Release of platelet activation factor from activated neutrophils. Transglutaminase-dependent enhancement of transbilayer movement across the plasma membrane.
Extracellular release of platelet activating factor (PAF) following synthesis in inflammatory cells is variable and modulated by a number of as yet undefined cellular mechanisms. Using human neutrophils loaded with the tritiated, nonmetabolizable PAF analog, 1-O-alkyl-2-N-methylcarbamyl-sn-glycero-3-phosphocholine (C-PAF), extracellular release was studied by techniques involving an albumin extraction method. Further modeling of plasma membrane events in the neutrophil was accomplished using movement across the membrane of erythrocyte ghosts. The data demonstrate that C-PAF release is dependent on cellular activation and is accompanied by alterations in the physical properties of the plasma membrane as measured by enhancement of merocyanine 540 (MC540) staining, as well as by bulk, nonspecific transbilayer movement of endogenous phospholipids as detected by the procoagulant activity of externalized phosphatidylserine (a phospholipid class usually sequestered in the plasma membrane inner leaflet). The finding that competitive inhibitors of transglutaminase significantly inhibited C-PAF release, enhancement of MC540 staining, and externalization of phosphatidylserine, strongly suggest a role for this enzyme in the enhancement of phospholipid transbilayer movement. Furthermore, the data suggest that recycling of C-PAF in the plasma membrane is likewise transglutaminase dependent and limits the net extracellular release of C-PAF which, like liberation of endogenously produced PAF, is dependent on extracellular "acceptors" and shown to be albumin concentration- and cell density-dependent.
Bratton
D L
DL
National Jewish Center for Immunology and Respiratory Medicine, Denver, Colorado 80206.
D002462:Cell Membrane / Q000378:metabolism; D003538:Cystamine / Q000494:pharmacology; D004910:Erythrocyte Membrane / Q000502:physiology; D006801:Humans; D066298:In Vitro Techniques; D015759:Ionomycin / Q000494:pharmacology; D007700:Kinetics; D008051:Lipid Bilayers; D008563:Membrane Lipids / Q000097:blood; D009240:N-Formylmethionine Leucyl-Phenylalanine / Q000494:pharmacology; D009504:Neutrophils / Q000187:drug effects / Q000201:enzymology / Q000502:physiology*; D010972:Platelet Activating Factor / Q000031:analogs & derivatives / Q000378:metabolism*; D012521:Sarcosine / Q000494:pharmacology; D012709:Serum Albumin / Q000494:pharmacology; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D013997:Time Factors; D011503:Transglutaminases / Q000378:metabolism*
D008051:Lipid Bilayers; D008563:Membrane Lipids; D010972:Platelet Activating Factor; D012709:Serum Albumin; D015759:Ionomycin; D009240:N-Formylmethionine Leucyl-Phenylalanine; D011503:Transglutaminases; D013755:Tetradecanoylphorbol Acetate; D003538:Cystamine; D012521:Sarcosine