Modulation of multiple neutrophil functions by preparative methods or trace concentrations of bacterial lipopolysaccharide

Human neutrophils were isolated from peripheral blood by four methods: 1) Ficoll-Hypaque gradients and erythrocyte lysis, 2) plasma-Percoll gradients, 3) a "lipopolysaccharide (LPS)-free" method yielding 85% neutrophils, and 4) by centrifugation of cells prepared by Method 3 through a plas...

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Published inThe American journal of pathology Vol. 119; no. 1; pp. 101 - 110
Main Authors Haslett, C, Guthrie, LA, Kopaniak, MM, Johnston, RB, Jr, Henson, PM
Format Journal Article
LanguageEnglish
Published Bethesda, MD ASIP 01.04.1985
American Society for Investigative Pathology
Subjects
Ammonium Chloride - pharmacology
Biological and medical sciences
Chemotaxis, Leukocyte - drug effects
Ficoll
Humans
Immunological methods for diagnosis and exploration
Immunopathology
Lipopolysaccharides - pharmacology
Medical sciences
Neutrophils - drug effects
Neutrophils - enzymology
Neutrophils - physiology
Neutrophils - ultrastructure
Other methods
Superoxides - metabolism
Human
Leucocyte function
Leukocyte
Neutrophile
Interaction
Lipopolysaccharide
Exploration
Isolation
Technique
Comparative study
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Summary:Human neutrophils were isolated from peripheral blood by four methods: 1) Ficoll-Hypaque gradients and erythrocyte lysis, 2) plasma-Percoll gradients, 3) a "lipopolysaccharide (LPS)-free" method yielding 85% neutrophils, and 4) by centrifugation of cells prepared by Method 3 through a plasma-Percoll gradient to produce pure neutrophils. The use of the Ficoll-Hypaque method resulted in spontaneous change of cell shape, enhanced formyl-methionyl-leucyl-phenylalanine (FMLP)-stimulated release of superoxide anion, increased release of lysosomal enzymes upon subsequent FMLP stimulation, and reduced chemotactic responsiveness, by comparison with the other methods. These effects were not due to erythrocyte lysis by NH4C1 but were reproduced by exposure of neutrophils prepared by the "LPS-free" method or the use of plasma-Percoll gradients to 10-100 ng/ml LPS. Neutrophil change of shape and stimulated O-2 production were particularly sensitive markers of these effects. The effects of trace concentrations of LPS in the modulation of neutrophil function may have relevance to the pathophysiology of endotoxemia and its resultant tissue injury.
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ISSN:0002-9440
1525-2191