IMMUNOLOGICAL GENERATION OF A PLATELET-ACTIVATING FACTOR AND A PLATELET-LYTIC FACTOR IN THE RAT

Abstract

Antigen challenge of the rat peritoneal cavity which was prepared with Ig[immunoglobulin]Ga-rich antiserum generated activities which released [14C]-serotonin from pre-labeled human platelets. After adsorption of these activities onto Amberlite XAD-8 and elution in 80% ethanol, 2 factors of differing polarity were resolved by chromatography on DEAE-cellulose in organic solvents. The activity eluting in the 7:1 chloroform:methanol solvent contained a platelet-lytic factor (PLF) assessed by the parallel release of lactic acid dehydrogenase and [14C]-serotonin; the cytotoxicity of this fraction was confirmed by phase-contrast microscopy examination which demonstrated fragmentation of the exposed platelets. The activity eluting in the 1:1 methanol:aqueous 1.0 M ammonium carbonate solvent was a platelet-activating factor (PAF) as defined by release of [14C]-serotonin without lactic acid dehydrogenase. The lytic and the activating principles were separable from slow reacting substance of anaphylaxis and polymorphonuclear leukocyte chemotactic activity, and each presented a single activity peak of differing mobility when chromatographed on silica gel H plates. Human eosinophil phospholipase D inactivated the lytic factor by more than 85% in 2 h at 37.degree. without affecting the activity of the activating factor. The release of [14C]-serotonin induced by the PAF was not affected by the absence of Ca from the medium or by elevations in the platelet concentrations of cyclic[c]AMP or cGMP that resulted from pre-incubation of platelets with prostaglandin D2 or sodium ascorbate, respectively.