpA and rgpB existed on 12.5- and 7.8-kilobase pair HindIII chromosomal fragments of P. gingivalis ATCC33277, respectively. rgpA and rgpB single mutants were constructed by mobilization of a suicide plasmid. Then, an rgpA rgpB double mutant was isolated by electroporation with a second suicide plasmid. No proteolytic activity for Arg-gingipain was observed in either the cell extract or the culture supernatant of the rgpA rgpB mutant. The chemiluminescence response of polymorphonuclear leukocytes, which is closely related to their bactericidal function, was not inhibited by the culture supernatant of the rgpA rgpA mutant, while the wild type parent showed a significant inhibition of the response. The result suggests that Arg-gingipain is responsible for disruption of the function of polymorphonuclear leukocytes. In addition, the rgpA rgpB double mutations caused a marked decrease in the hemagglutination of P. gingivalis, indicating that a major part of the hemagglutinin activity of the organism is associated with the two genes. These findings demonstrate that Arg-gingipain makes a significant contribution to the virulence of P. gingivalis.

Effect of priming polymorphonuclear leukocytes with cytokines (granulocyte-macrophage colony-stimulating factor [GM-CSF] and G-CSF) on the host resistance to Streptococcus pneumoniae in chinchillas infected with influenza A virus

Patients infected with influenza A virus (IAV) are at increased risk for bacterial superinfections, and this occurs in association with depressed polymorphonuclear leukocyte (PMNL) function. Recently, we reported that in vitro exposure of human PMNL to granulocyte-macrophage colony-stimulating factor (GM-CSF) reverses IAV-induced cell dysfunction. The present study used an established animal model of IAV infection to examine whether G-CSF and/or GM-CSF can overcome IAV- induced PMNL dysfunction and thereby prevent secondary infections. Preliminary studies determined a dosing schedule of these cytokines that caused significant priming of chinchilla PMNL. In subsequent studies, animals were inoculated intranasally with IAV (day 1) followed 3 days later by Streptococcus pneumoniae, and administered daily intraperitoneal injections with a cytokine or placebo on days 3 through 9. Animals had blood obtained on multiple occasions for PMNL studies, and were followed-up for evidence of pneumococcal disease. Both cytokines caused significant priming of the PMNL chemiluminescence response and this was associated with reversal of the IAV-induced PMNL dysfunction. However, neither cytokine decreased the incidence of pneumococcal disease.

Effect of Priming Polymorphonuclear Leukocytes With Cytokines (Granulocyte-Macrophage Colony-Stimulating Factor [GM-CSF] and G-CSF) on the Host Resistance to Streptococcus pneumoniae in Chinchillas Infected With Influenza A Virus

AbstractPatients infected with influenza A virus (IAV) are at increased risk for bacterial superinfections, and this occurs in association with depressed polymorphonuclear leukocyte (PMNL) function. Recently, we reported that in vitro exposure of human PMNL to granulocyte-macrophage colony-stimulating factor (GM-CSF) reverses IAV-induced cell dysfunction. The present study used an established animal model of IAV infection to examine whether G-CSF and/or GM-CSF can overcome IAV-induced PMNL dysfunction and thereby prevent secondary infections. Preliminary studies determined a dosing schedule of these cytokines that caused significant priming of chinchilla PMNL. In subsequent studies, animals were inoculated intranasally with IAV (day 1) followed 3 days later by Streptococcus pneumoniae, and administered daily intraperitoneal injections with a cytokine or placebo on days 3 through 9. Animals had blood obtained on multiple occasions for PMNL studies, and were followed-up for evidence of pneumococcal disease. Both cytokines caused significant priming of the PMNL chemiluminescence response and this was associated with reversal of the IAV-induced PMNL dysfunction. However, neither cytokine decreased the incidence of pneumococcal disease.

The Ability of Polymorphonuclear Leukocyte Priming Agents to Overcome Influenza A Virus-Induced Cell Dysfunction

The major mortality and morbidity resulting from influenza virus infections are due to secondary bacterial infections which occur in association with virus-induced inhibition of polymorphonuclear leukocyte (PMNL) function. The present study was undertaken to determine if compounds which prime PMNL function to subsequent stimulation with N-formylmethionyl-leucylphenylalanine (FMLP) or phorbol 12-myristate 13-acetate (PMA) can overcome influenza A virus (IAV)-induced inhibition of the PMNL chemiluminescence response to these stimuli. Granulocyte-macrophage colony stimulating factor (GM-CSF), guanosine triphosphate (GTP), and 1-oleoyl-2-acetylglycerol (OAG) were able to prime the PMNL response to FMLP and/or PMA and totally or partially overcome IAV-induced PMNL dysfunction in cells stimulated with FMLP or PMA. A direct correlation was found between the extent of PMNL priming due to GM-CSF, GTP, and OAG and the capacity of these compounds to overcome virus-induced PMNL dysfunction. The implications of these findings in regard to the mechanism by which priming agents overcome IAV-induced cell dysfunction and the potential of these compounds as therapeutic agents to treat secondary bacterial infections are discussed.

Phagocytosis of mastitis isolates of Staphylococcus aureus and expression of type 5 capsular polysaccharide are influenced by growth in the presence of milk

Phagocytosis by bovine polymorphonuclear granulocytes of seven capsular polysaccharide type 5 Staphylococcus aureus strains isolated from mastitis [corrected] was investigated by means of luminol-dependent chemiluminescence. Bacteria were grown on four different agar media (brain heart infusion, Columbia broth, modified staphylococcus medium 110, and skim milk) and were opsonized by normal bovine serum. When compared to growth on brain heart infusion agar, Columbia agar, and modified staphylococcus medium 110 agar, growth on skim milk agar rendered five of the strains more resistant to opsonization. The other two strains were resistant in all culture media used. Short periods of incubation in milk after growth on brain heart infusion agar did not augment resistance to phagocytosis, indicating that mere adsorption of milk components on bacteria was not responsible. The variability of the chemiluminescence response of polymorphonuclear leukocytes was pronounced among strains with each growth medium except milk. Growth on modified staphylococcus medium 110 and on milk agar favored the masking of teichoic acid, as shown by inagglutinability with rabbit antiserum. Interestingly, agglutination by a monoclonal antibody to capsular polysaccharide type 5 was optimal when bacteria were grown on skim milk agar. This suggests that capsular polysaccharide participated in the masking effect. These findings indicate that masking of the bacterial target of most of the naturally acquired opsonins present in normal bovine serum occurred when bacteria grew in the presence of milk, resulting in an increased resistance to phagocytosis by polymorphonuclear leukocytes.

Polymorphonuclear leukocyte chemiluminescence response in HLA-B27 positive and negative arthritic patients.

The polymorphonuclear leukocyte (PMN) chemiluminescence (CL) response to opsonized zymosan reflecting the C3b/C3bi-receptor-mediated PMN respiratory burst activation was studied in 99 arthritic patients typed for the HLA-B27 antigen. The results indicate that the B27 antigen per se is not associated with disturbances in PMN CL response. However, the erythrocyte sedimentation rate showed a correlation with CL, suggesting that the activity of arthritic disease may affect the PMN CL response.

Lipopolysaccharide-dependent enhancement of adherence-mediated chemiluminescence response of polymorphonuclear leukocytes.

Adherence of resting polymorphonuclear leukocytes to nylon fibre increased the chemiluminescence response (CL) from 99,400 to 910,300 cpm/25,000 PMNL. This effect could be amplified by lipopolysaccharide (LPS) priming of granulocytes in a dose-dependent fashion. The results of nylon fibre adherence experiments suggest an in vitro model that might approximate certain conditions of in vivo PMNL-endothelial adherence and respiratory burst activation, and these reactions of polymorphonuclear leukocytes may contribute to the pathomechanisms of the Adult Respiratory Distress Syndrome.

Inhibition of human neutrophil chemiluminescence by plasmid-mediated outer membrane proteins of Yersinia enterocolitica.

Recent studies have shown that the cell surface properties of Yersinia enterocolitica are altered by the presence of the virulence plasmid, which mediates temperature-inducible outer membrane proteins (OMP). We investigated the interaction of Y. enterocolitica with human polymorphonuclear leukocytes by monitoring luminol-enhanced chemiluminescence (CL) responses. A plasmid-bearing strain grown at 37 degrees C induced four- to sixfold less CL than did the same strain grown at 25 degrees C or a plasmidless, isogenic strain grown at either temperature. Inhibition of CL responses by whole cells was related to plasmid-mediated expression of OMP. The OMP alone could inhibit the CL response of polymorphonuclear leukocytes stimulated by either opsonized zymosan or whole cells of Y. enterocolitica. Pronase treatment of whole cells, which removed the plasmid-mediated OMP, resulted in partial but significant elimination of CL inhibition by whole cells and by OMP derived from them. Incubation with Y. enterocolitica for 60 min did not affect the viability of polymorphonuclear leukocytes. Our results suggest that the interaction of Y. enterocolitica with human polymorphonuclear leukocytes is directly affected by the plasmid-mediated OMP.

Comparison between the luminol-dependent chemiluminescence of polymorphonuclear leukocytes and of the myeloperoxidase-HOOH system: influence of pH, cations and protein.

Abstract— Measurements of luminol‐dependent chemiluminescence (CL) from normal human polymorphonuclear leukocytes (PMNL) were carried out and the results were compared to the CL obtained from a cell‐free myeloperoxidase(MPO)‐HOOH‐system.The chemotactic peptide formylmethionyl‐leucyl‐phenyl‐alanine (fMLP) induced a bimodal luminol‐dependent CL response in PMNL, while phorbol myristate acetate (PMA) induced a one‐peak response. Calcium was found to be required for the fMLP but not for the PMA induced response. The absence of glucose reduced the PMA but not the fMLP induced PMNL CL response. Neither calcium nor glucose had any effect on the light emitted from a cell‐free MPO‐HOOH‐system.The PMNL CL response to both fMLP and PMA was affected by the pH‐value, in that at a low pH (6.3) the response was decreased and at a high pH (8.0) the response was increased. These effects were also obtained in cell‐free MPO‐HOOH‐systems. The presence of human serum albumin (HSA) totally inhibited the CL of a cell‐free MPO‐HOOH‐system while, in PMNL CL, HSA reduced only the first phase of the fMLP induced response.The results are discussed in view of the fact that CL is increasingly used as a clinical instrument to study defects in PMNL function and the effects on PMNL function of various agents such as drugs.

Interaction between human polymorphonuclear leukocytes and Chlamydia trachomatis

The association of Chlamydia trachomatis serotypes E and L1 with human polymorphonuclear leukocytes (PMNL) and the ability of these chlamydiae to induce a chemiluminescence response in PMNL were recorded. C. trachomatis unequivocally activated a respiratory burst in PMNL. Despite similar association with PMNL, serotype E activated the PMNL significantly more than serotype L1. Type-specific, but not group-specific antisera increased the chemiluminescence but not association of chlamydiae with PMNL. These results indicate that recording of chemiluminescence may be of value to estimate potential antichlamydial activity of PMNL and that type-specific antibodies have opsonizing effect exerted subsequent to the attachment phase of chlamydiae to PMNL.

Interaction between human polymorphonuclear leukocytes andChlamydia trachomatis

The association of Chlamydia trachomatis serotypes E and L1 with human polymorphonuclear leukocytes (PMNL) and the ability of these chlamydiae to induce a chemiluminescence response in PMNL were recorded. C. trachomatis unequivocally activated a respiratory burst in PMNL. Despite similar association with PMNL, serotype E activated the PMNL significantly more than serotype L1. Type-specific, but not group-specific antisera increased the chemiluminescence but not association of chlamydiae with PMNL. These results indicate that recording of chemiluminescence may be of value to estimate potential antichlamydial activity of PMNL and that type-specific antibodies have opsonizing effect exerted subsequent to the attachment phase of chlamydiae to PMNL.

Effect of in vitro preincubation of polymorphonuclear leukocytes on formylmethionyl-leucyl-phenylalanine-induced chemiluminescence.

When polymorphonuclear leukocytes (PMNL) and soluble or particulate matter interact, the cells produce chemiluminescence. The chemotactic peptide formylmethionyl-leucyl-phenylalanine induced a two-peak chemiluminescence response in PMNL. The response was modified, both in magnitude and in the time course of the response, when PMNL were incubated at 22 degrees C for 15 to 120 min before the addition of stimulus. The cellular response to formylmethionyl-leucyl-phenylalanine was also changed by the addition of cell-free supernatants from stored PMNL suspensions to freshly prepared PMNL before the addition of formylmethionyl-leucyl-phenylalanine. This indicated that PMNL during storage released substances that changed the chemiluminescence response. A similar change could be obtained by addition of purified myeloperoxidase to freshly prepared PMNL before the addition of stimulus, indicating that the myeloperoxidase-hydrogen peroxide system can function as a modulator of the cellular response to chemotactic factors.