Transcellular metabolism of arachidonic acid: increased platelet thromboxane generation in the presence of activated polymorphonuclear leukocytes.
Human polymorphonuclear leukocytes (PMN) activated by n-formyl-methionyl-leucyl-phenylalanine (fMLP), in the presence of cytochalasin B, are able to induce activation of coincubated autologous platelets "via" cathepsin G released from the azurophilic granules. However, thromboxane (Tx) B2 production in this system cannot be completely explained by cathepsin G-stimulated platelet arachidonate metabolism. Indeed, the amount of TxB2 found in supernatants of platelet/PMN suspensions challenged with 1 mumol/L fMLP was twofold to fourfold higher than that measured when platelets were stimulated by supernatants from fMLP-activated PMN. In the present report, we analyzed the possibility that PMN-induced TxB2 production in this system is the result of transcellular metabolism of arachidonic acid (AA) between fMLP-activated PMN and cathepsin G-stimulated platelets. 3H-AA-labeled PMN were used to test if a transfer of AA or metabolite(s) occur from PMN to platelets. Our results showed that: (1) 3H-TxB2 and 3H-12-HHT are synthesized when 3H-AA-labeled PMN are activated mixed to unlabeled platelets; (2) total radioactivity released by fMLP-stimulated PMN is increased in the presence of platelets, whereas the membrane content of unesterified 3H-AA is reduced; (3) platelet cyclooxygenase inhibition completely prevents 3H-TxB2 synthesis; and (4) inhibition of cathepsin G-induced platelet activation with the antiprotease eglin C blocks the formation of 3H-TxB2. These data show that in the experimental system used, platelets use PMN-derived unmetabolized AA to synthesize TxB2.
Maugeri
N
N
Giulio Bizzozero Laboratory of Platelet and Leukocyte Pharmacology, Istituto di Richerche Farmacologiche Mario Negri, Consorzio Mario Negri Sud, Santa Maria Imbaro, Italy.
Evangelista
V
V
Piccardoni
P
P
Dell'Elba
G
G
Celardo
A
A
de Gaetano
G
G
Cerletti
C
C
D001095:Arachidonic Acids / Q000097:blood*; D001792:Blood Platelets / Q000187:drug effects / Q000378:metabolism* / Q000502:physiology; D056649:Cathepsin G; D002403:Cathepsins / Q000494:pharmacology; D002450:Cell Communication; D002851:Chromatography, High Pressure Liquid; D006801:Humans; D007700:Kinetics; D009240:N-Formylmethionine Leucyl-Phenylalanine / Q000494:pharmacology; D009504:Neutrophils / Q000502:physiology*; D015539:Platelet Activation / Q000187:drug effects; D012697:Serine Endopeptidases; D013929:Thromboxane B2 / Q000097:blood*
D001095:Arachidonic Acids; D013929:Thromboxane B2; D009240:N-Formylmethionine Leucyl-Phenylalanine; D002403:Cathepsins; D012697:Serine Endopeptidases; C534094:CTSG protein, human; D056649:Cathepsin G