PG Endoperoxide Research Articles

Inhibition of Platelet Aggregation by Sulfinpyrazone

The action mechanism of sulfinpyrazone (SUL) was examined with rabbit platelets by testing its influence on the aggregations induced by ADP and collagen, on platelet content of cyclic AMP, on the generation of labile aggregation-stimulating substance (LASS) by platelets, and on platelet aggregation induced by LASS. The generation of LASS was examined by incubating 20μg/ml of arachidonic acid (AA) with washed platelets in the presence or absence of SUL, and measuring platelet aggregation when the incubated mixture was added to platelet-rich plasma (PRP) containing 20μM indomethacin. SUL was administered in a dose of 200mg or 400mg to healthy adults, and plasma concentrations and aggregations induced by ADP, collagen, and epinephrine were measured before and 1, 2, 4, and 6 hours after administration.At 3mM concentration, SUL exhibited little inhibitory effect on ADP-induced aggregation of rabbit platelets; but at concentrations of 1mM and above SUL inhibited collagen-induced aggregation, showing suppression of the release reaction. At 10mM, SUL elicited no change in platelet levels of cyclic AMP, and did not promote the PGE1-induced increase in cyclic AMP. LASS-induced aggregation was slightly inhibited by 3mM SUL. LASS formation was markedly inhibited by 0.3mM SUL, and at higher concentrations the inhibition increased dose-dependently. After administration of SUL in a dose of 200mg, the plasma level of the drug reached a maximum (average 10.5μg/ml) after 1 hour and after 6 hours had fallen to one-third of that level (3.0μg). With a does of 400mg, the plasma levels reached a maximum (35.5μg/ml) after 2 hours and fell to approximately one-third of that level (12.7μg/ml) after 6 hours. The maximum inhibition of platelet aggregation was seen between 4 and 6 hours after administration, showing a discrepancy with the peak plasma level.SUL inhibited the secondary but not the primary aggregation induced by ADP, only slightly inhibited collagen-induced aggregation, but strongly suppressed epinephrine-induced aggregation. SUL also inhibited the biosyntheses of PG endoperoxides and thromboxane A2, and at high concetration showed a direct inhibitory effect against these mediators. The results can be interpreted in terms of the mechanism of inhibition of LASS generation, in view of the report that SUL, unlike aspirin, does not inhibit the activity of cyclo-oxygenase at concentrations that inhibit platelet aggregation.

Thrombin, collagen and A23187 stimulated endogenous platelet arachidonate metabolism: Differential inhibition by PGE 1, local anesthetics and a serine-protease inhibitor

The formation of malondialdehyde (MDA) and rabbit aorta contracting substance (RCS) induced by treatment of platelets with thrombin and collagen, but not that produced from exogenous arachidonic acid, is inhibited by prostaglandin E 1 (10 −8 − 10 −7M), the local anesthetics tetracaine, SKF 525-A and dibucaine (1 mM), and the serine-protease inhibitor phenylmethanesulfonyl fluoride (PMSF). The burst in oxygen consumption which accompanies platelet stimulation by thrombin and collagen in the presence of antimycin A, known to be due to the oxidation of endogenous arachidonate, is also markedly suppressed by PGE 1, tetracaine and PMSF. The inhibitory effect of PGE 1 is strongly potentiated by theophylline (1.0 mM). Addition of the Ca 2+ ionophore A23187 to platelet suspensions overcomes PGE 1 and PMSF inhibition of MDA and RCS formation, and induces a vigorous increase in O 2 consumption. Tetracaine and dibucaine, however, block the responses to A23187. Formation of MDA and RCS (a mixture of PG endoperoxides and TXA 2) due to stimulation by thrombin and collagen depends upon activation of Ca 2+-dependent phospholipase A 2 (PLA 2) to supply free arachidonate from specific membrane phospholipids. These experiments therefore indicate that increased cellular cAMP, induced by PGE 1, antagonizes the mobilization of the Ca 2+ which is normally required for PLA 2 activity. Thrombin-stimulated platelets exhibit enhanced 45Ca uptake which probably reflects exchange of extracellular Ca 2+ with an increased available pool of exchangeable intracellular Ca 2+. PGE 1 strongly suppresses this 45Ca uptake, providing more direct evidence supporting the view that cAMP prevents the rise in free cytoplasmic Ca 2+ induced by thrombin. Under conditions which make sufficient free cytoplasmic Ca 2+ available (i.e., A23187), despite high cellular cAMP, formation of RCS and MDA, and O 2 uptake are nearly normal indicating that activation of PLA 2 can occur. Local anesthetics on the other hand since they abolish the response to A23187 as well, appear to directly antagonize the ability of Ca 2+ to activate PLA 2. The effect of PMSF suggests that stimulus-specific proteases may be involved in the thrombin and collagen-induced activation of PLA 2 activity.

Microsomal preparations of normal bovine iris-ciliary body generate prostacyclin-like but not thromboxane-A 2-like activity

Indomethacin-treated bovine iris-ciliary body microsomes (IBIM) have been studied for their ability to convert PG endoperoxides into either thromboxance-A 2 (TxA 2)-like or prostacyclin (PGI 2)-like activity. The biological activity of the ocular tissue microsomes were compared with either indomethacin-treated human platelet microsomes (for TxA 2-like activity) or rabbit aorta microsomes (for PGI 2-like activity) under appropriate incubation conditions. No evidence could be found for the formation of TxA 2-like activity from PG endoperoxides by the IBIM. In contrasts, when the IBIM were incubated with PGH 2 for 1 min at 22°C without cofactors, PGI 2-like activity was produced, causing profound relaxation of the isolated dog coronary artery preparation without contracting the rabbit aorta and inhibiting arachidonic acid-induced platelet aggregation. Equivalent quantities of boiled IBIM failed to aleter the biological activity of PGH 2 under identical conditions. Tranylcypromine (500 μg/ml) completely abolished the appearance of PGI 2-like activity. Furthermore, the PGI 2-like activity found was stable for 10 min at 22°C at pH 8.5 but completely lost under similar conditions at pH 5.5. It is concluded than microsomal preparations of normal bovine iris-ciliary body can synthesize PGI 2-like activity in substantial amounts but not TxA 2-like activity.

Effects of prostaglandin endoperoxide analogs on contractile elements in lung and gastrointestinal tract

Prostaglandin endoperoxides are formed in the lung as intermediate compounds in the biosynthesis of prostaglandins and thromboxanes. The effects of different doses of two analogs of prostaglandin endoperoxide PGH 2 were compared with those of PGF 2α and PGE 2 on superfused preparations of isolated trachea, bronchiole, peripheral lung, pulmonary artery and gastrointestinal smooth-muscle tissues. Endoperoxide analogs induced contraction of all smooth-muscle structures in the lung and airways. Compared to PGF 2α, analog I was approximately 71 times as potent on guinea-pig trachea, 214 times as potent on guinea-pig lung, and 57 times as potent on guinea-pig polmunary artery. Analog II was moderately less potent on all tissues than analog I. On gastrointestinal smooth-muscle organs, the PGH 2 analogs were generally closer in activity to PGF 2α and PGE 2, or even weaker. The findings show that PG endoperoxide vessels, and suggest that the naturally occurring compounds may participate in the mediation of bronchoconstriction and pulmonary vasoconstriction in disease states.

Polymorphonuclear leukocytes produce thromboxane A 2-like activity during phagocytosis

Homogenates of phagocytosing polymorphonuclear leukocytes obtained from rabbit peritoneum were incubated with the prostaglandin endoperoxides PGG 2 or PGH 2. After 2 min at 0°C, incubation mixtures contained an increased rabbit aorta contracting activity. Ether extracts of incubation mixtures contained a substance which contracted the superfused strips of rabbit aorta and coeliac artery and had a half life which was similar to thromboxane A 2. The generation of thromboxane A 2-like activity from PG endoperoxides was prevented by boiling the homogenate prior to incubation, or by pretreatment with benzydamine, a drug which blocks thromboxane formation in platelets. Production of thromboxane A 2-like material by leukocyte homogenates was compared with platelet microsomal thromboxane synthetase.

Synthesis of 9,11-desoxy-9,11-vinyleno-PGF2a and its diastereoisomer, analogs of the PG endoperoxide(PGH2)

Synthesis of 9,11-desoxy-9,11-vinyleno-PGF2a and its diastereoisomer, analogs of the PG endoperoxide(PGH2)

Further studies on prostaglandin E 2 (PGE 2)-induced gonadotropin release: Comparison with the effect of 15-methyl PGE 2, PGAs, PGBs and prostaglandin endoperoxide analogs

It is known that PGE 2 is a potent stimulus of LH release. To determine if the effect of PGE 2 could be enhanced and/or prolonged by retarding its metabolic degradation, a derivative, 15-methyl PGE 2 (15-E 2) which is more slowly degraded than the natural compound was injected intravenously (i.v.) at various dose levels or into the third ventricle (3rd V) of ether-anesthetized, ovariectomized, estrogen (OVX, Eb)-treated rats and its effect on gonadotropin release was compared with that of PGE 2. Both PGs injected i.v. were equally effective in increasing plasma LH and maintaining the elevated levels, although 15-E 2 induced a larger and more sustained increase in plasma FSH than PGE 2. By contrast, 3rd V PGE 2 was clearly more effective than 3rd V 15-E 2 in releasing LH and to a lesser extent, FSH. The effect of 15-E 2 on LH was similar to that produced by 3rd V PGE 1 injected at a similar dose. However, its effect on FSH was greater than that of PGE 1. To evaluate the effect(s) of prostaglandins of the A and B series on gonadotropin release, PGA 1, PGA 2, PGB 1 or PGB 2 were injected intraventricularly in OVX, Eb-treated rats. PGBs were injected into conscious, free-moving rats. PGA 2 or PGB 2 increased plasma LH concnetrations although much less effectively than PGE 2. Third V PGA 1 or PGB 1 were ineffective. The 3rd V injection of two cyclic esters (U-44069 and U-46619), stable analogs of the PG endoperoxide PGG 2 and PGH 2, induced a small, transient increase in LH levels and did not alter plasma FSH in conscious, free-moving animals. PGE 2 injected intraventricularly at a similar dose was demonstrated to be much more potent than the analogs in stimulating LH and FSH release. The results indicate that: 1) 15-E 2, in spite of its described long-lasting activity, does not appear to be more potent than the natural compound in releasing LH, although when injected i.v., it appeared to induce a more sustained increase in plasma FSH; 2) although PGA 2 and PGB 2 can also act centrally to stimulate LH release, their low potency suggests that this is a pharmacological effect; and 3) the two analogs of PG endoperoxides tested proved to be poor stimuli for gonadotropin release. The significance of these findings is discussed.