Thrombin stimulates arachidonate metabolism in murine tumor cells.
Thrombin can be formed in the tumor cell microenvironment following activation of the clotting cascade by procoagulants of cancer or host cells. We have tested here the effects of thrombin, either "endogenous" or "exogenous" (see below), on arachidonate mobilization from membrane phospholipids of mouse mammary tumor virus-induced (MMTV) carcinoma cells. These tumor cells exhibit in vitro a tissue type procoagulant activity (130 thromboplastin units/10(4) cells) and are therefore able to induce thrombin formation in a plasmatic milieu. To verify the effect of thrombin formation by tumor cell procoagulant ("endogenous thrombin"), either human or mouse platelet-free plasma (20% in DMEM) was added to the cell layer (prelabelled for 5 hr with a trace amount (0.013 microM) of 3H-arachidonate) and the system was recalcified (15 mM CaCl2). Thin-layer radiochromatography of the culture medium showed a significant release of 3H-labelled arachidonate products PGE2, PGF2 alpha and 6-ketoPGF1 alpha after 1 hr of incubation. To verify the effect of thrombin formation from host sources ("exogenous thrombin"), either bovine or purified human alpha-thrombin (0.1-10 U/ml) was added to the cells for different periods (from 5 min to 20 hr). Exogenous thrombin stimulated arachidonate release and metabolism in a dose-related manner. With short labelling periods (0.013 microM 3H-arachidonate for 30 min-1 hr) thrombin stimulated the release of unmetabolized 3H-arachidonate, but not of 3H-arachidonate metabolites. These processes were inhibited by a specific inhibitor of thrombin enzymatic activity (alpha-NAPAP, 140 microM) and by a cyclo-oxygenase inhibitor (ASA 4mM). Tumor-associated procoagulants may thus contribute not only to fibrin deposition but also to generation of multipotent mediators such as arachidonate metabolites.
D000818:Animals; D016718:Arachidonic Acid; D001095:Arachidonic Acids / Q000378:metabolism*; D001780:Blood Coagulation Tests; D002478:Cells, Cultured; D002855:Chromatography, Thin Layer; D004305:Dose-Response Relationship, Drug; D008325:Mammary Neoplasms, Experimental / Q000378:metabolism*; D051379:Mice; D011451:Prostaglandin-Endoperoxide Synthases / Q000502:physiology; D013917:Thrombin / Q000494:pharmacology*; D014412:Tumor Virus Infections / Q000378:metabolism
D001095:Arachidonic Acids; D016718:Arachidonic Acid; D011451:Prostaglandin-Endoperoxide Synthases; D013917:Thrombin