Endotoxin binding to erythrocyte membrane and erythrocyte deformability in human sepsis and in vitro.
OBJECTIVE
Several studies have shown that lipopolysaccharide and lipid A impair red blood cell deformability and. However, it is unclear whether impaired red blood cell deformability is associated with binding of lipopolysaccharide to the red blood cell membrane.


METHODS
Analysis of hydroxymyristic acid content in red blood cell membranes and red blood cell deformation in patients with Gram-negative septicemia and after incubation of red blood cells from healthy adults with 100 microg of lipid A or 1 mg of lipopolysaccharide per milliliter of red blood cell in buffer solution and in whole blood. Hydroxymyristic acid is a fatty acid of the lipid A part of lipopolysaccharide in most Gram-negative bacteria.


METHODS
University research laboratories.


METHODS
Ten healthy adults and four patients with clinical and laboratory signs of septicemia.


METHODS
Blood sampling.


RESULTS
Red blood cell deformation was measured with a laser-diffraction shearing device (Rheodyn) and a computerized micropore filtration system (CTA). Lipopolysaccharide and lipid A binding to red blood cell membranes was studied by measuring the amide-linked hydroxymyristic acid by gas chromatography. The detection rates of hydroxymyristic acid were 82% for lipopolysaccharide and 79% for lipid A in buffer solution. In membranes of washed red blood cell, the detection rates of lipopolysaccharide and lipid A were 0.26 +/- 0.03% (2.6 +/- 0.3 microg/mL) and 1.3 +/- 0.5% (1.3 +/- 0.5 microg/mL), and in red blood cell membranes of whole blood the detection rates were 2.6% (25.5 microg/mL) and 4.1% (4.1 microg/mL), respectively. The lipopolysaccharide content in red blood cell membranes of septic patients ranged from 47 to 103 microg/mL of red blood cell. Red blood cell deformation in the Rheodyn and in the CTA were not influenced by lipopolysaccharide incubated with washed red blood cells. In the Rheodyn, red blood cell deformation was significantly decreased by 18% after lipid A incubation in washed red blood cells, by 26% after lipopolysaccharide incubation in whole blood, and by 31% in septic patients. Similar effects were observed when we used the CTA.


CONCLUSIONS
Red blood cell deformation is decreased in septic patients, after incubation of washed red blood cells with lipid A and of whole blood with lipopolysaccharide. Lipopolysaccharide did not influence red blood cell deformation after incubation with washed red blood cells. The decrease of red blood cell deformation was related to the amount of hydroxymyristic acid measured in red blood cell membranes, suggesting that endotoxin binding directly affects mechanical properties of red blood cells.
Pöschl
Johannes M B
JM
Division of Neonatology, Department of Pediatrics, University of Heidelberg Medical School, Germany. johannes_poeschl@med.uni-heidelberg.de
Leray
Claude
C
Ruef
Peter
P
Cazenave
Jean P
JP
Linderkamp
Otwin
O
D000328:Adult; D016470:Bacteremia / Q000097:blood* / Q000382:microbiology* / Q000503:physiopathology; D001426:Bacterial Proteins / Q000032:analysis*; D001665:Binding Sites; D015415:Biomarkers / Q000032:analysis; D016022:Case-Control Studies; D002849:Chromatography, Gas; D003577:Cytochrome P-450 Enzyme System / Q000032:analysis*; D004731:Endotoxins / Q000009:adverse effects*; D004907:Erythrocyte Deformability; D004910:Erythrocyte Membrane / Q000737:chemistry*; D016905:Gram-Negative Bacterial Infections / Q000097:blood* / Q000382:microbiology* / Q000503:physiopathology; D006801:Humans; D017078:Laser-Doppler Flowmetry; D008050:Lipid A / Q000009:adverse effects*; D008070:Lipopolysaccharides / Q000009:adverse effects*; D008833:Microcirculation; D008851:Micropore Filters; D006899:Mixed Function Oxygenases / Q000032:analysis*; D009251:NADPH-Ferrihemoprotein Reductase
D001426:Bacterial Proteins; D015415:Biomarkers; D004731:Endotoxins; D008050:Lipid A; D008070:Lipopolysaccharides; D003577:Cytochrome P-450 Enzyme System; D006899:Mixed Function Oxygenases; D009251:NADPH-Ferrihemoprotein Reductase; C052423:flavocytochrome P450 BM3 monoxygenases