title: Nanomechanics of the endothelial glycocalyx in experimental sepsis.
abstract: The endothelial glycocalyx (eGC), a carbohydrate-rich layer lining the luminal side of the endothelium, regulates vascular adhesiveness and permeability. Although central to the pathophysiology of vascular barrier dysfunction in sepsis, glycocalyx damage has been generally understudied, in part because of the aberrancy of in vitro preparations and its degradation during tissue handling. The aim of this study was to analyze inflammation-induced damage of the eGC on living endothelial cells by atomic-force microscopy (AFM) nanoindentation technique. AFM revealed the existence of a mature eGC on the luminal endothelial surface of freshly isolated rodent aorta preparations ex vivo, as well as on cultured human pulmonary microvascular endothelial cells (HPMEC) in vitro. AFM detected a marked reduction in glycocalyx thickness (266 ± 12 vs. 137 ± 17 nm, P<0.0001) and stiffness (0.34 ± 0.03 vs. 0.21 ± 0.01 pN/mn, P<0.0001) in septic mice (1 mg E. coli lipopolysaccharides (LPS)/kg BW i.p.) compared to controls. Corresponding in vitro experiments revealed that sepsis-associated mediators, such as thrombin, LPS or Tumor Necrosis Factor-α alone were sufficient to rapidly decrease eGC thickness (-50%, all P<0.0001) and stiffness (-20% P<0.0001) on HPMEC. In summary, AFM nanoindentation is a promising novel approach to uncover mechanisms involved in deterioration and refurbishment of the eGC in sepsis.
authors.affiliation: Department of Medicine D, Division of General Internal Medicine, Nephrology, and Rheumatology, University Hospital, Muenster, Muenster, Germany.
authors.forename: Anne
authors.initials: A
authors.lastname: Wiesinger
authors.forename: Wladimir
authors.initials: W
authors.lastname: Peters
authors.forename: Daniel
authors.initials: D
authors.lastname: Chappell
authors.forename: Dominik
authors.initials: D
authors.lastname: Kentrup
authors.forename: Stefan
authors.initials: S
authors.lastname: Reuter
authors.forename: Hermann
authors.initials: H
authors.lastname: Pavenstädt
authors.forename: Hans
authors.initials: H
authors.lastname: Oberleithner
authors.forename: Philipp
authors.initials: P
authors.lastname: Kümpers
mesh_terms: D000818:Animals; D001011:Aorta / Q000473:pathology / Q000648:ultrastructure; D002470:Cell Survival; D004727:Endothelium / Q000473:pathology* / Q000648:ultrastructure*; D019446:Endotoxemia / Q000382:microbiology / Q000473:pathology; D019276:Glycocalyx / Q000737:chemistry*; D006801:Humans; D008297:Male; D008810:Mice, Inbred C57BL; D018625:Microscopy, Atomic Force; D053758:Nanoparticles / Q000737:chemistry* / Q000648:ultrastructure; D018805:Sepsis / Q000473:pathology*; D013997:Time Factors