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While hemolysis in patients with sepsis is associated with increased mortality its mechanisms are unknown and Toll-like receptor (TLR)-4 mediated effects, complement-mediated hemolysis, or direct cell membrane effects are all conceivable mechanisms. In this study, we tested the hypotheses that toxic lipopolysaccharide (LPS) as well as non-toxic RS-LPS evokes hemolysis (1) by direct membrane effects, and (2) independent of the complement system and TLR-4 activation. We found, that incubation with LPS resulted in a marked time and concentration dependent increase of free hemoglobin concentration and LDH activity in whole blood and washed red cells. Red cell integrity was diminished as shown by decreased osmotic resistance, formation of schistocytes and rolls, and a decrease in red cell membrane stiffness. Non-toxic RS-LPS inhibited the LPS-evoked increase in TNF-α concentration demonstrating its TLR-4 antagonism, but augmented LPS-induced increase in supernatant hemoglobin concentration and membrane disturbances. Removal of plasma components in washed red cell assays failed to attenuate hemolysis. In summary, this study demonstrates direct physicochemical interactions of LPS with red cell membranes resulting in hemolysis under in vitro conditions. It might thus be hypothesized, that not all effects of LPS are mediated by TLR and may explain LPS toxicity in cells missing TLR.

Bacterial lipopolysaccharides are believed to have a toxic effect on human cell membranes. In this study, the influence of a lipopolysaccharide (LPS) from Escherichia coli on the structure, the dynamics and the mechanical strength of phospholipid membranes are monitored by nuclear magnetic resonance spectroscopy (NMR) and by atomic force microscopy (AFM). Model membranes are formed from 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC) and are either prepared as multilamellar bulk samples or multilamellar vesicles. Field gradient NMR data directly prove the rapid integration of LPS into DMPC membranes. Solid state NMR experiments primarily detect decreasing molecular order parameters with increasing LPS content. This is accompanied by a mechanical softening of the membrane bilayers as is shown by AFM indentation measurements. Altogether, the data prove that lipopolysaccharide molecules quickly insert into phospholipid bilayers, increase membrane fluctuation amplitudes and significantly weaken their mechanical stiffness.

No abstract

Choice chamber preference tests were used to determine if camel crickets, Ceuthophilus secretus, use pheromones in forming aggregations. Insects occurred significantly more often on choice chamber sides having filter paper scented by exposure to conspecifics than they did on unconditioned filter paper. Insects also formed aggregations significantly more quickly when exposed to conspecifics as opposed to nonconspecific scents in the choice chambers. These data demonstrate that C. secretus use an aggregation pheromone. Insects occurred significantly more often on sides conditioned with ether extracts of whole insect bodies as opposed to unconditioned sides. Insects also occurred significantly more often on filter paper conditioned with abdomen extracts as opposed to unconditioned filter paper, but no preference was shown for extracts from the head and thorax region. These data demonstrate that the abdomen is the generalized site of aggregation pheromone production. Insects which had both antennae removed were unable to form aggregations in response to conspecific scent, thus indicating that the antennae are the site of aggregation pheromone reception.