title: Aspects of the haemolytic reaction induced by Kanagawa haemolysin of Vibrio parahaemolyticus.
abstract: Vibrio parahaemolyticus, an important enteric pathogen, produces toxin (Kanagawa haemolysin, KH), the presence of which correlates well with pathogenicity. KH induced lysis of human red blood cells (HRBC); the kinetics were strongly dependent on KH concentration (0-1 HU/ml) and rather independent of target cell concentration [0.5 < or = haematocrit (%) < or = 6] and the ratio KH:HRBC. The suggestion that KH-induced haemolysis is due to colloid osmosis is supported by results indicating: (1) osmotic protection (by suspension in iso-osmotic choline chloride, D-sorbitol or L-valine, or MOPS-buffered saline with added sucrose), (2) a cell volume increase prior to lysis, and (3) an increase in HRBC cation (86Rb+) influx after KH addition, indicating raised passive cation permeation. The effect of temperature on KH-induced haemolysis indicates the importance of processes other than the action of a simple water-filled pore, because of the high activation energy [53.30 +/- 2.79 kJ (mol.)-1] involved. Although haemolytic rate was attenuated by washout after 5 min KH exposure, the KH-induced lesion itself was not susceptible to washout by either extracellular volume expansion (at constant osmolarity) or centrifugation/resuspension. This suggests that HRBC binding of KH from aqueous solution still continues after 5 min exposure at 37 degrees C. Pre-vortexing KH with dibutyl phthalate (DBP) dramatically reduced the haemolytic activity of the aqueous toxin preparation, suggesting a protein-lipid interaction, which may support the contention that KH can move from a hydrophilic to a hydrophobic environment. Two features were identified that are characteristic of highly purified TDH preparations: (1) thermostability of haemolysin, and (2) monovalent cation selectivity series of lesion: Cs+ > Li+ > K+ > Rb+ > Na+, confirming that TDH is the important leak-inducing agent of KH.
authors.affiliation: University Laboratory of Physiology, University of Oxford, U.K.
authors.forename: J S
authors.initials: JS
authors.lastname: Huntley
authors.forename: A C
authors.initials: AC
authors.lastname: Hall
mesh_terms: D000328:Adult; D002414:Cations, Monovalent / Q000378:metabolism; D048429:Cell Size / Q000187:drug effects / Q000502:physiology; D002794:Choline / Q000494:pharmacology; D004912:Erythrocytes / Q000166:cytology / Q000187:drug effects*; D006400:Hematocrit; D006460:Hemolysin Proteins / Q000633:toxicity*; D006461:Hemolysis / Q000502:physiology*; D006801:Humans; D007553:Isotope Labeling; D007700:Kinetics; D009995:Osmosis; D012413:Rubidium / Q000378:metabolism; D013012:Sorbitol / Q000494:pharmacology; D013696:Temperature; D014633:Valine / Q000494:pharmacology; D014736:Vibrio parahaemolyticus / Q000378:metabolism*
chemical_list: D002414:Cations, Monovalent; D006460:Hemolysin Proteins; D013012:Sorbitol; D014633:Valine; D012413:Rubidium; D002794:Choline