leukotriene-b4 and Immunologic-Deficiency-Syndromes

Polymorphonuclear granulocytes (PMN) play a key role in host defense against microbial infections. After severe trauma PMN show cellular dysfunctions including chemotactic migration, phagocytosis, and bacterial killing. In these settings the contribution of the cellular maturation stage compared to functional activities has not been investigated. Polymorphonuclear granulocytes are potent producers of lipid mediators via the 5-lipoxygenase (5-LO) pathway (leukotrienes, LTs) which exert important proinflammatory and immunoregulatory activities. We analyzed leukotriene generation from PMN-fractions (N = 23) of 15 polytrauma patients in comparison to 17 healthy donor cell fractions and correlated this lipid mediator release to the hematopoietic maturation stage of respective PMN. Polymorphonuclear granulocytes were isolated from EDTA-anticoagulated peripheral blood employing a one step procedure based on a discontinuous double Ficoll-gradient. Cells (5 x 10(6)/500 microl phosphate-buffered saline) were stimulated for 20 min at 37 degrees C with 1 microM Ca-ionophor A23187 in the presence of 1 mM Ca++ and 0.5 mM Mg++. Leukotrienes were analyzed by reversed-phase HPLC. Expression of 5-lipoxygenase (5-LO) was additionally determined by Western blot. Maturation stage of PMN was quantitated by Pappenheim-staining of cell smears. After polytrauma the generation of leukotrienes from PMN was individually diminished. Synthesis of enzymatically formed metabolites (LTB4, OH-LTB4 and COOH-LTB4) was concomitantly reduced. The decresaed leukotriene synthesis strongly correlated (r2 = 0.907, P < 0.0001) to the occurrence of immature PMN (mostly band cells). The expression of 5-lipoxygenase in PMN fractions consisting mainly of band cells was decreased. Our results provide evidence that posttraumatic granulocyte dysfunction is partly due to immature functional cell capacities.

Topics: Adolescent; Adult; Arachidonate 5-Lipoxygenase; Blotting, Western; Calcimycin; Calcium Signaling; Cell Differentiation; Cellular Senescence; Chromatography, High Pressure Liquid; Enzyme Induction; Female; Humans; Immunologic Deficiency Syndromes; Inflammation Mediators; Ionophores; Leukotriene B4; Leukotrienes; Male; Middle Aged; Multiple Trauma; Neutrophils

We have identified a patient with a number of neutrophil dysfunctions. The patient was a female baby who lived for 8 months. During her life, she developed severe bacterial infections and showed omphalitis, impaired wound healing, and a pronounced leukocytosis. She was not a patient with leukocyte adhesion deficiency, because all leukocyte CD18 complex proteins were expressed at normal levels. Yet, neutrophil polarization and chemotaxis to platelet-activating factor, leukotriene B4, or formyl-methionyl-leucyl-phenylalanine (FMLP) were completely absent. We found a strong defect in actin polymerization in response to chemotactic stimuli, but only a retarded or even normal reaction with other stimuli. This indicates that the cellular dysfunctions were not due to an intrinsic defect in actin metabolism. Instead, the regulation of actin polymerization with chemotactic stimuli seemed to be defective. We concentrated on FMLP-induced responses in the patient's neutrophils. Functions dependent on activation of complement receptor type 3, such as aggregation or adherence to endothelial cells, were normally induced. Binding to serum-coated coverslips was normal in cell number; however, spreading was not observed. Exocytosis from the specific granules was readily induced. In contrast, FMLP failed to induce a respiratory burst activity or degranulation of the azurophil granules. FMLP induced a normal increase in free intracellular Ca2+, but a decreased formation of diglycerides (especially the 1-O-alkyl,2-acyl compounds). Thus, we have described a patient whose neutrophils show a severe defect in functional activation via chemotaxin receptors, resulting in a selective absence of NADPH oxidase activity, exocytosis from the azurophil granules, and actin polymerization. Our findings show that actin polymerization for neutrophil spreading and locomotion is regulated differently from that for phagocytosis. Also, the release of azurophil and specific granule contents is clearly shown to be regulated in a different way.

Topics: Actins; Antigens, CD; Calcimycin; Calcium; CD18 Antigens; CD4 Antigens; CD8 Antigens; Cell Adhesion; Cell Aggregation; Chemotaxis, Leukocyte; Cytochalasin B; Endothelium, Vascular; Female; Humans; Immunologic Deficiency Syndromes; In Vitro Techniques; Infant, Newborn; Kinetics; Leukocyte Count; Leukotriene B4; N-Formylmethionine Leucyl-Phenylalanine; Neutrophils; Oxygen Consumption; Platelet Activating Factor; Reference Values; Sepsis; Syndrome; T-Lymphocyte Subsets