NITRATES AND EICOSANOIDS Nitrates and endothelial prostacyclin production: studies in vitro

The hypothesis that nitrates evoke prostacyclin production by vascular endothelium has been reevaluated on cultured umbilical vein endothelial cells and in vascular fragments, both obtained from humans. Endothelial cell monolayers (passages 1 and 2) were washed free of culture medium and exposed for 3 to 5 min to buffer or nitroglycerin (NTG), isosorbide dinitrate (ISDN), or isosorbide-5mononitrate (ISMN) over a range of concentrations (10-9M to 10-6M) encompassing those usually attained in vivo, with or without 25 gM sodium arachidonate. Basal prostacyclin production, measured by radioimmunoassay of the stable metabolite 6-keto-PGF,a, depended on cell density in the endothelial monolayer (being higher in preconfluent cultures) and on incubation time. Basal prostacyclin, however, was not altered by incubation with NTG (3.3 + 2.0 pg/1000 cells without drug vs 3.9 + 3.8 pg/1000 cells with drug, mean + SD), ISDN (3.1 + 1.9 vs 3.1 + 2.2), or ISMN (2.0 0.9 vs 2.3 1.5) at 10-7M (all differences NS). Also, long-term incubation (2. 6, and 24 hr) with ISDN and ISMN did not alter prostacyclin production over control. Over a 30-fold increase (p < .001) in prostacyclin production was obtained with arachidonate stimulation, but incubation with nitrates did not significantly modify the stimulated production. Saphenous vein. mesenteric artery, and atrial appendage fragments incubated at 370 C for 20 min in a shaking water bath with a control buffer procluced 27.8 + 13.9, 189.7 -+ 75.2, and 662.3 -+ 390.6 pg 6-keto-PGF,Jm/giz tissue, respectively. Arachidonate (25 gM) increased prostacyclin production by these tissues to 158.8 ± 120.6 (p < .001). 381.0 + 48.2 (p < .01), and 885.6 -+ 291.4 (NS). respectively. However no significant variation from control prostacyclin was obtained with 20 min exposure to NTG, ISDN, or ISMN in a range from 10'M to 10-6M. No dose-effect relationship for these drugs on prostacyclin production was found. These data suggest that nitrates do not increase endothelial prostacyclin production. Circulation 71, No. 1, 176-182, 1985. NITRATES are widely used and universally accepted drugs in cardiovascular therapy. but their mechanism of action on the vessel wall has not been fully clarnfied.' Furthermore, nitrates exert extravascular actions such as inhibition of platelet function, 4 the mechanism for which is also unknown. Considerable interest has therefore been raised by reports that nitroglycerin (NTG) stimulates the production of prostacyclin (PGI) by vascular endothelium.' It has been hypothesized that the production of prostacyclin, From the Division of Hematology-Oncolog, Cornell Universitv Medical College. New York. Supported by a Thronmbosis SCOR grant (HL 18828) from the National Institutes of Health (Dr. Weksler). Dr. De Caterina, on leave of absence from the Institute of Clinical Physiolooy. C.N.R.. Pisa, Italy. is the recipient of a Fogarty International Research Fellowship from the Public Health Service. Department of Health and Human Sciences, U.S.A. Address for correspondence: Ratfaele De Caterina. M.D.. Division of Hematology-Oncology, Cornell Universitv Medical College. 1300 York Ave.. New York, NY 10021. Received July 16, 1984: revision accepted Oct. 4. 1984. through its vasodilator and antiaggregatory properties, accounts for at least some of the beneficial effects of nitrates.' However, contradictory reports on this subject have also appeared.9 "' Validation of the hypothesis that nitroderivatives affect vascular prostacyclin production is both of theoretical and practical relevance. Nitrates and drugs that inhibit the prostaglandin system. such as nonsteroidal anti-inflammatory agents, rank among the most frequently used drugs in medical therapy and are often used together. If the hypothesis that nitrates can act via stimulation of prostacyclin holds. their combined use would be expected to be detrimental.' We therefore decided to reexamine this issue systematically using various preparations of vascular endothelium and studying longacting nitrates as well as NTG.