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A Zatta

Publications and source records attributed to A Zatta.

At least 19 recordsLinked to original sources
PubMed1999

Differential inhibition of polymorphonuclear leucocyte functions by cloricromene.

Leucocytes play an essential role in the pathogenesis of ischaemia and reperfusion injury and inhibition of their adhesion and of mediator release can reduce vascular and tissue damage. Previous studies have shown that cloricromene modifies several granulocyte as well as monocyte/macrophage functions and it has been shown that cloricromene administration exerts a clear protective action in several experimental models of ischaemia. The present work describes new data on polymorphonuclear leukocyte (PMN) inhibition exerted by cloricromene and compares these observations with earlier ones. Human washed PMN and human whole blood (HWB) were studied in vitro upon stimulation with f-MLP in the presence of cytochalasin B, with opsonized zymosan and with a phorbol ester (PMA). Amongst free radicals, superoxide anions were chosen as index of oxidative burst. Phagocytosis and beta-glucuronidase, as lysosomal release indicators, were measured to characterize PMN function: cloricromene inhibited concentration-dependently all the parameters upon stimulation by each activator tested. Experiments performed in rabbit whole blood (RWB) showed that cloricromene inhibited free radical generation with IC50 values similar to those obtained in human whole blood. Comparing the action of cloricromene on human cells in different tests, we found that some parameters were more sensitive than others, even when the same stimulus was used. In particular, free radical generation was inhibited by cloricromene with IC50 values below 36 microM, while other functions, like lysosomal release and phagocytosis were inhibited with IC50 values over 100 microM. These data confirm that cloricromene exerts a notable inhibitory effect on PMN and may explain the activity of the compound, observed in vivo in several experimental models of ischaemia-reperfusion and shock.

Animals↗
PubMed1993

Platelet aggregation induced by the endoperoxide analogue U46619 is inhibited by polymorphonuclear leukocyte ADPase activity.

Platelet activation by the stable endoperoxide analogue U46619 is mediated largely by ADP released from platelet-dense granules. Polymorphonuclear leukocytes (PMNs) endowed with ecto-ADPase activity may operate as antiaggregatory cells in platelet aggregation induced by U46619. Unstimulated PMNs were effective in reducing aggregation when platelets were stimulated by threshold concentrations of U46619, whereas at higher concentrations of the stimulus, PMN activation is required. Evidence that the inhibition was mediated by PMN ecto-ADPase activity was obtained by high-performance liquid chromatography analysis, indicating that PMNs were able to efficiently metabolize platelet-active ADP into AMP. Moreover, PMN-derived supernatants were able to inhibit platelet aggregation, suggesting that under this circumstance the inhibition was exerted by an uncharacterized, releasable ADPase activity. This study supports the hypothesis that, besides nitric oxide and hydrogen peroxide, ADPase activity may represent another PMN-mediated pathway capable of regulating platelet activity in areas of reduced blood flow, such as those found in conditions of myocardial ischemia.

Adenosine Diphosphate↗
PubMed1993

Hematologic and biochemical profiles of selectively bred WHHL rabbits.

In Watanabe heritable hyperlipidemic (WHHL) rabbits, hypercholesterolemia and hypertriglyceridemia develop from birth, because of a deficiency of low-density lipoprotein receptors, and are followed by a consequent early development of aortic atherosclerosis. This closely resembles human familial hypercholesterolemia. Starting in 1984, we have developed a closed colony by breeding two male and two female homozygous WHHL rabbits, obtained from Japan (Dr. Watanabe, Kobe University). In our facility, the application of a selective breeding program, strictly based on mating parents that both have high serum lipid concentrations, has produced markedly elevated cholesterol (701 +/- 172 mg/dl, mean +/- SD) and triglyceride (780 +/- 325 mg/dl) concentrations in weaning rabbits. Clinical chemical analysis revealed no kidney or liver function abnormalities even in animals with extremely high lipid concentrations, and hematologic profiles were very similar in WHHL and age-matched New Zealand White rabbits, with the exception of platelet count, which was significantly higher in WHHL rabbits. Platelet aggregation induced by collagen and platelet-activating factor was significantly reduced in WHHL rabbits, whereas thrombin and prothrombin times appeared normal when compared with those in New Zealand White rabbits.

Animal Husbandry↗
PubMed1991

Polymorphonuclear leukocyte-dependent modulation of platelet function: effect of cloricromene.

Cloricromene inhibits the activation of platelets and polymorphonuclear leukocytes (PMN) induced by specific stimuli. We report here that cloricromene caused a concentration-dependent reduction of platelet aggregation, beta-thromboglobulin (beta-TG) release and cytoplasmic Ca2+ movements induced by PMN stimulated with N-formyl-L-methionyl-L-leucyl-L-phenylalanine (fMLP), and enhanced the inhibitory action of unstimulated PMN on platelet responses to the stable endoperoxide analog U46619. These effects may be of pharmacological interest in view of the multiple interactions between platelets and leukocytes that occur in thrombotic and inflammatory diseases.

15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5↗
PubMed1990

Cloricromene inhibits the activation of human platelets by ADP alone or in combination with adrenaline.

Cloricromene may inhibit platelet activation induced by several agonists. In this study we report that ADP and adrenaline synergistically promote platelet aggregation and cytoplasmic Ca2+ movements in aequorin-loaded platelets. Cloricromene caused dose-dependent reduction in platelet aggregation and cytoplasmic Ca2+ movements after exposure of the cells to a low concentration of ADP (2 microM) or to a combination of ADP (2 microM) and adrenaline (10 microM). Cloricromene's inhibitory action may be of considerable pharmacological interest since platelet activation by a combination of agonists may mimic the conditions under which thrombosis occurs in vivo.

Adenosine Diphosphate↗
PubMed1990

Inhibition of platelet function by polymorphonuclear leukocytes.

Human polymorphonuclear leukocytes (PMNs) were evaluated for their ability to modulate platelet response induced by collagen, thrombin, platelet-activating factor and the stable analog of cyclic endoperoxides U46619. Platelet aggregation was first evaluated in whole blood and in leukocyte-depleted whole blood by the impedance method. This novel approach highlighted the inhibitory role of leukocytes on platelet aggregation in whole blood. The inhibitory role of PMNs on platelet function was subsequently evaluated on washed cells. PMN inhibition of platelet aggregation and beta-thromboglobulin release was more evident with threshold concentrations of stimuli. The inhibition also depended on the number of PMNs incubated in mixed cellular suspensions. Higher concentrations of stimuli may overcome the PMN-dependent inhibition. Under this condition, preincubation of cells with N-formyl-methionyl-leucyl-phenylalanine (a specific PMN agonist) restored the inhibitory effect of PMNs on platelet aggregation in whole blood and in mixed cellular suspensions. Not only PMNs, but also PMN-derived supernatants, dose-dependently inhibited U46619-induced platelet aggregation, suggesting that the inhibition observed may be exerted by chemically stable compound(s). Cytoplasmic Ca2+ movement was measured in aequorin-loaded platelets exposed to thrombin or U46619 to see whether cytoplasmic Ca2+ levels were affected by PMN. Ca2+ levels were similar in the presence or absence of PMNs, suggesting that inhibition may be related to a subsequent platelet response step. A series of bioassay experiments showed that PMNs were able to remove and/or convert adenosine diphosphate available for platelet aggregation but not to reduce U46619 availability. Our findings suggest that (1) unstimulated PMNs may release factor(s) that inhibit platelet aggregation and beta-thromboglobulin release; (2) this in itself is sufficient to block the platelet response to a threshold concentration of stimuli; (3) release of the same or other inhibitory mediators from stimulated PMNs may have to be greater to inhibit platelet response to higher concentrations of stimuli. Data presented here suggest that adenosine diphosphatase activity and chemically stable, as yet unidentified, compounds besides previously well-characterized labile compounds such as nitric oxide and arachidonic acid metabolites are responsible for the PMN-dependent mechanism of inhibition of platelet response that could be relevant in physiopathologic conditions.

Adenosine Diphosphate↗
PubMed1989

Molecular aspects of cloricromene (AD6) distribution in human platelets and its pharmacological effects.

Cloricromene (AD6) is an investigational drug which inhibits platelet aggregation and release reaction. We studied the relationship between its action and its distribution and metabolism in platelets. Incubation of anticoagulated whole blood or platelet-rich plasma (PRP) without exogenous aggregating agents resulted in a progressive decrease of platelet count with a concomitant increase of beta-thromboglobulin (BTG) release. AD6 (20-50 mumols/l), but not acetylsalicylic acid (ASA), incubated with whole blood or PRP, prevented the fall in platelet count and the release of BTG for at least 150 min. Moreover, incubation of PRP with AD6 (50 mumols/l) and subsequent stimulation by ADP at threshold concentrations resulted in a significant reduction (about 30%) in aggregation for at least 90 min. AD6 (20 mumols/l) added to PRP was rapidly metabolized by hydrolysis of an ester bond to AD6 acid, a stable catabolite pharmacologically inactive in platelets. Significant amounts of AD6 acid (up to 13.26 +/- 2.80 pmol/10(6) platelets) were associated with the platelets after incubation either at 37 degrees C or 4 degrees C. The amount of AD6 acid in the platelet pellet was proportional to AD6 concentration (2 to 100 mumols/l). PRP incubation with AD6 acid (20 mumols/l) resulted in very low levels (less than 1 pmol/10(6) platelets) of the same compound in the platelet pellet after 1, 5 or 30 min. These data suggest that AD6 is taken up as an ester and converted to its acid catabolite with a consequent long-lasting inhibition of platelet function.

Adenosine Diphosphate↗
PubMed1988

Stenosis and vascular damage as a cause of thrombosis in the dog femoral artery.

We describe here an experimental model of peripheral arterial thrombosis and the effect of several drugs which are known to affect vessel and platelet biological functions. A similar method has been previously applied by us and others on dog coronary arteries. Male Beagle dogs, under pentobarbital anesthesia, were instrumented to measure arterial pressure, heart rate, ECG, femoral blood flow and expired CO2. A segment of the femoral artery was squeezed with forceps to damage the endothelium, and a plastic cylinder was placed around the vessel in the area of the damage. The cylinders had a length of 2 mm and an internal diameter of 1.6-1.8 mm. Under these circumstances blood flow in the stenosed artery was reduced by about 60-70% from control value and showed cyclic blood flow variations (CBFV). CBFV eventually led either to a total occlusion of the vessel (documented by blood flow measurement and by angiographic analysis), or to a spontaneous partial restoration of flow, followed by another decrease, in a repetitive fashion. Drug effect was monitored by observing the changes in frequency and amplitude of CBFV. Ketanserin (0.25 mg/kg), dazmegrel (0.5 mg/kg), and chlorpromazine (0.5 mg/kg), abolished or greatly reduced CBFV in all the experiments, while acetylsalycilic acid (ASA, 10 mg/kg) reduced or abolished CBFV in 60% of the treated dogs. Heparin (50 I.U./kg), dipyridamole (1.0 mg/kg) and prazosin (0.1 mg/kg) did not change CBFV. These results emphasize the importance of serotonin and thromboxane as mediators of vascular occlusion in this particular experimental model. This approach provides a reproducible in vivo preparation to study the pharmacological control of peripheral arterial thrombosis.

Angiography↗
PubMed1987

At low extracellular calcium concentration platelet activating factor induces beta-thromboglobulin release from human platelets through thromboxane-endoperoxides formation.

The aim of this study was to investigate the mechanism involved in beta-thromboglobulin (BTG) release induced by platelet activating factor (PAF) in human platelet-rich plasma (PRP) and washed platelets (WP) during aggregation. PAF was used in PRP at increasing concentrations starting at its threshold concentration for irreversible aggregation (TAC: 90-150 nM). In citrated PRP, PAF induced release of BTG (80-95% of total content) and thromboxane B2 (TXB2) formation (30-40 pmol/ml). At low PAF concentrations aggregation and BTG release were blocked by apyrase (a scavenger of ADP), by ASA (an inhibitor of cyclooxygenase) and by BM 13177 (a thromboxane receptor antagonist). Higher concentrations of PAF overcame the effect of apyrase, but only induced reversible aggregation and minor release in the presence of ASA or BM 13177. In heparinized PRP, PAF induced full irreversible aggregation, but only very low BTG release (about 25% of total content) and thromboxane synthesis (2-3 pmol/ml). WP resuspended in the presence of 2 mmol/l Ca2+ seldom responded to PAF alone, as previously shown by others, but full aggregation could be induced by concomitant addition of subthreshold concentrations of PAF (25-50 nM) and epinephrine (1 microM). In these conditions average BTG release from WP was less than 20% of the amount releasable by thrombin. In contrast, when WP were resuspended in the absence of Ca2+, stimulation by PAF + EPI induced sustained BTG release (40-50% of total content) and TXB2 synthesis (15-20 pmol/ml). We conclude that at low Ca2+ concentration PAF induces BTG release mainly through thromboxane-endoperoxides formation. In contrast, when [Ca2+] is normal, PAF does not or weakly induces thromboxane formation and BTG release.

Blood Platelets↗
PubMed1986

The effect of AD6 (8-monochloro-3-beta-diethylamino-ethyl-4-methyl-7-ethoxy- carbonylmethoxycoumarin) on washed human platelet aggregation induced by platelet activating factor (PAF) and epinephrine.

The effect of AD6 (8-monochloro-3-beta-diethylamino-ethyl-4-methyl-7- ethoxycarbonylmethoxycoumarin) was studied in vitro on washed human platelet aggregation and beta-thromboglobulin (beta TG) release induced by PAF alone or in combination with epinephrine. AD6 caused concentration-dependent inhibition of aggregation and beta TG release. Acetylsalicylic acid and the ADP scavenger apyrase were ineffective on the same parameters, while the specific PAF antagonist CV 3988, the Ca2+ antagonist diltiazem and the cyclo-oxygenase-lipoxygenase inhibitors nordihydroguaiaretic acid and BW 755C inhibited aggregation. The results indicate that AD6 is an inhibitor of platelet aggregation in vitro with a mode of action different from cyclo-oxygenase blockade.

4,5-Dihydro-1-(3-(trifluoromethyl)phenyl)-1H-pyraz↗
PubMed1986

Action of AD6 (8-monochloro-3-beta-diethylaminoethyl-4-methyl-7-ethoxycarbonylmet hox y coumarin) on human platelets in vitro.

The action of AD6 was tested in vitro on human platelets by measuring beta-thromboglobulin (BTG), platelet factor 4 (PF4) and thromboxane B2 (TXB2) release as well as aggregation. BTG and PF4 release from blood anticoagulated with sodium citrate was inhibited by AD6 during a 3 h incubation. Platelet rich plasma (PRP) was stimulated with ADP, collagen, sodium arachidonate, PAF, A23187 and epinephrine, while resuspended washed platelets (WP) were stimulated by thrombin. AD6 (5-100 microM) inhibited dose dependently aggregation, BTG, PF4 and TXB2 release induced by threshold concentration of all the tested aggregating agents; however AD6 action could be overcome by increasing the concentration of the stimulating agents. After cyclo-oxygenase blockade by acetylsalicylic acid (ASA), PRP was stimulated by a supramaximal concentration of PAF. Under these circumstances we could observe a reversible aggregation and a partial release of BTG and PF4, AD6 was able to further reduce aggregation and release. Cyclic AMP accumulation induced in WP by prostacyclin was not modified by AD6 (100 microM), while theophylline greatly potentiated prostacyclin action. We conclude that AD6 is an inhibitor of platelet activation in vitro. Its mode of action is different from cyclo-oxygenase blockade and provides inhibition of platelet activation by a number of different stimuli.

Anticoagulants↗
PubMed1985

Disappearance of human platelet factor 4 (PF4) in rabbits: does an immediate component exist?

Twelve male New Zealand rabbits were injected with 21 micrograms/kg of human platelet factor 4 antigen (PF4). The decay of the protein followed a monoexponential curve for the first 5 mins, with a half-life (t 1/2) of 1.94 mins and a calculated concentration at 0 time (CO) of 79.4 ng/ml. Five rabbits were pre-treated with heparin (2.500 I.U. i.v.) and 3 mins later were injected with the same amount of PF4. PF4 decay followed a monoexponential curve with a t 1/2 of 25.3 mins, and with CO of 380.8 ng/ml. This value is not greatly different from the one calculated assuming an immediate and uniform distribution in plasma (482.7 ng/ml for a plasma volume of 43.5 ml/kg). The 12 rabbits injected with PF4 were divided in 3 groups, in which heparin was given at 10', 30' or 60' after PF4, respectively. After heparin the peak levels of PF4 were 139.9 ng/ml, 65.3 ng/ml and 52.7 ng/ml, respectively. The following monoexponential PF4 decay had t 1/2 of 20.7, 25.6 and 26.0 mins, respectively. In a separate group of 4 animals, we studied heparin decay after an intravenous bolus of 2.500 I.U. Heparin decay could not be described by a monoexponential equation and was different from the decay of PF4 injected after heparin. On the basis of the present data we suggest the presence of an immediate component of PF4 decay, most likely due to uptake by the tissues. Heparin pretreatment may avoid this uptake process.

Animals↗
PubMed1984

Clearance and in vivo release by heparin of human platelet factor 4 (PF4) in the rabbit.

13 male New Zealand rabbits were injected with two different doses (25 micrograms/Kg and 100 micrograms/Kg) of human platelet factor 4 antigen (PF4). The disappearance of the protein was extremely fast with an half-life for the fast component of 1.07 +/- 0.16 and 1.76 +/- 0.11 min respectively. The half-life for the slow component, detectable only with the highest dosage, was 18.8 min. The administration of 2500 I.U. of heparin 30 min after PF4 administration induced a partial release of the injected protein and its clearance from plasma was slow, with half-life of 23.3 +/- 5.9 min and 30.9 +/- 2.19 min respectively.

Animals↗