Cellular activating properties and morphology of membrane-bound and purified meningococcal lipopolysaccharide

Neisseria meningitidis, the cause of epidemic meningitis and acute lethal s epsis, synthesizes surplus lipopolysaccharides (LPSs) during growth, which are released as outer membrane vesicles (OMV) or 'blebs'. Meningococcal dis ease severity is related to plasma LPS levels. We have compared the biologi cal activities of native outer membrane vesicles (nOMV) to those of purifie d Nm-LPS (Nm-LPS) and LPS-depleted OMV (dOMV) prepared from N. meningitidis . The LPS content of nOMV was determined spectrophotometrically by quantify ing KDO and by silver-stained SDS-PAGE gels. The morphology of the preparat ions was studied by transmission electron microscopy. The Li,Limulus amoebo cyte lysate (LAL) assay was used to quantify LPS in the plasma solutions. T he preparations were diluted in endotoxin-free heparin plasma to equal amou nts of LPS (w/w) in the range 50-5000 pg/ml, The biological reactivity was tested by: (i) a monocyte target-assay (monocyte purity greater than or equ al to 96%); and (ii) a whole blood model, measuring the secretion of TNF-al pha and IL-6 induction of procoagulant activity in monocytes (PCA). In both models, nOMV induced dose-dependent cell responses (TNF-alpha, IL-6, PCA) similar to purified Nm-LPS, whereas dOMV induced minimal responses. However , LAL activity was significantly higher for nOMV than fur purified Nm-LPS a nd dOMV. The cellular responses of purified Nm-LPS and nOMV were reduced (> 95%) by a specific anti-CD14-antibody.