Cellular effects of endotoxin in vitro: mobility of endotoxin in the plasma membrane of hepatocytes and neuroblastoma cells.
Lipopolysaccharide labeled with fluorescein isothiocyanate (FITC-LPS) was used to examine interactions between endotoxin and plasma membrane in isolated rat hepatocytes and mouse neuroblastoma NB41A3 cells. At the same endotoxin to cell ratio, hepatocytes bound more toxin than did neuroblastoma cells. At a dose of 12 micrograms/mg dry wt, a bound mobile fraction of between 60 and 75% of FITC-LPS was found on hepatocytes at 25 degrees C with a lateral diffusion coefficient (D) of 4.0 X 10(-9) cm2/s. In neuroblastoma cells, the mobile fraction was larger (85-90%), with D 1.0 X 10(-8) cm2/s. D was temperature-dependent between 10 and 37 degrees C and increased from 1.8 X 10(-9) to 1.0 X 10(-8) cm2/s in hepatocytes and from 9.4 X 10(-9) to 1.9 X 10(-8) cm2/s in neuroblastoma cells. In both types of cell, nonviable (cells which did not exclude Trypan blue) as compared to viable cells showed different recovery patterns and 100% of the probe molecules were mobile. These results suggest that: (1) endotoxin binding to mammalian cells consists of two subpopulations with different mobilities; (2) binding of the immobile fraction is dependent on cellular integrity; and (3) the differences in binding, lateral mobility, and size of the immobile fraction in hepatocytes and neuroblastoma cells may be due to variations in membrane composition and/or number of binding sites.
D000818:Animals; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D004731:Endotoxins / Q000378:metabolism*; D016650:Fluorescein-5-isothiocyanate; D005452:Fluoresceins / Q000378:metabolism / Q000633:toxicity*; D066298:In Vitro Techniques; D007700:Kinetics; D008070:Lipopolysaccharides / Q000378:metabolism / Q000633:toxicity*; D008099:Liver / Q000378:metabolism*; D008297:Male; D008560:Membrane Fluidity; D051379:Mice; D009447:Neuroblastoma / Q000378:metabolism*; D051381:Rats; D011919:Rats, Inbred Strains; D013050:Spectrometry, Fluorescence; D013861:Thiocyanates / Q000378:metabolism / Q000633:toxicity*
D004731:Endotoxins; D005452:Fluoresceins; D008070:Lipopolysaccharides; D013861:Thiocyanates; D016650:Fluorescein-5-isothiocyanate