Studies on antiplatelet effects of OP-41483, a prostaglandin I2 analog, in experimental animals. II. Mechanism of its antiplatelet effect

The mechanism for the inhibition of platelet functions by a prostaglandin I2 analog, OP-41483, was studied with guinea pig platelets. OP-41483, and PGI2 as well, inhibited aggregation, ADP release and thromboxane formation of platelets with IC50 values of 4.3-5.8 ng/ml and 0.6-0.9 ng/ml, respectively. The ligand binding study using [3H]-OP-41483 suggested that OP-41483 bound with different affinities to two classes of binding sites on platelets. The dissociation constant of OP-41483 for the higher affinity site corresponded to the IC50 values of its antiplatelet effect. PGI2 as well as OP-41483 displaced [3H]-OP-41483 previously bound to platelets, thus indicating that both agents exerted their antiplatelet effects by binding to the same site on platelets. OP-41483 and PGI2 activated adenylate cyclase and raised cyclic AMP levels in platelets. However, their inhibitory effect on platelet aggregation was not fully antagonized by an adenylate cyclase inhibitor, 2',5'-dideoxyadenosine (DDA), at a concentration completely inhibiting the increase of cyclic AMP. Moreover, this DDA-resistant effect of OP-41483 disappeared in the presence of calcium chloride (10(-4)-10(-3) M). OP-41483 and PGI2 inhibited thrombin-induced Ca++ influx into platelets. The inhibition of Ca++ influx was not reversed by DDA. Based on these results, we speculate that the inhibitory effects of OP-41483 and PGI2 on platelet functions are produced through dual mechanisms: one mediated by activation of adenylate cyclase and the other by an inhibition of Ca++ influx; and these two mechanisms seem to be independent of each other.     

Studies on antiplatelet effect of OP-41483, a prostaglandin I2 analog, in experimental animals. I. Effect on platelet function and thrombosis

Antiplatelet and antithrombotic effects of OP-41483, a PGl2 analog, were studied in experimental animals, and the following results were obtained: 1) With 10 min-intravenous infusion to guinea pigs, OP-41483 inhibited platelet adhesiveness and platelet aggregation at 300-1000 ng/kg/min and 1000 ng/kg/min, respectively. In these effects, OP-41483 was 1-3 times more potent than carbacyclin and 3 times less potent than PGl2. 2) With oral administration to guinea pigs, OP-41483 given as its alpha-cyclodextrin clathrate (OP-41483 alpha-CD) inhibited platelet adhesiveness at doses higher than 1.0 mg/kg (expressed in terms of OP-41483), whereas PGl2 and carbacyclin did not at 10 mg/kg. OP-41483 alpha-CD also inhibited platelet aggregation after a single dose of 3 mg/kg and repeated doses of 3 mg/kg/day for 7 days. 3) In the electrically induced thrombosis model of guinea pig mesenteric artery, OP-41483 (300-1000 ng/kg/min, i.v.-infusion) and OP-41483 alpha-CD (1.0-3.0 mg/kg, p.o.) inhibited thrombus formation, but heparin (1.0-10 U/kg/min, i.v.-infusion) did not. 4) In the rabbit extracorporeal circulation thrombosis model, OP-41483 (100 and 300 ng/kg/min, i.v.-infusion) inhibited thrombus formation in the extracorporeal shunt and prevented the decrease in platelet count, hematocrit and fibrinogen level in circulating blood. Heparin (1.0-3.0 U/kg/min, i.v.-infusion) also inhibited the thrombus formation and the decrease in fibrinogen level, but did not inhibit the decrease in hematocrit and platelet count.     

Inhibitory effect of idebenone (CV-2619), a novel compound, on vascular lesions in hypertensive rats

The effects of a novel compound, 6-(10-hydroxydecyl)-2,3-dimethoxy-5-methyl-1,4-benzoquinone (idebenone, CV-2619), on cerebral and renal vascular changes were examined in stroke-prone spontaneously hypertensive rats (SHRSP) and in rats with experimentally induced hypertension. CV-2619 (35 mg/kg/day, p.o.) significantly inhibited the onset of cerebrovascular lesions (stroke) and the elevation of blood pressure in SHRSP with mild hypertension. A higher dose (2 X 50 mg/kg/day, p.o.) clearly delayed the onset of both stroke and proteinuria without any effect on the blood pressure in SHRSP with severe hypertension. In DOCA-salt hypertensive rats, CV-2619 (2 X 5 or 2 X 25 mg/kg/day, p.o.) dose-dependently inhibited decreases in body weight and water balance and the development of cerebral and renal vascular changes. These results suggest that CV-2619 inhibits the development of stroke and renal vascular lesions in hypertensive rats.     

Long-lasting antinociceptive effect of RC-160, a somatostatin analog, in mice and rats.

The effect of RC-160 (D-Phe-Cys-Tyr-D-Trp-Lys-Val-Cys-Trp-NH2) was studied in mice by means of the hot plate test and in rats with the tail flick test. In mice, 512 micrograms/kg (s.c.) induced an antinociceptive effect maximal (+88 +/- 17%) at the 6th hour after injection but still significant at the 24th hour; 16 micrograms/kg (s.c.) was active for 3 h; 64, 256 and 1024 micrograms/kg were active 30 min, 3 and 24 h after injection. The effects of these last three doses were not different. The antinociceptive action was as high at 24 h as at 3 h (e.g. for 512 micrograms/kg, 44 +/- 9% and 51 +/- 12%, respectively); 48 h after injection the scores of the treated groups were not different from the score for saline treatment in the tail flick test, 512 micrograms/kg induced a significant antinociceptive effect for 12 h (maximal at the 3rd hour: +60%). No behavioral changes, and particularly no motor activity modifications were observed.     

Inhibitory effect of cycloheximide on gastric secretion in rats.

Cycloheximide administered to rats in non-toxic doses immediately after pyloric ligation results in a lower incidence of experimental gastric ulcers. The effect is paralleled by an impaired secretion of hydrochloric acid (more than 90 per cent inhibition). Simultaneously the activity of pepsin in gastric juice and in homogenates of gastric mucosa was reduced by 60 per cent and mucoproteins were lowered by 30 per cent. The basal and pentagastrin-stimulated secretion were inhibited by cycloheximide to the same extent.     

The in vitro and ex vivo antiplatelet effect of TRK-100, a stable prostacyclin analog, in several species

Effect of TRK-100, a stable PGI2 analog, on platelet function was tested in vitro and ex vivo. TRK-100 at the dose range of 0.5-300 nM inhibited platelet aggregation induced by arachidonic acid, adenosine 5'-diphosphate and collagen in several species including human platelets. The potency of TRK-100 was 1/2 to 1/5 that of PGI2. The effect was strong in human and cat platelets. In conscious rabbits and rats, oral TRK-100 at the dose range of 0.1-1 mg/kg inhibited ex vivo platelet aggregation up to 80% in the rat and 70% in the rabbit, and the effect lasted over 5 hr. However, in both species, the effect on blood pressure was minimal. In anesthetized rabbits, inhibition of platelet aggregation was the same level as in the conscious animal, but blood pressure depression was observed. Cyclic AMP levels of human platelets, 2 min after incubation, was elevated up to 2.4 microM/10(9) platelets by 100 ng/ml of PGI2 and 1.5 microM by 100 ng/ml of TRK-100. It was shown that TRK-100 has a potent antiplatelet effect both in vitro and ex vivo in many species through elevation of platelet cAMP. These results suggest that TRK-100 may be a potential oral antithrombotic drug.     

Inhibitory effect of tin on intestinal calcium absorption in rats.

The effect of tin on intestinal calcium absorption was studied in rats given stannous chloride (Sn, 30 mg/kg every 12 hr) orally for 3 days. The increase of serum calcium concentration after a single oral administration of calcium chloride (Ca, 500 mg/kg) was significantly inhibited by the prior administration of tin. Calcium concentrations in duodenal mucosa were also reduced to one-third of control values by the administration of tin. The calcium-binding activity in the supernatant fraction of 38,000g of the mucosal homogenate was decreased to one-quarter of control values, and alkaline phosphatase activity in the mucosal homogenate was markedly lowered by the administration of tin. These results suggest that the oral administration of tin inhibited the duodenal active transport of calcium.     

Inhibitory effect of curcumin, an anti-inflammatory agent, on vascular smooth muscle cell proliferation.

The effects of curcumin, an anti-inflammatory agent from Curcuma longa, on the proliferation of blood mononuclear cells and vascular smooth muscle cells were studied. Proliferative responses were determined from the uptake of tritiated thymidine. In human peripheral blood mononuclear cells, curcumin dose dependently inhibited the responses to phytohemagglutinin and mixed lymphocyte reaction at the dose ranges of 10(-6) to 3 x 10(-5) and 3 x 10(-6) to 3 x 10(-5) M, respectively. Curcumin (10(-6) to 10(-4) M) dose dependently inhibited the proliferation of rabbit vascular smooth muscle cells stimulated by fetal calf serum. Curcumin had a greater inhibitory effect on platelet-derived growth factor-stimulated proliferation than on serum-stimulated proliferation. Cinnamic acid, coumaric acid and ferulic acid were much less effective than curcumin as inhibitors of serum-induced smooth muscle cell proliferation, suggesting that the cinnamic acid and ferulic acid moieties alone are not sufficient for activity, and that the characteristics of the diferuloylmethane molecule itself are necessary for activity. Curcumin may be useful as a new template for the development of better remedies for the prevention of the pathological changes of atherosclerosis and restenosis.     

Inhibitory effect of ethanol on endothelium-dependent vascular responsiveness

The effect of ethanol was studied on the endothelium-dependent vascular responses in isolated rat aortic strips. Ethanol depressed the endothelium-dependent relaxation induced by acetylcholine and ATP but not that induced by the calcium ionophore, A23187. Endothelium-independent relaxation in response to sodium nitroprusside, a soluble guanylate cyclase activator, was not depressed by ethanol. On the other hand, ethanol significantly enhanced the contractile response to clonidine, an alpha 2-adrenoceptor agonist, in endothelium-intact strips and depressed it in endothelium-denuded strips. These results suggest that ethanol can inhibit endothelium-dependent relaxation by acting on endothelial cells but not on smooth muscle cells, and can also suppress an inhibitory effect of the endothelium on alpha 2-adrenoceptor-mediated vasoconstriction.     

阿藿烯对大鼠抗血栓活性作用研究

目的:研究阿藿烯对大鼠血栓形成和凝血功能的影响,评价其抗血栓活性。方法:将SD大鼠随机分为空白对照组、血塞通阳性药组(50mg/kg)、阿藿烯低剂量组(25mg/kg)、中剂量组(50mg/kg)、高剂量组(75mg/kg),各组动物连续灌胃5d,末次给药2h后建立下腔静脉结扎模型、FeCl3致动脉血栓模型、断尾流血时间测定模型测定阿藿烯对血栓形成和凝血的影响;并测定各组大鼠给药前及给药后2h血浆凝血酶原时间(PT)、活化部分凝血酶时间(APTT)。结果:阿藿烯能抑制动静脉血栓的形成,使血栓重量降低,且高剂量阿霍烯对动静脉血栓形成的抑制率均大于40%;同时有效延长大鼠断尾流血时间,并能使大鼠血浆APTT、PT明显延长(P<0.05),且都具有剂量依赖关系。结论:阿藿烯有抗血栓活性。     

Inhibitory effect of a new steroidal saponin,OSW-1, on ovarian functions in rats

1. This study was undertaken to determine the effects of OSW-1 (3β, 16β, 17α-trihydroxycholest-5-en-22-one 16-O-(2-O-4-methoxybenzoyl-β-D-xylopyranosyl)-(1→3)-(2-O-acetyl-α-L-arabinopyranoside)) on the pituitary-ovarian system and the functions of aortic smooth muscle.
2. A single s.c. injection of OSW-1 (9 μg kg⁻¹) on the morning of pro-oestrus inhibited the occurrence of the expected next pro-oestrus, whereas administration of OSW-1 at a dose of 4.5 μg kg⁻¹ did not affect the oestrous cycle. OSW-1 treatment on the day of dioestrus-l did not affect the oestrous cycle.
3. At doses of 4.5 and 9 μg kg⁻¹ OSW-1, the serum oestradiol (E₂) levels at the expected next pro-oestrus were significantly lower than in control (pro-oestrus). The serum luteinizing hormone (LH) levels 4 days after 9 μg kg⁻¹ OSW-1 treatment were also markedly lower than those of control. OSW-1 (4.5 μg kg⁻¹) did not affect the levels of inhibin, progesterone and gonadotrophins on the same day.
4. OSW-1 did not inhibit the preovulatory LH surge which occurs on the afternoon of pro-oestrus day.
5. The expression of mRNA coding for the cholesterol side chain cleavage cytochrome P-450 (p450scc), an ovarian steroidal limiting enzyme, was suppressed at 24 and 96 h after OSW-1 treatment.
6. Administration of OSW-1 (9 μg kg⁻¹) tended to reduce the relaxation of isolated thoracic aorta ring preparations induced by acetylcholine, while there was no difference in the relaxation induced by sodium nitroprusside.
7. Our results show that OSW-1 inhibits ovarian E₂ secretion and that the decrease in E₂ secretion may contribute to its effects on the oestrous cycle and the sensitivity of the thoracic aorta to relaxation. The decrease in the levels of ovarian steroids induced by OSW-1 may be due to its direct inhibitory action on the gene expression of the steroidal enzyme and on the proliferation of granulosa cells in the ovary.

     

Inhibitory effect of indomethacin farnesil, a novel antiinflammatory prodrug, on carrageenin-induced inflammation in rats

Antiinflammatory effects of indomethacin farnesil (IMF), a novel prodrug of indomethacin, was examined after both oral and local administration. In the air pouch carrageenin-induced inflammation, an oral dose of IMF exerted dose dependent inhibitory effects on the accumulation of inflammatory exudate fluid and the migration of leukocytes into the exudate. Both the increased vascular permeability and the prostaglandin E2 levels in the exudate fluid were reduced by IMF. Significant levels of free indomethacin were detected in the pouch fluid. In spite of the inability of IMF to inhibit prostaglandin synthesis in a cell free cyclooxygenase system, IMF injected locally inhibited carrageenin paw edema, and the inhibitory effect was comparable to that of indomethacin itself. When injected locally into the paw together with carrageenin, 14C-IMF was effectively converted to its active metabolite, indomethacin. The indomethacin concentration in the paw tissue was comparable to that of indomethacin injected paws with the same molar dose of free indomethacin.     

Salutary effects of CG-4203, a novel, stable prostacyclin analog, in hemorrhagic shock

Prostacyclin (PGI2) is a potent vasodilator, an inhibitor of platelet aggregation, and a membrane-stabilizing agent that has been shown to exert beneficial effects in a variety of models of ischemia and circulatory shock. However, the use of PGI2 is limited by its instability and rapid biodegradation. We studied the effects of a novel, stable prostacyclin analog, CG-4203, in a murine model of hemorrhagic shock. Hemorrhaged rats treated with CG-4203 maintained postreinfusion mean arterial blood pressure (MABP) at significantly higher values than rats receiving only the vehicle (final MABP 101 +/- 3 vs. 75 +/- 5 mm Hg, p less than 0.01). CG-4203 was also found to attenuate the increase in plasma cathepsin D activity (p less than 0.01), as well as the plasma accumulation of free amino-nitrogen compounds (p less than 0.05). Furthermore, the plasma activity of a myocardial depressant factor (MDF) was significantly lower in CG-4203-treated hemorrhaged rats than in rats receiving the vehicle (25 +/- 2 vs. 54 +/- 7 U/ml, p less than 0.01). In addition, CG-4203 exerted an anti-proteolytic action in pancreatic homogenates and inhibited platelet aggregation in platelet-rich plasma. However, CG-4203, at concentrations expected during treatment of shock, failed to have an immediate or delayed vasodilator effect in rat aortic rings, and thus vasodilation is not an important aspect of the antishock effects of CG-4203. Our results suggest that inhibition of platelet aggregation, as well as the antiproteolytic and membrane-stabilizing actions, could mediate the beneficial effects of CG-4203 in hemorrhagic shock.     

The effect of nifedipine,nimodipine and nisoldipine on agonistand trauma-stimulated vascular prostacyclin synthesis in vitro

Summary The present study shows that nifedipine, nimodipine and nisoldipine inhibit in vitro agonist-induced prostacyclin (PGI₂) release from rat aortic rings. The agonists used were: U46619 (a thromboxane A₂ analogue); A23187 (a calcium ionophore) and adrenaline.In contrast, these calcium channel blockers did not inhibit in vitro trauma-or arachidonic acid (AA)-induced PGI₂ release. Therefore, in vitro PGI₂ release, models may be calcium channel dependent (adrenaline, U46619, A23187) or independent (trauma, AA), a property which is relevant to calcium channel blocker research. It is suggested that calcium channel dependent PGI₂ release is relevant to modulating the relaxation phase of muscle contraction, while calcium channel independent PGI₂ release is relevant to limiting the extension of thrombi following local vascular trauma.     

利洛司酮对大鼠异位子宫内膜的抑制作用(英文)

目的 :观察利洛司酮对大鼠异位子宫内膜的抑制作用。方法 :用外科自体移植术建立大鼠子宫内膜异位症模型。灌胃给药 ,qd× 2 1d ,测量给药前、后异位内膜体积 ,计算药物对异位内膜的抑制率。结果 :用药后对照组大鼠异位子宫内膜比用药前略增大 ;给予利洛司酮 2 0 ,80mg·kg- 1和米非司酮 50mg·kg- 1后 ,异位内膜的体积比用药前明显缩小 ,抑制率依次为 (80± 2 7) % ,(88± 2 2 ) %和 (82± 31) %。结论 :利洛司酮和米非司酮一样明显抑制大鼠异位子宫内膜的生长     

Beneficial effects of a new prostacyclin analogue, KP-10614, on acute myocardial infarction in rats.

The potential therapeutic value of a new prostacyclin analogue, (4z, 16s)-4,5,18,18,19,19-hexadehydro-16,20-dimethyl-delta 6(9a)- 9-(O)-methano-PGI1 (KP-10614), was studied in acute myocardial infarction in rats. Myocardial infarction was induced by ligation of the left coronary artery and ischemic injury was followed up to 4 h. The infarct size, evaluated by the area unstained by 2,3,5-triphenyltetrazolium chloride, reached 41.1 +/- 1.4% of the left ventricle at 4 h. KP-10614 (3 ng/kg/min x 4 h) reduced the infarct size at 4 h significantly (26.5 +/- 2.9%). At the same dose, KP-10614 inhibited ADP-induced ex vivo platelet aggregation significantly (21.5 +/- 4.0% of the control aggregation), but did not alter the arterial blood pressure or heart rate. To assess the role of platelets in myocardial infarction, circulating platelets were reduced by about 95% with rabbit antiserum to rat platelets. In platelet-depleted rats, the infarct size decreased significantly to 24.1 +/- 4.6% of the left ventricle at 4 h. These results suggest that platelets play an important role in expression of myocardial ischemic injury resulting from coronary artery occlusion in rats, and the ability of KP-10614 to decrease the infarct size appeared to be attributable, at least in part, to the inhibition of platelet aggregation or cellular metabolic effects produced by platelets at the site of tissue injury.     

Inhibitory effect of oxcarbazepine on high-frequency firing in peripheral nerve fibers

We assessed the effects of oxcarbazepine, an antiepileptic derivative of carbamazepine, on discharges in single cutaneous afferent fibers produced by repetitive high-frequency stimulation (mimicking the abnormal excitation of peripheral nerves in neuropathic pain and paresthesia). After intravenous administration of oxcarbazepine, the later responses in the train dropped out without the earlier ones being affected and, thus, the total number of spikes decreased. The latency of the responses to an individual pulse was unchanged. These results, which indicate that oxcarbazepine inhibits the generation of high-frequency firing without affecting impulse conduction, suggest that this drug may be useful against neuropathic pain and paresthesia.     

Role of sulfhydryl groups in the stimulatory effect of captopril on vascular prostacyclin synthesis

The effect of captopril on vascular prostacyclin production was studied, evaluating which of its components--sulfhydryl (SH) group or proline--is responsible for this effect. Rat aortas were incubated with captopril (10-100 microM), 2-mercaptoethanol or proline (10 microM), and captopril plus the SH-binding reagents N-ethylmaleimide or ethacrynic acid (50 microM). Prostacyclin was measured by radioimmunoassay of 6-keto-prostaglandin F1 alpha. Captopril stimulated prostacyclin production. This effect was associated with an enhanced conversion of arachidonate to prostacyclin and was not related to bradykinin. Since 2-mercaptoethanol increased vascular prostacyclin per se and proline did not, the stimulatory effect of captopril appears to be dependent upon the SH group; in addition, both SH blockers, N-ethylmaleimide and ethacrynic acid, antagonized this effect. This study shows that captopril stimulates vascular prostacyclin synthesis directly and that the SH group plays a key role in this action. This stimulation of prostacyclin synthesis may contribute to the antihypertensive action of captopril.     

Inhibitory effect of AD-1590, a non-steroidal anti-inflammatory drug, on allergic inflammation in mice and rats

Effect of AD-1590 on allergic inflammations was investigated. AD-1590 and indomethacin at an oral dosage as high as 32 mg/kg did not show any significant inhibitory activity on rat passive cutaneous anaphylaxis, a type-I allergy, although prednisolone and cyproheptadine produced strong inhibition. Against rat adjuvant arthritis, type-III and -IV allergies, AD-1590 showed potent prophylactic (2 and 4 mg/kg/day) and therapeutic (0.4-1 mg/kg/day) effects when given orally once a day for 3 weeks beginning from just before and for 1 week starting from 14 to 18 days after adjuvant inoculation, respectively; however, its prophylactic and therapeutic potencies were about one-fourth and one-fifth, respectively, that of indomethacin. The arthritis was strongly inhibited with prophylactic treatment of prednisolone (1 mg/kg/day) or cyproheptadine (40 mg/kg/day). On the other hand, prednisolone (ED50 = 0.0119 mg/ear, topical) showed strong activity in inhibiting mouse contact hypersensitivity to oxazolone (ear edema), a type-IV allergy, but cyproheptadine only had weak activity. AD-1590 (0.318 mg/ear) and indomethacin (0.699 mg/ear) produced rather strong inhibition; in particular, AD-1590 produced almost complete inhibition at high dosages, whereas most of the non-steroidal anti-inflammatory drugs (NSAID) tested showed weak inhibition or a partial inhibition of about 50% even at the highest dosage. The oral potency of AD-1590 was about 2 and 100 times those of indomethacin and ibuprofen, respectively. These results demonstrate that in allergic inflammation, the pharmacological properties of AD-1590 are somewhat different from those of other NSAID and different from those of prednisolone and cyproheptadine.