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M Chignard

Publications and source records attributed to M Chignard.

At least 37 records · Page 2Linked to original sources
PubMed1994

Proteinase 3. A neutrophil proteinase with activity on platelets.

Purified proteinase 3 (PR3) devoid of any elastase (HLE) and cathepsin G (Cat.G) contaminants, was prepared from azurophilic granules of human polymorphonuclear neutrophils by using a novel procedure. Although unable to induce platelet activation (up to 25 micrograms/ml) by itself, PR3 at a concentration as low as 2.5 micrograms/ml enhanced the platelet response to a concomitantly added threshold concentration of Cat.G, a recognized platelet agonist. In the presence of 10 micrograms/ml PR3, aggregation and degranulation of platelets induced by Cat.G were 43.2 +/- 5.9% and 27.1 +/- 1.9% as compared with 5.5 +/- 2.9% and 4.2 +/- 1.5% (n = 4) for Cat.G alone. This enhancing effect by PR3 was also observed with collagen and a cyclic endoperoxide analogue, and was inhibited by eglin C and elafin, two PR3 inhibitors. Associated with the removal of activity by a anti-PR3 mAb and the lack of effect of the secretory leukocyte proteinase inhibitor, these data demonstrated that the effect is specifically related to the enzymatic activity of PR3. It is hypothesized that this mechanism could play a role in the polymorphonuclear neutrophil-mediated platelet activation, an event already known to be dependent on Cat.G and HLE. This is supported by the fact that the association of PR3 and HLE, at concentrations ineffective by themselves, was able to potentiate Cat.G-induced platelet activation.

Blood Platelets↗
PubMed1994

Modulation by superoxide anions of neutrophil-mediated platelet activation.

When polymorphonuclear neutrophil-platelet suspensions were stimulated by 0.5 microM N-formyl-Met-Leu-Phe in the presence of 40 U/mL of superoxide dismutase, a significant reduction of platelet secretion was observed (51.4 +/- 6.3% vs 62.4 +/- 4.6% for control; mean +/- SEM; N = 6; P < 0.01). This was due to the superoxide anion scavenging property of superoxide dismutase since neutrophil degranulation, cathepsin G and elastase enzymatic activities (the two main mediators of this cell-to-cell interaction) and platelet reactivity were not affected. Involvement of superoxide anions was confirmed using leukotriene B4, a neutrophil agonist which induces degranulation with minimal superoxide anion production. Indeed, serotonin release induced by this agonist was unchanged whether superoxide dismutase was added or not.

Cathepsin G↗
PubMed1994

Regulation of the structure and activity of platelet adhesion receptors by leukocyte proteinases.

Two major membrane receptors implicated in the adhesive properties of blood platelets are the GPIb-IX complex, a receptor for subendothelial von Willebrand factor, and the alpha IIb beta 3 integrin, the receptor for plasma fibrinogen. We have evaluated how the biological activities of these receptors can be potentially modulated through limited proteolysis when platelets are exposed to the serine-proteinases secreted by activated polymorphonuclear neutrophils, i.e., leukocyte elastase (EL) and cathepsin G (CG). CG can activate the alpha IIb beta 3 integrin through intracellular metabolic pathways, but has no direct proteolytic activity on the receptor subunits. By contrast, EL does not activate the platelet metabolism, but specifically cleaves a short peptide sequence within the alpha IIb subunit, and this cleavage occurs in parallel with an up-regulation of the activity of the fibrinogen receptor. On another hand, both EL and CG cleave the amino-terminal portion of the GPIb alpha subunit of the GPIb-IX receptor, eliminating the binding site for von Willebrand factor and diminishing the capacity of platelets to interact with this adhesion protein. Thus, neutrophil proteinases have the potential to regulate the activity of platelet adhesion receptors, and such experimental observations may prove to be relevant in vivo in various pathological conditions.

Endopeptidases↗
PubMed1994

Progressive inactivation of cathepsin G and elastase released from activated neutrophils in vitro: lack of participation of the neutrophil alpha 1-proteinase inhibitor.

Addition of platelets to activated human polymorphonuclear neutrophils (PMNs) led to their aggregation and degranulation, with these responses decreasing as a function of the time interval between PMN activation and platelet addition. Thus, for a 15-second interval platelet aggregation and serotonin release reached 51.3% +/- 6.7% (n = 11) and 64.3% +/- 4.9% (n = 8), respectively, but after a 5-minute interval they were totally absent. This effect was correlated with the decrease in enzymatic activities of elastase (HLE) and cathepsin G (CAT-G) that were released on PMN activation and responsible for the activation of nearby platelets (r = 0.86 and 0.90 for CAT-G and HLE, respectively; p < 0.05). Because it has been recently shown that PMNs express an alpha 1-proteinase inhibitor (alpha 1-Pl) gene and secrete this antiproteinase at their surface, we investigated whether the PMN alpha 1-Pl could regulate the biologic activities of both proteinases. Although superoxide anions oxidize alpha 1-Pl and reduce its affinity for CAT-G and HLE, maneuvers aimed at modifying their concentrations did not modify the loss of CAT-G and HLE. In fact, the progressive decrease of the two proteinase enzymatic activities followed the same pattern whether PMNs were present or not. It is concluded that PMN alpha 1-Pl is not involved in the time-dependent inactivation of CAT-G and HLE released from activated PMNs.

Blood Platelets↗
PubMed1993

Inhibition by human leukocyte elastase of neutrophil-mediated platelet activation.

When human polymorphonuclear neutrophils and platelets were incubated with human leukocyte elastase before N-formyl-Met-Leu-Phe (FMLP) challenge, a time- and concentration-dependent inhibition of the resulting platelet activation was observed. Thus, when the mixed cell suspension was preincubated for 6 min with 1 microM elastase before stimulation of neutrophils with 0.5 microM FMLP, resulting aggregations and serotonin releases were respectively only 4.4 +/- 4.1% (n = 4) and 1.6 +/- 2.4% (n = 4) as compared to 41.6 +/- 5.2% (n = 9) and 71.3 +/- 16.0 (n = 9) for controls. A direct inhibitory action of elastase on neutrophil activation was ruled out, as well as a breakdown of cathepsin G, a mediator involved in neutrophil-mediated platelet activation. In fact, we demonstrated that the target for the inhibitory effect of elastase in such a cell-to-cell cooperation system was the platelet. This phenomenon is likely to play a role under in vivo conditions in pathologies in which a significant granulocytic proteolytic activity has been detected in the plasma.

Blood Platelets↗
PubMed1993

Enhancement of cathepsin G-induced platelet activation by leukocyte elastase: consequence for the neutrophil-mediated platelet activation.

We have focused our interest on the platelet-activating properties of two polymorphonuclear neutrophil (PMN)-derived proteinases, namely elastase (HLE) and cathepsin G (Cat.G). First of all, we observed that whereas HLE was unable to trigger platelet activation by itself, it enhanced platelet activation induced by Cat.G when both proteinases were added simultaneously. It has been recently described that, upon stimulation, PMN released Cat.G, which in turn activated surrounding platelets. Thus, we looked for a combined effect of Cat.G and HLE during this cell-to-cell interaction. When PMN (5 x 10(6)/mL) were stimulated by 0.5 mumol/L N-formyl-Met-Leu-Phe, they released 237.9 +/- 49.1 nmol/L Cat.G and 381.7 +/- 28.0 nmol/L HLE. Such a concentration of purified Cat.G (240 nmol/L) induced only a moderate platelet activation when added to a PMN-platelet mixture. However, when Cat.G (240 nmol/L) and HLE (380 nmol/L) were added together, the resulting platelet activation was strictly comparable to that corresponding to the addition of N-formyl-Met-Leu-Phe (P > .05) in terms of aggregation, dense and alpha granule secretion, and thromboxane B2 production. In fact, Elafin, a specific HLE inhibitor, when added to the PMN-platelet cooperation system triggered by N-formyl-Met-Leu-Phe, prevented platelet activation within the same range of concentrations as for inhibition of HLE activity. In conclusion, we now show that not only Cat.G, but also HLE is involved in the PMN-mediated platelet activation.

Amino Acid Sequence↗
PubMed1993

Inhibition by recombinant SLPI and half-SLPI (Asn55-Ala107) of elastase and cathepsin G activities: consequence for neutrophil-platelet cooperation.

1. The capacity of recombinant human secretory leukocyte proteinase inhibitor (SLPI) to inhibit human leukocyte elastase (HLE) and cathepsin G (Cat G) was investigated and compared with a recombinant truncated form (carboxyl-terminal domain, Asn55-Ala107) called 1/2 SLPI. 2. Both compounds were efficient when tested against enzymatic activities of purified HLE and Cat G indicating that the HLE- and Cat G-inhibitory sites were preserved in the truncated form. SLPI and 1/2 SLPI also affected platelet activation induced by 0.2 microM Cat G (IC50 = 112 +/- 13 nM for SLPI and 280 +/- 12 nM for 1/2 SLPI). 3. The effects of SLPI and 1/2 SLPI were then tested against polymorphonuclear neutrophil (PMN)-mediated platelet activation, a cell-to-cell interaction mediated by HLE and Cat G released from PMN. In this experimental system, addition of SLPI or 1/2 SLPI before N-formyl-Met-Leu-Phe (fMLP) led to the inhibition of the resulting platelet activation. As was the case for Cat G enzymatic activity and Cat G-induced platelet activation, SLPI was more efficient than 1/2 SLPI (IC50 = 676 +/- 69 nM vs 1121 +/- 150 nM). 4. The ratio of the IC50 against PMN-mediated platelet activation compared to purified Cat G-mediated platelet activation was 6.03 for SLPI and 4.32 for 1/2 SLPI. This difference may be due to the smaller size of the truncated form which could allow this molecule to diffuse more easily between PMN and platelets. 5. In conclusion, 1/2 SLPI could be a promising candidate in the treatment of pathological states linked to inflammation in which participation of HLE and Cat G has been evoked.

Amino Acid Sequence↗
PubMed1993

Leucocyte elastase-mediated release of von Willebrand factor from cultured endothelial cells.

The aim of this study was to investigate a possible activation of human endothelial cells in monolayer culture by a purified neutrophil-derived proteinase, i.e. elastase. Cells were isolated from human umbilical cord veins, and incubated either in primary culture or after two passages, in the presence of various concentrations of this proteinase. Although a lack of prostacyclin formation was noted, elastase induced a large release of von Willebrand factor (vWf) in a concentration-dependent manner. Thus, upon incubation with 1 microgram.ml-1 elastase for 30 min, 70 mU.ml-1 vWf were detected in the incubation medium, as compared to 5 mU.ml-1 for control. Using cells in primary culture, a fivefold higher concentration of vWf was recovered following incubation with 10 micrograms.ml-1 elastase than with 0.5 IU.ml-1 thrombin. This effect was linked to the enzymatic activity of elastase and not to its cationic charge, as deduced from the inhibition by eglin C, and the lack of effect of the phenylmethylsulphonyl fluoride (PMSF) treated proteinase. We conclude that vWf release was not due to cell activation, since cytoplasmic calcium mobilization was absent, and inhibition of protein kinase C did not modify the response. In fact, this release was the consequence of cell damage, since concentrations of vWf recovered correlated with cell lysis. These results support the hypothesis that the high level of plasma vWf in patients with sepsis or adult respiratory distress syndrome could result from damage to the endothelial cells by elastase released from activated neutrophils.

Calcium↗
PubMed1992

Activation and damage of cultured airway epithelial cells by human elastase and cathepsin G.

Accumulation of polymorphonuclear neutrophils (PMN) and epithelium damage have often been described during airway inflammation. We studied the effects of two PMN-derived proteinases, namely elastase and cathepsin G, on guinea-pig tracheal epithelial cells in culture. Both proteinases activated tracheal epithelial cells in terms of prostaglandin (PG) E2 production. A concentration- and time-dependent effect was observed with 10 micrograms/ml and 6 h as the optimal conditions for both enzymes. Optical microscopic studies confirmed an effect on tracheal epithelial cells as intercellular gaps were observed upon incubation of the monolayers with proteinases. A small cytotoxic effect was observed after 1 h incubation but remained stable up to 6 h. This cytotoxic effect, more pronounced with elastase than with cathepsin G, was dissociated from PGE2 formation.

Animals↗
PubMed1992

Combined activation of platelets by cathepsin G and platelet activating factor, two neutrophil-derived agonists.

In this paper we have studied the combined effects on platelet activation, of two polymorphonuclear neutrophil (PMN)-derived agonists, namely platelet-activating factor (PAF) and cathepsin G (Cat.G), used at threshold concentrations. Our results showed that the order of agonist addition was a determinant factor since the addition of Cat.G prior to PAF induced a full platelet activation while the reverse combination had no effects. The successive challenge of platelets by Cat.G and then PAF induced a strong aggregation accompanied by an enhancement of alpha and dense granule secretion. The observed phenomenon was also dependent on the time interval between agonist addition. It was significant at 30 s (P less than 0.05) and plateaued over 1-2 min. Platelet activation resulting from the combination Cat.G-PAF can be described as a function of PAF concentrations, the synergism being significant between 10 nM and 1 microM. The mechanism by which Cat.G primes platelets remains to be elucidated. However, some points have been examined and have led us to conclude that an increase in expression and/or affinity of PAF receptors, [Ca2+]i movements, protein kinase C activation and phospholipase A2 pathway are not involved. Whatever the biochemical mechanism underlying this synergism which involved PMN and platelets, it may constitute a link between the inflammatory and haemostatic processes in response to tissue damage.

Cathepsin G↗
PubMed1992

Heparin inhibits neutrophil-induced platelet activation via cathepsin G.

Activated human polymorphonuclear neutrophils (PMNs) induce platelet stimulation via cathepsin G (cat G), a platelet-activating cationic serine proteinase released from the azurophilic granules. Heparin inhibited up to 100% of aggregation and serotonin release triggered by 180 nmol/L of purified cat G in a concentration-dependent manner between 10 and 70 mU/ml. When tested against the enzymatic activity of 180 nmol/L cat G (hydrolysis of N-succinyl-Ala-Ala-Pro-Phe-p-nitroanilide), inhibition by heparin never exceeded 60% (up to 100 U/ml). Inhibition was most probably related to electrostatic interactions between the cationic cat G and the anionic heparin, because addition of 180 mU/ml of protamine sulfate restored platelet activation. Low molecular weight heparin, CY 216, used between 5 and 50 mU/ml, also inhibited cat G-induced platelet activation. When purified PMNs and washed platelets were mixed together and challenged with a PMN agonist (FMLP at 1 mumol/L), platelet activation was observed. Pretreatment of the mixed cell population with 300 mU/ml heparin prevented platelet activation. This was illustrated by electron microscopy studies. The present data, apart from confirming a participation of cat G in the PMN-platelet interaction, bring evidence that heparin has potent antiproteinase effect in a biologic model.

Blood Platelets↗
PubMed1991

Tumor necrosis factor-alpha enhances platelet activation via cathepsin G released from neutrophils.

In this paper we show that TNF-alpha enhances platelet activation. Experiments were performed on a human polymorphonuclear neutrophil (PMN)-platelet cooperation system in which PMN, stimulated by FMLP, release cathepsin G (Cat.G), a serine proteinase responsible for the activation of nearby platelets. Pretreatment of the mixed cell suspension with 5 ng/ml TNF-alpha resulted in a strong platelet activation (37.7 +/- 3.2% aggregation; 46.0 +/- 14.4% serotonin release) in response to a weak concentration of FMLP (1.25 x 10(-8) M) inducing by itself only 7.7 +/- 4.0% of aggregation and 3.8 +/- 4.1% of serotonin release (mean +/- SD; n = 10). This effect was concentration dependent (maximum between 5 and 10 ng/ml) and was optimal for a brief preincubation time (5 min). Under these experimental conditions the target of TNF-alpha was PMN, as shown by beta-glucuronidase release. The observed potentiation was modified neither by 0.1 mM acetyl salicylic acid (a cyclo-oxygenase inhibitor) nor by 0.1 mM BN 52021 (a platelet-activating factor antagonist), while such a phenomenon was fully inhibited by 20 micrograms/ml eglin C, a strong and specific inhibitor of the human granulocytic proteinases, elastase and Cat.G. In fact, full inhibition was also observed with 300 nM alpha-1-antichymotrypsin, a specific inhibitor of Cat.G. This clear-cut evidence of Cat.G involvement was substantiated by the enhancement of Cat.G release from FMLP-activated PMN primed with TNF-alpha. These results demonstrate that the priming of PMN by TNF-alpha may modulate the activation of other inflammatory cells, particularly of platelets. It is hypothesized that this phenomenon could contribute to pulmonary pathologies, and more specifically to the adult respiratory distress syndrome, a disease for which PMN, platelet and TNF-alpha involvement has been proposed.

Cathepsin G↗
PubMed1991

Interference of anti-inflammatory and anti-asthmatic drugs with neutrophil-mediated platelet activation: singularity of azelastine.

1. The capacity of various drugs (acetylsalicylic acid (ASA), ketoprofen, diclofenac, piroxicam, BW 755C, BW A4C, nedocromil sodium and azelastine) to inhibit human polymorphonuclear neutrophil (PMN)-mediated platelet activation was investigated. In this model, stimulated PMN release cathepsin G (Cat G), a serine proteinase which, in turn, induces platelet activation. 2. Among the different tested drugs, azelastine (100 microM for 1 min) was the only one able to prevent platelet aggregation. The cyclo-oxygenase inhibitors were all inactive, although used at effective concentrations as judged by inhibition of thromboxane B2 (TxB2) formation. Inhibition of platelet aggregation by azelastine was concentration-dependent, the range of active concentrations being of 20-70 microM. Release from platelets of 5-hydroxytryptamine was also inhibited at 30 microM and above, but never reached 100%. 3. The inhibition by azelastine is due to an effect on both cells. Indeed, beta-glucuronidase release from activated PMN and platelet activation by purified Cat G were both affected. 4. However, used at high concentrations (greater than 100 microM) azelastine was toxic since it released significant amounts of lactate dehydrogenase (LDH) from PMN and platelets. 5. These results show the capacity of azelastine, an anti-allergic and anti-asthmatic compound, to inhibit the cell-to-cell communication between PMN and platelets, an effect which may be relevant for its therapeutic efficacy or for a new application in diseases in which PMN and platelets are involved.

Anti-Inflammatory Agents, Non-Steroidal↗
PubMed1990

Cooperation between platelets and neutrophils for paf-acether (platelet-activating factor) formation.

Association of platelets and neutrophils is frequently observed within thrombi or inflammatory sites. Interactions between these two cell populations have been reported for the production of several mediators of inflammation such as hydrogen peroxides or leukotrienes. Another potential mediator of thrombosis and inflammation is paf-acether, which is synthesized by activated platelets and neutrophils. Since platelets form and release large amounts of the paf-acether precursor lyso paf-acether, platelet and neutrophil cooperation for paf-acether biosynthesis was investigated. Purified human neutrophils (4 x 10(6)/ml) stimulated by opsonized zymosan (ZC, 1 mg/ml) formed 4.5 +/- 2.5 ng/ml paf-acether. Human washed platelets (3 x 10(8)/ml) stimulated with thrombin (1 IU/ml) formed 0.60 +/- 0.43 ng/ml paf-acether. Platelets and neutrophils, incubated together and both stimulated by their specific agonist, formed more than twice as much paf-acether as did platelets and neutrophils separately (10.90 +/- 4.25 ng/ml, n = 6, P less than .001). The formation of lyso paf-acether and the release of lysozyme and LDH were unchanged under the cooperation conditions. The formation of paf-acether almost doubled (10.24 +/- 3.81 ng/ml paf-acether vs. 5.30 +/- 2.23, P less than .05, n = 4) when ZC-stimulated neutrophils were incubated with supernatants from thrombin-stimulated platelets as well as with synthetic lyso paf-acether. Extracted and purified lyso paf-acether from thrombin-stimulated platelets led to an increase of biosynthesis of paf-acether by neutrophils (13.86 +/- 2.26 ng/ml paf-acether vs. 5.76 +/- 0.38, P less than .05, n = 3). These results indicate that a cooperation between platelets and neutrophils exists for paf-acether formation. The phenomenon depends on a platelet-derived soluble factor, possibly lyso paf-acether. This cell-to-cell interaction is of interest since paf-acether is formed by and acting on platelets and neutrophils and represents a molecular basis for potent amplification of inflammatory reactions.

Animals↗
PubMed1990

Activation of human platelets by C5a-stimulated neutrophils: a role for cathepsin G.

Human platelets can be stimulated by recombinant human fifth component of complement (rhC5a) in the presence of human neutrophils. After challenge with N-formyl-Met-Leu-Phe or rhC5a, concentrated neutrophils release cathepsin G into the supernatant. The concentrations of cathepsin G recovered by titration of the enzymatic activity correlate with the capability of these supernatants to induce platelet stimulation as measured by serotonin release. Cathepsin G purified from neutrophil granules triggered platelet aggregation and serotonin release independent of arachidonic acid metabolites and platelet-activating factor formation. A concentration of 100 nM of cathepsin G, which was reached in the surrounding space of activated neutrophils, induced a 50% platelet stimulation. Three distinct antiproteinases were tested against cathepsin G-induced platelet activation. Z-Gly-Leu-Phe-CH2Cl, a specific inhibitor of cathepsin G enzymatic activity, proved to be nonspecific in our biological system. By contrast, alpha 1-antichymotrypsin and alpha 1-antitrypsin displayed specific activities. The physiological specific inhibitor of cathepsin G, alpha 1-antichymotrypsin, was the most potent and was used in the rhC5a-induced neutrophils-mediated platelet activation. A complete inhibition was achieved, showing that release of cathepsin G from neutrophils accounts for platelet activation. Such a chain of events involving C5a, neutrophils, cathepsin G, and platelets may be of relevance in certain inflammatory states, particularly the adult respiratory distress syndrome.

Amino Acid Sequence↗
PubMed1990

Interference of recombinant eglin C, a proteinase inhibitor extracted from leeches, with neutrophil-mediated platelet activation.

Eglin C is an inhibitor of two serine proteinase-derived polymorphonuclear leucocytes (PMN) i.e., elastase and cathepsin G. Since the latter has recently been shown to be involved in the activation of platelets by stimulated PMN, the effects of recombinant eglin C in the PMN-platelet cooperation model were studied. First, the inhibitory capacity of eglin C against purified cathepsin G was measured spectrophotometrically by following hydrolysis of a specific synthetic substrate, N-succinyl-ala-ala-pro-phe-para-nitroanilide. The inhibition of the enzymatic activity of 180 nM (5 micrograms/ml) cathepsin G was directly proportional to eglin C concentration and reached 100% with 2 micrograms/ml (240 nM). Platelet activation generated by a submaximal concentration of cathepsin G (200 nM) was also totally suppressed by 2 micrograms/ml of eglin C. Inhibition was specific (a 100 times higher concentration of eglin C did not alter platelet activation induced by thrombin), and surmountable (an increase of cathepsin G concentration reduced the eglin C effect). Thus, the mechanism of inhibition by eglin C of cathepsin G-induced platelet activation could be explained by a stoichiometric relation between eglin C and cathepsin G as previously described. Investigations were then performed with the PMN-platelet cooperation model, using two distinct stimuli, N-formylmethionylleucylphenylalanine (FMLP) or recombinant human C5a, at submaximal concentrations, 2.10(-7) M and 10(-7) M, respectively. A concentration-dependent inhibition of platelet activation aggregation and serotonin release-lambda was observed. Eglin C used at 10 micrograms/ml and 25 micrograms/ml totally blocked the platelet responses induced by recombinant human C5a and FMLP, respectively. Leucotriene B4, but also thromboxane B2 production measured by radioimmunoassays, were observed under FMLP activation. In the presence of eglin C, thromboxane B2 formation was totally suppressed, whereas leucotriene B4 synthesis was still effective. In fact, the mechanism of inhibition of eglin C is located neither on PMN (leucotriene B4 formation by FMLP-activated PMN was not affected), nor on platelets (response to thrombin was unchanged). The target is most probably cathepsin G since eglin C suppressed thromboxane B2 formation by platelets challenged by this serine proteinase. These results constitute an argument in favor of the implication of cathepsin G in the PMN-mediated platelet activation. Moreover, they reinforce the hypothesis that this mechanism could be operating under in vivo pathologic conditions, since eglin C is capable of preventing or ameliorating some experimental pulmonary diseases.

Animals↗
PubMed1989

Reduced sensitivity of human platelets to PAF-acether following ticlopidine intake.

We investigated in 9 patients the effect of a 7-day treatment by Ticlopidine (250 mg b.i.d.) on washed platelets activation by PAF-acether in comparison with adenosine 5'-diphosphate (ADP) and arachidonic acid (AA). Aggregations induced by ADP were totally suppressed upon drug administration. AA-induced aggregations were partly but significantly inhibited (p less than 0.05). Responses of platelets to PAF-acether before treatment differed from patient to patient. A paired Student's test and a two-way analysis of variance showed a significant inhibitory effect of Ticlopidine treatment on PAF-acether-induced aggregation. The inhibitory effect of Ticlopidine or its metabolite(s) was evidenced after platelet washing procedure, suggesting a persistent effect of this drug on platelet after administration of the drug has been stopped.

Adenosine Diphosphate↗