Claes Dahlgren

Characteristics of the granulocyte chemiluminescence reaction following an interaction between human neutrophils and Salmonella typhimurium bacteria.

The production of reactive oxygen metabolites by neutrophils is thought to play a key role in the host defence against invading microorganisms. The production of these oxidative metabolites can be measured as chemiluminescence. In this study, two strains of Salmonella typhimurium were used as stimuli, and the opsonin‐independent CL response from neutrophils challenged with these bacteria was investigated. The strains used, S. typimurium 395 MS and a rough (Rd Epi‐2) mutant 395 MR 10, differ with respect to physicochemical surface characteristics. When neutrophils were exposed to the phagocytic prey, only the MR 10 bacteria induced a CL response. The response induced by the MS bacteria was less than 2% of that induced by MR 10. In order to study the relation between intra and extracellularly generated CL, systems were used which selectively inhibit the intra and extracellular CL, respectively. Using these systems it was found that a predominant part of the response was of intracellular origin. When the neutrophils were treated with cytochalasin B (5ug) before the addition of the bacteria, the CL response was reduced to around 37% of the value obtained from untreated cells, and the relation between the extra and the intracellular parts of the response was changed. The mechanism(s) and biological consequences of the extracellular and intracellular generation of oxygen metabolites, respectively, are discussed.

Chemotaxis of polymorphonuclear leukocytes in response to surface-bound complement-derived chemoattractants generated in situ.

Polymorphonuclear leukocytes (PMNLs) were allowed to migrate on slides with fixed yeast particles dotted about on the surface. Locomotion was quantified by counting the number of yeast particles in association with a PMNL. Yeast particles that differed in their ability to consume hemolytic complement differed also in their ability to generate chemoattractants. Addition of a complement source to yeast particles able to activate the complement system resulted in a chemotactic response, as well as when fluid-phase attractants were removed prior to the measurement of PMNL chemotaxis, indicating that the chemoattractants generated were adsorbed to the surface. Using an immunofluorescence technique, it was found that complement factor 5 coated a circular area around each yeast particle, provided that the particles were able to activate the complement system.

Physicochemical surface changes on phagocytic cells during differentiation in relation to chemotaxis and phagocytosis.

During cell differentiation induced by DMSO, the HL-60 cells exhibit certain surface alterations such as increased hydrophobic interaction. Similar changes have been observed in activated alveolar macrophages, and granulocytes exposed to chemoattractant in vivo and in vitro. Whether these surface changes are directly linked to the enhanced chemotactic, oxidative and phagocytic responsiveness is unclear, although increased liability to hydrophobic interaction promotes phagocytic recognition in several systems. From the present data it is evident that oxidative and phagocytic responsiveness are acquired at different stages of differentiation. Detecting physicochemical differences in the surface properties of the HL-60 cells during differentiation may furthermore provide a useful tool for studying subpopulations of cells during differentiation and activation.

Function of exudate neutrophils from skin in psoriasis

Human neutrophils harvested from skin chambers containing autologous serum in psoriatic patients were compared with peripheral blood neutrophils by examining migration, phagocytosis, and oxidative activation. Random migration was reduced in exudate cells, whereas the chemotactic response to yeast-activated serum was evident not only in blood neutrophils, but also in exudate cells. The metabolic activation of exudate neutrophils, when stimulated with formyl-methionyl-leucyl-phenylalanine and measured as chemiluminescence, was enhanced by 50-400% compared to blood neutrophils. The chemiluminescence response to phorbol myristate acetate was on the other hand reduced to 35%. In the phagocytic assay, using C3bi- and IgG-opsonized yeast particles, exudate neutrophils from non-psoriatic healthy controls showed enhanced uptake of C3bi-coated yeast compared to blood neutrophils. In psoriatics, the blood neutrophils showed enhanced C3bi-mediated phagocytosis compared to non-psoriatic control cells. No further increase in C3bi-mediated phagocytosis was then seen in exudate cells from these patients. IgG-mediated phagocytosis was in contrast to C3bi similar between blood and exudate neutrophils in psoriatics and non-psoriatic controls. These experiments show that during exudation priming of different receptor-mediated processes can occur. However, no significant difference was observed between different functional capacities in exudate neutrophils from psoriasis patients and non-psoriatic controls.

Superoxide production and chemiluminescence induced in differentiated HL-60 cells by the chemoattractant formyl-methionyl-leucyl-phenylalanine

Superoxide production and chemiluminescence induced in differentiated HL-60 cells by the chemoattractant formylmethionyl-leucyl-phenylalanine: In order to study the generation of oxidative metabolites in relation to cell differentiation, dimethyl sulfoxide (DMSO) and retinoic acid (RA) differentiated HL-60 cells were stimulated with the chemotactic peptide formylmethionyl-leucyl-phenylalanine (FMLP). The oxidative response was measured as luminol-dependent chemiluminescence, lucigenin-dependent chemiluminescence, and cytochrome c reduction. Cells grown in the presence of DMSO or RA progressively expressed morphological changes, and when the mature cells were exposed to FMLP the cells produced oxidative metabolites. Quantitatively the HL-60 cells grown in the presence of DMSO gave rise to the most pronounced response. No correlation was obtained between superoxide production, luminol-chemiluminescence and lucigenin-dependent chemiluminescence, indicating that different aspects of the oxidative response are elucidated by the three different methods. Furthermore, the experiments show that DMSO and RA-induced differentiation of HL-60 cells leads to granulocyte-like cells with different abilities to produce oxidative metabolites, possibly due to differences in receptor function.

Modulation of Granulocyte Response to the Chemoattractant F-Met-Leu-Phe

Cells involved in host defense, including macrophages and granulocytes, change their surface and functional properties during maturation (1,2,3). In addition, mature granulocytes are able to change their surface structure and functional characteristics in response to mediators of the inflammatory process (4,5,6,7).