Feng Nien Ko

Inhibition of platelet aggregation by some flavonoids

The inhibitory effects of five flavonoids on the aggregation and secretion of platelets were studied. These flavonoids inhibited markedly platelet aggregation and ATP release of rabbit platelets induced by arachidonic acid or collagen, and slightly those by platelet-activating factor. ADP-induced platelet aggregation was also suppressed by myricetin, fisetin and quercetin. The IC50 on arachidonic acid-induced platelet aggregation was: fisetin, 22 microM; kaempferol, 20 microM; quercetin, 13 microM; morin, 150 microM less than IC50 less than 300 microM. The thromboxane B2 formations were also inhibited by flavonoids in platelets challenged with arachidonic acid. Fisetin, kaempferol, morin and quercetin antagonized the aggregation of washed platelets induced by U46619, a thromboxane A2/prostaglandin endoperoxides mimetic receptor agonist. In human platelet-rich plasma, quercetin prevented the secondary aggregation and blocked ATP release from platelets induced by epinephrine or ADP. These results demonstrate that the major antiplatelet effect of flavonoids tested may be due to both the inhibition of thromboxane formation and thromboxane receptor antagonism.

Clinical implications and factors related to left atrial spontaneous echo contrast in chronic nonvalvular atrial fibrillation

The mechanisms leading to formation of spontaneous echo contrast (SEC), a smoke-like echo on echocardiography, are still controversial. To further explore the clinical implications and factors related to SEC formation, the correlation among echocardiographic variables, hematologic parameters or platelet aggregability, and the occurrence of SEC was studied in 119 patients with chronic nonvalvular atrial fibrillation. There were 75 men and 44 women with a mean age of 65 +/- 10 years (range 38-88). Left atrial SEC was detected in 39 patients (33%) by transesophageal echocardiography. Patients with history of systemic embolism were more frequently found to have left atrial SEC and left atrial thrombus by univariate analysis. Multivariate analysis showed that left atrial SEC (p < 0.001) was the only independent predictor of history of systemic embolism. Age, sex, left atrial or left ventricular dimension, left ventricular ejection fraction, antiplatelet or anticoagulant therapy and the percentage of lone atrial fibrillation were not significantly different between patients with and without left atrial SEC. Among the hematologic parameters, higher hematocrit was found in patients with left atrial SEC, while white blood cell and platelet counts were comparable in both groups. Platelet aggregability with different concentrations of inducers, adenosine diphosphate and collagen, was evaluated by the turbidimetric method in 15 patients with left atrial SEC and in 42 patients without left atrial SEC who were not receiving antiplatelet or anticoagulant therapy. No significant difference was found in platelet aggregability using four inducer concentrations between two groups of patients.(ABSTRACT TRUNCATED AT 250 WORDS)

Vasorelaxation of rat thoracic aorta caused by osthole isolated from Angelica pubescens.

The pharmacological effects of osthole on isolated rat thoracic aorta were examined. Osthole inhibited norepinephrine (NE, 3 microM)-induced phasic and tonic contractions in rat thoracic aorta in a concentration-dependent manner (40-200 microM). The tonic contraction elicited by NE was also relaxed by the addition of osthole. This relaxing effect of osthole was not affected by indomethacin (20 microM) and was still observed in endothelium-denuded rat aorta. Methylene blue (50 microM) partially antagonized this relaxing effect of osthole. In high-K+ medium (80 mM), the Ca2+ (0.03-3 mM)-induced vasocontraction was inhibited concentration dependently by osthole (20-100 microM). Addition of osthole (100 microM) at the plateau of the K+ (80 mM)-induced contraction caused relaxation. Methylene blue (50 microM) did not antagonize this relaxation. In Ca(2+)-free medium, the caffeine (10 mM)-induced phasic contraction was also suppressed by osthole in a concentration-dependent manner. Although the cAMP level was not changed by osthole, the cGMP level of rat aorta was increased by osthole in a concentration-dependent manner. The increase in cGMP level caused by osthole was completely blocked by methylene blue. [3H]Inositol monophosphate formation caused by NE was not affected by osthole at a concentration of 200 microM. The 45Ca2+ influx elicited by either NE or high K+ was inhibited by osthole in a concentration-dependent manner. It is concluded that osthole relaxes rat thoracic aorta by virtue of its Ca(2+)-channel blocking properties and by elevating cGMP levels in vascular smooth muscle.

The relaxant action of osthole isolated from Angelica pubescens in guinea-pig trachea

The effect of osthole, isolated from Angelica pubescens, on the contraction of guinea-pig trachea was studied. Osthole (25–100 μmol/l), theophylline (10–1000 μmol/l) and higher concentrations of nifedipine (0.1–100 μmol/l) suppressed the contraction response curves of tracheal smooth muscle caused by carbachol, prostaglandin F2α (PGF2α), U46619 (thromboxane A2 analogue) and leukotriene C4 (LTC4) in a concentration-dependent manner. The contraction caused by high K+ (120 mmol/1) and cumulative concentrations of CaCl2 (0.03–3 mmol/1) was also inhibited concentration-dependently by osthole (25–100 μmol/l), theophyl line(10–1000 μmol/l) and lower concentrations of nifedipine (0.01–0.1 μmol/l). The relaxant actions of osthole were not affected by propranolol (1 μmol/l), glibenclamide (10 μmol/l) or removal of tracheal epithelium. Osthole (100 μmol/l) was still effective in causing tracheal relaxation in the presence of nifedipine (1 μmol/l). In Ca2+-free- and EGTA (0.2 mmol/1)-containing medium, the relaxing effect of osthole was more potent than in normal Krebs solution. Osthole (25 and 50 μmol/l) caused 2.9 and 6.5, or 3.0 and 5.6 fold, respectively, increase in potency of forskolin or sodium nitroprusside in causing tracheal relaxation but did not affect that by cromakalim. Osthole (50 μmol/l) enhanced the increase in tissue cAMP and cGMP levels induced by forskolin and sodium nitroprusside, respectively, and in higher concentrations (100 and 250 μmol/l), itself increased markedly tissue cAMP and cGMP contents. Osthole (10–250 mol/l) inhibited the activity of cAMP and cGMP phosphodiesterases in a concentration-dependent manner. It is concluded that osthole exerts a nonspecific relaxant effect on the trachealis by inhibiting the cAMP and cGMP phosphodiesterases.

Vasorelaxing effect in rat thoracic aorta caused by fraxinellone and dictamine isolated from the Chinese herb Dictamnus dasycarpus Turcz: comparison with cromakalim and Ca2+ channel blockers

SummaryThe components of Dictamnus dasycarpus Turcz were tested for their vasorelaxing effect on the rat aorta, and fraxinellone and dictamine were shown to be effective vasorelaxants. In high K+ (60 mmol/l) medium, Ca2+ (0.03 to 3 mmol/l)-induced vasoconstriction was inhibited concentration-dependently by both agents. The IC50 for fraxinellone and dictamine were calculated to be about 25 μmol/l and 15 μmol/l (for Ca2+) concentration of (1 mmol/l), respectively. Cromakalim (0.2–10) μmol/l relaxed aortic rings precontracted with 15 but not 60 mmol/l of K+. Fraxinellone and verapamil were more potent and effective in producing relaxation in 60 mmol/l than in 15 mmol/l K+-induced contraction. However, dictamine was more potent in producing relaxation in 5 mmol/l K+-induced contraction. Nifedipine (1 μmol/l), dictamine (100 μmol/l) and fraxinellone (100 μmol/l) relaxed the aortic contraction caused by KCl or Bay K 8644. The tonic contraction elicited by nor adrenaline (NA, 3 μmol/l) was also relaxed by dictamine (500 μmol/l), but not by fraxinellone (500 μmol/l) in the nifedipine (1 μmol/l)-treated aorta. This relaxing effect of dictamine persisted in endothelium-denuded aorta. Glibenclamide (10 μmol/l) shifted the concentration-relaxation curve of cromakalim, but not that of dictamine, to the right in rat aortic rings precontracted with NA. Dictamine (500 μmol/l) did not affect tonic contraction of NA which are reduced by H-7 (1 μmol/l) in Ca2+ depleted medium. In conclusion, fraxinellone is a selective blocker of voltage-dependent Ca2+ channel, while dictamine relaxed the rat aorta by suppressing the Ca2+ influx through both voltage-dependent and receptor-operated Ca2+ channels.

PAF antagonism in vitro and in vivo by aglafoline from Aglaia elliptifolia Merr

Aglafoline, isolated from Aglaia elliptifolia Merr, inhibited in a selective and concentration-dependent manner the aggregation and ATP release reaction induced in washed rabbit platelets by PAF (platelet-activating factor). The IC50 values of aglafoline, BN52021 and kadsurenone on PAF (3.6 nM)-induced platelet aggregation were about 50, 12 and 18 microM, respectively. Aglafoline also inhibited [3H]PAF (3.6 nM) binding to washed rabbit platelets with an IC50 value of 17.8 +/- 2.6 microM. The concentration-response curve of PAF-induced platelet aggregation was shifted to the right by aglafoline with pA2 and pA10 values of 5.97 and 5.04, respectively. Although thromboxane B2 formation caused by collagen and thrombin was partially suppressed by aglafoline, thromboxane B2 formation caused by ionophore A23187 and arachidonic acid was not affected. Aglafoline inhibited the [3H]inositol monophosphate formation caused by PAF but not that caused by collagen or thrombin in the presence of indomethacin (20 microM). The cAMP content of washed rabbit platelets was not affected by aglafoline. Rat femoral intravenous administration of aglafoline (10 mg/kg) did not affect blood pressure. However, aglafoline (10 mg/kg) both prophylactically and therapeutically antagonized PAF (2.5 micrograms/kg)-induced hypotensive shock in rats. Intravenous PAF (30 ng/kg) caused severe bronchoconstriction in guinea pigs. This effect was completely blocked by aglafoline. This implies aglafoline is an effective PAF antagonist not only in vitro, but also in vivo.

Cinnamophilin, a novel thromboxane A2 receptor antagonist, isolated from Cinnamomum philippinense

The pharmacological activity of cinnamophilin ((8R,8S)-4,4-dihydroxy-3,3-dimethoxy-7-oxo-8,8-neolignan), isolated from Cinnamomum philippinense, was studied in isolated rat aorta, guinea-pig trachea and rabbit platelets. Cinnamophilin was found to be a thromboxane A2 receptor blocking agent in these tissues as revealed by its competitive antagonism of the U-46619 (9,11-dideoxymethanoepoxy-9 alpha,11 alpha-prostaglandin F2 alpha)-induced contraction of rat aorta and guinea-pig trachea and aggregation of rabbit platelets with pA2 values of 7.3 +/- 0.2, 5.2 +/- 0.1 and 6.3 +/- 0.3, respectively. Protection against the irreversible vasoconstriction of rat aorta caused by U-46619 (0.05 microM) was obtained by cinnamophilin (10 microM) but not by caffeine (25 mM). Cinnamophilin (1-15 microM) also possessed voltage-dependent Ca2+ channel blocking action, judging from its antagonism of the high K+ (60 mM)- and Bay K 8644 (0.1 microM)-induced contraction in rat thoracic aorta. Cinnamophilin (30 microM) produced a slight relaxation of noradrenaline (3 microM)-induced tonic contractions, and this relaxing effect was abolished in the presence of nifedipine (1 microM). Nifedipine (10 microM) sufficient to inhibit high K(+)-induced contractions failed to attenuate the contractile response to U-46619. A high concentration of cinnamophilin (100 microM) did not affect the aortic contraction induced by endothelin-1, angiotensin II, carbachol or serotonin. Neither cAMP nor cGMP in rat aorta was increased by cinnamophilin. These results indicate that cinnamophilin is a selective thromboxane A2 receptor antagonist especially in rat aorta, and also possesses voltage-dependent Ca2+ channel blocking properties.

Antiplatelet effects of clausine-D isolated from Clausena excavata

Clausine-D inhibited concentration-dependently the aggregation and release reaction of washed rabbit platelets induced by arachidonic acid and collagen, without affecting those induced by U46619, PAF and thrombin. The IC50 values of clausine-D on arachidonic acid- and collagen-induced platelet aggregation were calculated to be 9.0 +/- 1.1 and 58.9 +/- 0.9 microM, respectively. Thromboxane B2 and prostaglandin D2 formation in platelets caused by arachidonic acid were also suppressed. Clausine-D inhibited increased intracellular concentration of calcium in platelets caused by arachidonic acid and collagen, and also abolished the generation of inositol monophosphate caused by arachidonic acid, but not that by collagen, U46619, PAF and thrombin. In human citrated platelet-rich plasma, clausine-D inhibited the secondary phase, but not the primary phase, of aggregation induced by epinephrine and ADP. These results indicate that the antiplatelet effect of clausine-D is due to inhibition of the formation of thromboxane A2.

Antiplatelet effects of protopine isolated from Corydalis tubers

Protopine inhibited the aggregation and ATP release of rabbit platelets induced by ADP, arachidonic acid, PAF, collagen and ionophore A23187. Although the platelet aggregation caused by thrombin was not inhibited by protopine (100 micrograms/ml), the release reaction was partially suppressed. In rabbit platelet-rich plasma, protopine also inhibited the platelet aggregation caused by ADP, arachidonic acid, PAF and collagen. The thromboxane B2 formation of washed platelets caused by arachidonic acid, collagen, ionophore A23187 and thrombin was suppressed by protopine. Protopine inhibited the intracellular calcium increase caused by arachidonic acid in quin-2/AM loaded rabbit platelets. In the presence of indomethacin, the intracellular calcium increase caused by collagen and PAF was completely suppressed by protopine, and the intracellular calcium increase caused by thrombin was partially inhibited. The phosphoinositides breakdown caused by collagen and PAF was inhibited by protopine, but that by thrombin was not affected significantly. Protopine did not cause the elevation of cyclic AMP level of platelets. It is concluded that the antiplatelet effects of protopine is due to inhibition on thromboxane formation and phosphoinositides breakdown and then lead to the decrease of intracellular calcium concentration.

The relaxant actions on guinea-pig trachealis of atherosperminine isolated from Fissistigma glaucescens

The pharmacological activity of atherosperminine, isolated from Fissistigma glaucescens, was determined in isolated guinea-pig trachealis. Atherosperminine (25-100 microM) and theophylline (10-1000 microM) both inhibited the contractile response caused by carbachol, prostaglandin F2 alpha (PGF2 alpha), U46619 (thromboxane A2 analogue), leukotriene C4 (LTC4) and Ca2+ (in the presence of 120 mM KCl) in a concentration-dependent manner. The inhibition was characterized by a rightwards shift of the concentration-response curves with suppression of the maximal contraction. Propranolol (1 microM), glibenclamide (10 microM) and removal of tracheal epithelium did not modify the relaxant action of atherosperminine. Atherosperminine (25 and 50 microM) caused a 2.4- and 5.0-fold, respectively, potentiation of the action of forskolin to cause tracheal relaxation but did not potentiate the action of sodium nitroprusside or cromakalim. Atherosperminine (50 microM) potentiated the action of forskolin to increase tissue cAMP content and, in higher concentrations (100 and 250 microM), itself increased tissue cAMP but not cGMP content. Atherosperminine markedly inhibited cAMP phosphodiesterase but not cGMP phosphodiesterase in homogenates of guinea-pig trachealis. It is concluded that atherosperminine exerts a non-specific relaxant effect on the trachealis. Its major mechanism of action appears to be inhibition of cAMP phosphodiesterase, perhaps with a minor effect on cGMP phosphodiesterase at higher concentrations.