Biomedical subjects

M E van der Tol

Publications and source records attributed to M E van der Tol.

12 recordsLinked to original sources

PubMed1998

A synthetic lipopolysaccharide-binding peptide based on amino acids 27-39 of serum amyloid P component inhibits lipopolysaccharide-induced responses in human blood.

LPS-binding proteins in plasma play an important role in modifying LPS toxicity. Significant properties have already been attributed to the LPS-binding protein (LBP). It accelerates LPS toxicity as well as incorporation into high-density lipoproteins, leading to neutralization of LPS in serum. A search for other LPS-binding components in serum, using LPS-coated magnetic beads, revealed a new LPS-binding protein. N-terminal microsequencing identified this protein as serum amyloid P component (SAP). Purified SAP bound to smooth and rough types of LPS via the lipid A part. SAP inhibited the binding of FITC-labeled ReLPS (LPS from Salmonella minnesota strain R595) to human monocytes and the ReLPS-induced priming of the oxidative burst of human neutrophils only in the presence of low concentrations of LBP. In search for the LPS binding site of SAP, we found that pep27-39, a 13-mer peptide consisting of amino acids 27-39 of SAP, competitively inhibited the binding of LPS to SAP. In addition, pep27-39 significantly inhibited ReLPS-induced responses in phagocytes in the presence of serum, as well as in human whole blood. Carboxamidomethylated pep27-39 showed an even more pronounced reduction of the ReLPS-induced priming of phagocytes in human blood. Performing gel filtration of FITC-labeled ReLPS incubated with soluble CD14, we showed that SAP could not prevent binding of LPS to soluble CD14, in contrast to pep27-39. The ability of pep27-39 to antagonize specifically the effects of LPS in the complex environment of human blood suggests that pep27-39 may be a novel therapeutic agent in the treatment of gram-negative sepsis.

Acute-Phase Proteins↗

PubMed1991

Tumour necrosis factor triggers granulocytes to internalize complement-coated virus particles.

Monocytes produced tumour necrosis factor-alpha (TNF-alpha) upon interaction with infective herpes simplex virus (HSV). Therefore, TNF-alpha and its action were examined in the regulation of anti-viral functions of human polymorphonuclear leucocytes (PMN). The uptake of fluorescein-labelled HSV by human PMN was monitored using flow cytometric analysis. As shown in earlier work, complement-coated herpes virions were bound to PMN but were not internalized. However, after priming with recombinant human TNF-alpha, complement-coated virions were taken up by human PMN. This complement receptor-mediated internalization is a new observation, both because it affects PMN and it is induced by a recombinant cytokine. Recombinant human granulocyte-macrophage colony-stimulating factor (GM-CSF), another priming factor of PMN, was unable to induce this phenomenon. By contrast, other parameters of PMN anti-viral defence were enhanced to the same extent by both TNF-alpha and GM-CSF. PMN primed by either TNF-alpha or GM-CSF showed both an enhanced respiratory burst and an increased membrane potential depolarization when triggered by immune complexes containing HSV, antibody and complement. Both primers rendered an enhanced uptake of HSV in the presence of specific antibody and in the presence of both antibody and complement, but they caused no effect on the rate or quantity of processing of phagocytosed HSV particles by PMN. We propose that TNF-alpha and GM-CSF act as effective modulators of PMH to enhance virus removal, especially in inflammatory sites. We tentatively conclude that TNF-alpha together with PMN and complement represent a novel non-specific defence mechanism against HSV.

Complement System Proteins↗

PubMed1990

Degradation of herpes simplex virions by human polymorphonuclear leukocytes and monocytes.

The degradation of herpes simplex virus particles after uptake by phagocytes was studied, but, since lysis of the phagocyte also resulted in damage to the viral envelope, measurement of viral infectivity as a criterion of viral degradation after phagocytosis was not possible. Therefore we focused on later events in viral destruction, namely the degradation of macromolecules. We have demonstrated that polymorphonuclear leukocytes (PMN) and monocytes (MN) can rapidly degrade the membrane proteins of the phagocytosed herpes-virus virions. PMN and MN from a patient with chronic granulomatous disease showed a similar rate of degradation compared to PMN and MN from healthy donors, which excludes an important role for toxic oxygen species in viral protein degradation. Experiments using toxic oxygen species-generating systems supported this observation. In contrast to PMN, MN are also effective in the digestion of viral DNA. We conclude that PMN and MN are able to neutralize large amounts of phagocytosed HSV, so their role in antiviral defence has again been demonstrated.

DNA, Viral↗

PubMed1989

Quantitation of conjugate formation between human polymorphonuclear leukocytes and antibody-coated target cells by flow cytometry: the role of Fc receptor and LFA-1 antigen.

The specific binding of human polymorphonuclear leukocytes (PMN) to antibody-coated target cells was characterized by flow cytometry. PMN were labeled with phycoerythrin-E (PE) via a granulocyte-specific monoclonal antibody (leu-M1) and mixed with fluorescein isothiocyanate-labeled K562 tumor cells sensitized with rabbit antiserum. Specific conjugates were formed as analyzed by two-color fluorescence in a flow cytometer. The formation of stable conjugates was dependent on initiation of contact, temperature, time, and antiserum concentration. Studies with inhibitors implicate that microfilaments, but not microtubules, Ca2+, Mg2+, or energy-dependent processes were a prerequisite for binding of PMN to the antibody-coated target cells. No conjugates were formed when uncoated target cells were used or when the experiment was performed in the presence of protein A, indicating that binding was specifically mediated through Fc receptors (FcR). Monoclonal antibodies against the FcRII and FcRIII were used to address the role of these receptors in conjugation. One of the two anti-FcRIII antibodies and an anti-FcRII antibody effectively prevented conjugation. A monoclonal antibody directed against the common beta-chain of the adhesion molecule family and a combination of antibodies against the alpha-chain of LFA-1 and Mo-1 also blocked conjugation when target cells were sensitized under suboptimal conditions. The antibody against the beta-chain also diminished killing of antibody-coated K562, as measured by chromium release when included in the cytotoxicity assay. These results indicate that flow cytometry permits accurate quantitation and characterization of the binding between PMN and antibody-coated target cells, which in principle, can be prevented by monoclonal antibodies against surface receptors. Binding is primarily established by both the FcRII and FcRIII. Adhesion-associated molecules on the PMN surface contribute to optimal binding.

Antibodies, Monoclonal↗

PubMed1989

Phagocytosis of herpes simplex virus by human granulocytes and monocytes.

Polymorphonuclear leukocytes (PMN) can mediate cytotoxic reactions against virus infected targets cells. We observed very efficient binding of PMN to HSV-infected fibroblasts when loaded with HSV-specific antibodies. Using electron microscopy, infected fibroblasts were found to be totally surrounded by PMN and the phagocytosis of virions and fragments of infected cells was demonstrated. To quantify and study this phenomenon, and to compare PMN with monocytes, we developed radiometric and fluorometric phagocytosis assays. Leukocytes were mixed with [3H]glucosamine- or FITC-labeled virus and incubated at 37 degrees C. PMN associated radioactivity or fluorescence per cell as measured by flow cytometry was determined. PMN phagocytosis was dependent on the presence of specific anti-HSV antibodies and could be enhanced by addition of complement. Monocytes were also able to phagocytize virions; however, the rate of uptake was less than that for PMN. Under optimal conditions the total amount of herpes simplex particles that could be associated with one PMN or monocyte was about 10,000. PMN and monocytes are capable of phagocytosis of HSV. This may be an important factor in preventing the spread of infection in vivo.

Antibodies, Viral↗

PubMed1989

Complement-mediated phagocytosis of herpes simplex virus by granulocytes. Binding or ingestion.

The role of complement receptors in phagocytosis of herpes simplex virus (HSV) by PMN was examined. Complement components were deposited on the surface of the virus particle in the presence or absence of specific anti-HSV antibodies. Flow cytometry was used to analyze the phagocytosis of fluorescence-labeled viruses and demonstrated that although a virion is able to associate with PMN in the presence of complement alone, the granulocyte is not triggered to mount a metabolic burst. Efficient stimulation of PMN occurs when complexes are formed consisting of virus, specific antibodies, and complement. To address the question whether the viruses were inside or outside the cell, a combined enhancement/quenching method was developed using ammonium chloride as a lysosomotropic agent and trypan blue as a quenching dye. The data indicate that Fc receptor-mediated phagocytosis by PMN results in the ingestion of all cell-associated herpes virions. Interactions of virions through PMN-complement receptors CR1 and CR3 results solely in binding to the PMN but not in internalization. Interactions via both complement and Fc receptors cause synergistic stimulation of the PMN and result in very efficient association of viruses, greater than 80% of which were inside the cell.

Complement System Proteins↗

PubMed1986

Human polymorphonuclear leukocytes release leukotriene B4 during phagocytosis of Staphylococcus aureus.

We studied release of leukotriene B4 (LTB4) by human polymorphonuclear leukocytes (PMNs) during phagocytosis of staphylococci in the presence or absence of arachidonic acid. The 12 X 10(7) PMNs incubated with 3 X 10(9) opsonized S. aureus and 50 microM arachidonic acid released 1.45 +/- 0.42 nmol LTB4. No LTB4 was detected after stimulation of PMNs with S. aureus or arachidonic acid by themselves. However, by increasing the concentration of arachidonic acid to 200 or 400 microM, 1.22 +/- 0.45 and 1.98 +/- 0.49 nmol LTB4, respectively, was released by PMNs. The effect of different bacteria-PMN ratios on LTB4 production was also studied. LTB4 varied from 0.3 to 2.0 nmol when bacteria/PMN ratios increased from 5 to 50 (respectively) in the presence of 50 microM arachidonic acid. Thus, phagocytizing PMNs produce LTB4 in the presence of arachidonic acid, and its production is dependent on the number of bacteria phagocytized.

Arachidonic Acids↗

PubMed1985

Interactions between human polymorphonuclear leukocytes and influenza virus.

The effects of influenza virus A (H3N2) on several functions of human polymorphonuclear leukocytes (PMN) were examined. Incubation of PMN with virus induced chemiluminescence, aggregation, and degranulation of the leukocytes. The amount of chemiluminescence generated increased from 1 X 10(6) to 6 X 10(6) cpm when 2.5 X 10(6) to 2 X 10(7) virus particles were added to 2.5 X 10(6) PMN. Maximal aggregation occurred within 2 min and the response depended on the amount of virus added to the PMN. Release of acid phosphatase by virus-treated PMN was 62 +/- 12% within 1 h compared with 7 +/- 7% by control PMN (P less than 0.005). Incubation of PMN with influenza virus resulted in a diminished phagocytic activity of the phagocytes. PMN from a patient with chronic granulomatous disease were similarly affected. It was thus concluded that the observed defect in phagocytic activity was not due to the reactive oxygen species generated by the PMN during incubation with virus.

Cell Aggregation↗

PubMed1984

Aggregation of human polymorphonuclear leucocytes during phagocytosis of bacteria.

The process of aggregation of human polymorphonuclear leucocytes (PMN) during the uptake of bacteria was studied. Radiolabelled S. aureus were opsonized in different sera, washed, resuspended in buffer and added to the PMN. Uptake of the bacteria and aggregation of the PMN were measured simultaneously. Maximal aggregation occurred within 6 min, when 5 X 10(6) PMN had phagocytosed 2.5 X 10(8) S. aureus. Also the effects of serum concentrations and different sera for opsonization of the bacteria on PMN aggregation were studied. Despite normal uptake, aggregation of PMN was low when bacteria were opsonized in complement-deficient sera. Furthermore when PMN were treated with pronase to inactivate complement receptors on the cell surface of the PMN, and bacteria preopsonized in immune serum were added, no change in uptake occurred, although the degree of aggregation halved compared to control PMN. So, interaction between the bacteria and the complement receptor of the PMN cell membrane is needed for triggering the process of aggregation. By using dansylcadaverin and diphenylamine to modulate lysosomal enzyme release, azide or PMN from a chronic granulomatous disease patient to study the effect of the formation of oxygen species, and theophylline, DB-cAMP or 8 Br-cAMP to increase cAMP levels, it was concluded that aggregation of PMN during phagocytosis was not dependent on oxygen metabolism, degranulation or cAMP levels of PMN.

Adult↗

PubMed1983

Escherichia coli lipopolysaccharides diminish and enhance cell function of human polymorphonuclear leukocytes.

The effects of the lipopolysaccharide (LPS) of Escherichia coli J5 and 0111B4 on the function of human polymorphonuclear leukocytes (PMN) were tested. E. coli J5 is a UDP-galactose-4-epimerase-deficient mutant of E. coli 0111B4, and its LPS, therefore, contains mainly lipid A, as it lacks the polysaccharide side chains. PMN which had been incubated with J5 LPS showed decreased phagocytic, chemotactic, and metabolic activities as compared with control PMN. In contrast, incubation of PMN with 0111B4 LPS had no effect or even an enhancing effect on PMN function. When lipid A and the polysaccharide fraction were isolated from 0111B4 LPS, it was shown that lipid A had the same deleterious effect on PMN function as did J5 LPS and that the LPS fraction had no effect. When PMN were incubated with J5 LPS or lipid A, it could be shown that these structures were able to induce PMN to generate superoxide and chemiluminescence. 0111B4 LPS and the polysaccharide component were able to generate a metabolic burst by the PMN to a lesser extent. The induced defects in PMN function by J5 LPS could be prevented when polymyxin B or an oxygen-radical scavenger was present. We hypothesize that the lipid A portion of LPS is toxic for PMN due to the induction of toxic oxygen species by the PMN. These toxic oxygen species destroy the phagocytic, chemotactic, and metabolic activities of the PMN.

Chemotaxis, Leukocyte↗

PubMed1980

Interactions of phagocytic and bacterial cells in patients with bacteremia caused by gram-negative rods.

The phagocytic and bactericidal functions of polymorphonuclear leukocytes and monocytes and the opsonic activity of serum from patients with gram-negative bacteremia were compared with those of cells and serum from healthy donors and control patients. Leukocytes from five of 20 patients showed diminished phagocytic capacity. Leukocytes from three of 12 patients had decreased chemotactic activity. Eleven of 37 blood culture isolates were inefficiently phagocytized after opsonization in homologous patient serum. However, in no instance was the opsonic capacity of patient serum significantly lower than that of control serum. In the serum of some patients, an increase in heat-stable opsonins was found during the course of infection. Resistance to opsonization of strains of Escherichia coli correlated with the presence of K capsular polysaccharide. It was concluded that both impaired leukocyte function and ineffective opsonization play a role in the pathogenesis of gram-negative bacteremia. Heat-stable opsonins (presumably specific antibodies) appear to be necessary for effective phagocytosis of bacilli that cause gram-negative bacteremia.

Antigens, Bacterial↗

PubMed1979

Role of Escherichia coli K capsular antigens during complement activation, C3 fixation, and opsonization.

Escherichia coli strains with K capsular polysaccharides are relatively resistant to phagocytosis by polymorphonuclear leukocytes, in contrast to E. coli strains without K antigens. This inhibition of phagocytosis is related to an impaired recognition of the K+ strains by the phagocytes due to ineffective opsonization. All five strains without K antigens were readily phagocytized after opsonization in 5% normal serum, compared with no uptake of the K+ strains. Evidence is presented that the decreased opsonization of the K+ strains in normal serum is caused by a low rate of complement activation of the strains, with subsequent absence of C3b fixation or C3d fixation or both to the cell wall of the bacteria. After removal of the K+ antigens by heating of a K+ E. coli strain, the strain was able to activate complement, to bind C3b or C3d or both, and to become opsonized. Complement was then activated via the classical and alternative pathways, which was comparable to the complement consumption by K- E. coli.

Antigens, Bacterial↗