MilliporeSigma
  • Home
  • Search Results
  • Mechanisms of leukocyte distribution during sepsis: an experimental study on the interdependence of cell activation, shear stress and endothelial injury.

Mechanisms of leukocyte distribution during sepsis: an experimental study on the interdependence of cell activation, shear stress and endothelial injury.

Critical care (London, England) (2010-11-10)
Annette Ploppa, Volker Schmidt, Andreas Hientz, Joerg Reutershan, Helene A Haeberle, Boris Nohé
ABSTRACT

This study was carried out to determine whether interactions of cell activation, shear stress and platelets at sites of endothelial injury explain the paradoxical maldistribution of activated leukocytes during sepsis away from local sites of infection towards disseminated leukocyte accumulation at remote sites. Human umbilical venous endothelial cells (HUVEC) and polymorphonuclear neutrophils (PMN) were activated with lipopolysaccharide at 100 and 10 ng/ml to achieve adhesion molecule patterns as have been reported from the hyper- and hypo-inflammatory stage of sepsis. To examine effects of leukocyte activation on leukocyte-endothelial interactions, activated HUVEC were perfused with activated and non-activated neutrophils in a parallel plate flow chamber. Adhesion molecule expression and function were assessed by flow cytometry and blocking antibodies. In a subset of experiments the sub-endothelial matrix was exposed and covered with platelets to account for the effects of endothelial injury. To investigate interactions of these effects with flow, all experiments were done at various shear stress levels (3 to 0.25 dyne/cm(2)). Leukocyte-endothelial interactions were analyzed by videomicroscopy and analysis of covariance. Activation of neutrophils rendered adhesion increasingly dependent on shear stress reduction. At normal shear stress, shedding of L-selectin decreased adhesion by 56%. Increased rolling fractions of activated PMN at low shear stress revealed impaired integrin affinity despite numerical up-regulation of CD11b. On sub-maximally activated, intact HUVEC shear stress became the prevailing determinant of adhesion. Presence of a platelet-covered injury with high surface density of P-selectin was the strongest variable for adhesion. When compared to maximally activated HUVEC, platelets increased neutrophil adhesion by 2.7-fold. At sub-maximal activation a 10-fold increase was observed (P < 0.05 for all). L-selectin shedding and integrin dysfunction render leukocyte adhesion increasingly susceptible to shear stress and alternative adhesion receptors. In combination, these effects inhibit recruitment to normally perfused sites with intact endothelium and favor maldistribution towards sites with compromised perfusion or endothelial injury.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
Medium 199, Modified, with Earle′s salts, without L-glutamine, sodium bicarbonate, and phenol red, powder, suitable for cell culture
Sigma-Aldrich
Medium 199, With Earle′s salts and sodium bicarbonate, without L-glutamine, liquid, sterile-filtered, suitable for cell culture
Sigma-Aldrich
Medium 199, 10 ×, With Hanks′ salts, without L-glutamine and sodium bicarbonate, liquid, sterile-filtered, suitable for cell culture
Sigma-Aldrich
Medium 199, With Earle′s salts, L-glutamine and sodium bicarbonate, liquid, sterile-filtered, suitable for cell culture
Sigma-Aldrich
Medium 199, HEPES Modification, with Earle′s salts, L-glutamine and 25 mM HEPES, without sodium bicarbonate, powder, suitable for cell culture
Sigma-Aldrich
Medium 199, With Hanks′ salts and L-glutamine, without sodium bicarbonate, powder, suitable for cell culture
Sigma-Aldrich
Medium 199, With Hanks′ salts and sodium bicarbonate, without L-glutamine, liquid, sterile-filtered, suitable for cell culture
Sigma-Aldrich
Medium 199, HEPES Modification, with Earle′s salts, 25 mM HEPES and sodium bicarbonate, without L-glutamine, liquid, sterile-filtered, suitable for cell culture
Sigma-Aldrich
Medium 199, 10 ×, With Earle′s salts, without L-glutamine and sodium bicarbonate, liquid, sterile-filtered, suitable for cell culture
Sigma-Aldrich
Medium 199, With Earle′s salts and L-glutamine, without sodium bicarbonate, powder, suitable for cell culture
Sigma-Aldrich
Lipopolysaccharides from Escherichia coli O26:B6, γ-irradiated, BioXtra, suitable for cell culture
Sigma-Aldrich
Lipopolysaccharides from Escherichia coli O26:B6, ≥10,000 EU/mg, purified by phenol extraction
Sigma-Aldrich
Lipopolysaccharides from Escherichia coli O26:B6, purified by gel-filtration chromatography
Sigma-Aldrich
Lipopolysaccharides from Escherichia coli O26:B6, purified by trichloroacetic acid extraction
Sigma-Aldrich
Lipopolysaccharides from Escherichia coli O26:B6, Ready Made solution, 1 mg/mL, 0.2 μm filtered