Efficacy of the Latham Blood Processor To Perform Plateletpheresis

Platelets were separated during extracorporeal circulation by the method of Tullis et al . in the Latham Blood Processor equipped with a 375 ml disposable centrifuge bowl. The average yield of platelets recovered from each 710 ml unit of blood was 0.665 × 10¹¹ ± 0.244 × 10¹¹ (± 1 S. D.) N = 105, corresponding to an efficiency of 45.8 ± 12.4 per cent.
Platelets were also separated during extracorporeal circulation by a modification of the method. In this technic the final platelet concentrate contained about 50 ml of red blood cells that were removed by an additional centrifugation. The average quantity of platelets recovered from each 710 ml of blood was 1.23 × 10¹¹ ± 0.285 × 10¹¹ (± I S. D.) corresponding to an efficiency of removal of 69.5 ± 14.6 per cent (± I S. D.). The modified method permitted a collection of large numbers of platelets from single donors in a relatively short time.
Donor effects of the procedure were insignificant. Mild to moderate donor reaction consisting of chilling and hypotensive reactions occurred in three of 145 donors. Transient decreases occurred in hematocrit and platelet count. No untoward effects were observed following repeated procedures.     

Platelet count and the risk for thrombosis and death in the elderly

Summary.  Aim: Our aim was to examine the association between platelet count and the incidence of myocardial infarction, ischemic stroke, hemorrhagic stroke, venous thrombosis, and mortality. Methods and results: Platelet count was measured at baseline in 1989–1990 and at 3 years follow-up, or at baseline (for a newly recruited group) in 1992–1993 in 5766 community-dwelling individuals aged 65 years and older (mean age at baseline, 73 years). During 12–15 years of follow-up, there were 821 incident myocardial infarctions, 807 ischemic strokes, 161 hemorrhagic strokes, 159 venous thrombotic events, and 3413 participants died. Platelet count was not associated with the occurrence of myocardial infarction, ischemic or hemorrhagic stroke, venous thrombosis, or cardiovascular mortality. Non-cardiovascular mortality was higher among both participants with low and with high platelet count. Adjusted non-cardiovascular mortality rates for platelet counts below 100, 100–199, 300–399, and above 400 × 10⁹ L⁻¹ relative to the reference mortality rate in participants with platelet count values between 200 and 299 × 10⁹ L⁻¹ were 1.89 (1.21–2.96), 1.08 (0.98–1.20), 1.20 (1.06–1.37), and 1.47 (1.14–1.90), respectively. Conclusion: Platelet counts were not associated with vascular outcomes but low and high platelet counts were associated with non-cardiovascular mortality, including cancer mortality.     

How factor VIIa works in hemophilia

Summary.  The influence of elevated platelet concentration and recombinant factor VIIa (rFVIIa) on thrombin generation at 5 pM tissue factor (TF) in a synthetic mixture corresponding to hemophilia B (SHB) and 'acquired' hemophilia B blood (AHBB) produced in vitro by an antifactor IX antibody was evaluated. (a) Thrombin generation in SHB and AHBB was delayed and reduced; (b) with 10 nM rFVIIa or 5× normal platelets (10 × 10⁸/mL) SHB and AHBB showed a slight increase in thrombin generation; (c) in the absence of TF, almost no thrombin generation was detected in SHB and AHBB in the presence of 10 nM rFVIIa and 10 × 10⁸/mL activated platelets (5× normal); (d) with TF, 10 nM rFVIIa and 3–5× normal nonactivated platelets (6–10 × 10⁸/mL), thrombin levels approaching normal values were attained. FVIIa appears to function effectively and locally by the combined effect of TF expression and platelet accumulation at the site of a vascular lesion.     

P-selectin antagonism reduces thrombus formation in humans

Summary.  Background:  Interaction of P-selectin with its glycoprotein ligand (P-selectin glycoprotein ligand type 1) mediates inflammatory processes that may also include vascular thrombosis. Platelet P-selectin expression is increased in patients with coronary heart disease, and its antagonism represents a potential future therapeutic target for the prevention and treatment of atherothrombosis. Aim:  To investigate the effects of the novel small molecule P-selectin antagonist PSI-697 on thrombus formation in humans. Methods and Results:  In a double-blind randomized crossover study, thrombus formation was measured in 12 healthy volunteers, using the Badimon ex vivo perfusion chamber under conditions of low and high shear stress. Saline placebo, low-dose (2  m ) and high-dose (20  m ) PSI-697 and the glycoprotein IIb–IIIa receptor antagonist tirofiban (50 ng mL⁻¹) were administered into the extracorporeal circuit prior to the perfusion chamber. As compared with saline placebo, blockade of platelet glycoprotein IIb–IIIa receptor with tirofiban produced 28% and 56% reductions in thrombus formation in the low-shear and high-shear chambers, respectively. PSI-697 caused a dose-dependent, but more modest, reduction in thrombus formation. Low-dose PSI-796 (2  m ) reduced total thrombus area by 14% ( P  = 0.04) and 30% ( P  = 0.0002) in the low-shear and high-shear chambers, respectively. At the high dose (20  m ), PSI-697 reduced total thrombus area by 18% ( P  = 0.0094) and 41% ( P  = 0.0008) in the low-shear and high-shear chambers, respectively. Conclusions:  P-selectin antagonism with PSI-697 reduces ex vivo thrombus formation in humans. These findings provide further evidence that P-selectin antagonism may be a potential target for the prevention and treatment of cardiovascular disease.     

Human platelet glycoprotein VI function is antagonized by monoclonal antibody-derived Fab fragments

Summary.  Platelet interactions with adhesive ligands exposed at sites of vascular injury initiate the normal hemostatic response but may also lead to arterial thrombosis. Platelet membrane glycoprotein (GP)VI is a key receptor for collagen. Impairment of GPVI function in mice results in a long-term antithrombotic protection and prevents neointimal hyperplasia following arterial injury. On the other hand, GPVI deficiency in humans or mice does not result in serious bleeding tendencies. Blocking GPVI function may thus represent a new and safe antithrombotic approach, but no specific, potent anti-GPVI directed at the human receptor is yet available. Our aim was to produce accessible antagonists of human GPVI to evaluate the consequences of GPVI blockade. Amongst several monoclonal antibodies to the extracellular domain of human GPVI, one, 9O12.2, was selected for its capacity to disrupt the interaction of GPVI with collagen in a purified system and to prevent the adhesion of cells expressing recombinant GPVI to collagen and collagen-related peptides (CRP). While 9O12.2 IgGs induced platelet activation by a mechanism involving GPVI and FcγRIIA, 9O12.2 Fab fragments completely blocked collagen-induced platelet aggregation and secretion from 5 µg mL⁻¹ and fully prevented CRP-induced activation from 1.5 µg mL⁻¹. 9O12.2 Fabs also inhibited the procoagulant activity of collagen-stimulated platelets and platelet adhesion to collagen in static conditions. Furthermore, 9O12.2 Fabs impaired platelet adhesion, and prevented thrombi formation under arterial flow conditions. We thus describe here for the first time a functional monoclonal antibody to human GPVI and demonstrate its effect on collagen-induced platelet aggregation and procoagulant activity, and on thrombus growth.     

Cisplatin-induced renal effects and thromboxane A2 receptor blockade

The aim of this study was to evaluate the renal protective effect of linotroban, a thromboxane A₂ receptor antagonist, in 25 patients with malignant tumours scheduled for cisplatin therapy. Cisplatin was administered 1 h after the start of a 24-h continuous infusion of linotroban or placebo. Glomerular filtration rate and effective renal plasma flow were measured. Infusions of cisplatin decreased glomerular filtration rate by 17 ± 25 mL min⁻¹ ( P  = 0.049 vs. baseline) and effective renal plasma flow by 94 ± 150 mL min⁻¹ ( P  = 0.049 vs. baseline) in the placebo group. In the linotroban group a decrease in glomerular filtration rate by 11 ± 18 mL min⁻¹ ( P  = 0.050 vs. baseline) and in effective renal plasma flow by 26 ± 63 mL min⁻¹ ( P  = 0.2 vs. baseline) was noted. However, no difference was noted between groups in response to treatment. Our findings indicate that linotroban may not be useful for prevention of cisplatin's acute nephrotoxic effects.     

A snake venom metalloproteinase, kistomin, cleaves platelet glycoprotein VI and impairs platelet functions

Summary.  Background and objectives:  Injuries to the vessel wall and subsequent exposure of the matrix of the subendothelial layer resulted in thrombus formation. Platelet glycoprotein (GP) Ib and VI play a crucial role in matrix-induced activation and aggregation of platelets. Methods and results:  In the present study, we reported that the GPIb-cleaving snake venom metalloproteinase (SVMP), kistomin, inhibited collagen-induced platelet aggregation. Moreover, kistomin inhibited platelet aggregation induced by convulxin (CVX, a GPVI agonist) and a GPVI-specific antibody in a concentration and time-dependent manner. Kistomin treatment decreased platelet GPVI but not integrin α2β1 and αIIbβ3, accompanied with the formation of GPVI cleavage fragments, as determined by flow cytometric and Western blot analyses. In addition, intact platelet GPVI and recombinant GPVI were digested by kistomin to release 25- and 35-kDa fragments, suggesting that kistomin cleaved GPVI near the mucin-like region. We designed four synthetic peptides ranging from Leu¹⁸⁰ to Asn²⁴⁹ as the substrates for kistomin and found that kistomin cleaved these synthetic peptides at FSE²⁰⁵/A²⁰⁶TA and NKV²¹⁸/F²¹⁹TT, as analyzed by MALDI-TOF-MS. In addition, GPVI-specific antibody-induced tyrosine kinase phosphorylation in platelets was reduced after kistomin pretreatment, and platelet adhesion to collagen but not to fibrinogen was attenuated by kistomin. Conclusions:  We provided here the first evidence that a P-I snake venom metalloproteinase, kistomin, inhibits the interaction between collagen and platelet GPVI through its proteolytic activity on GPVI, thus providing an alternative strategy for developing new anti-thrombotic agents.     

Stimulation of arginine vasopressin secretion by a small increase in blood ionized calcium in normal men

Evidence has been provided for an increase in baseline serum corticotrophin (ACTH) levels in response to a rise in circulating ionized calcium (Ca_i) levels within the physiological range. In order to establish whether small Ca_i increments are also able to modify the basal secretion of arginine vasopressin (AVP), we infused calcium gluconate through an intravenous infusion pump in eight healthy male subjects (25–31 years old). Serum Ca_i, ACTH and AVP concentrations were measured every 10 min over an infusion period lasting 90 min. A significant progressive rise in serum Ca_i (baseline: 42 ± 0.9 mg dL⁻¹; 90 min: 47.2 ± 0.9 mg dL⁻¹, P  < 0.001), ACTH (baseline: 30.7 ± 1.3 pg mL⁻¹; mean peak at 80 min: 37.4 ± 2.4 pg mL⁻¹, P  < 0.01) and AVP levels (baseline: 2.1 ± 0.6 pg mL⁻¹; mean peak at 80 min: 3.2 ± 0.5 pg mL⁻¹, P  < 0.01) was observed during calcium infusion. Furthermore, a significant positive correlation ( r  = 0.71; P  < 0.001) was observed between ACTH and AVP responses to calcium infusion at 60, 70, 80 and 90 min. These data demonstrate that AVP secretion is stimulated by a slight rapid increase in serum Ca_i levels even though absolute serum Ca_i levels remain within the normal range. In addition, the positive correlation between Ca_i-induced ACTH and AVP increments suggests that AVP plays a releasing role on ACTH secretion during calcium infusion.     

Prolonged procoagulant activity on overstretch-injured coronary arteries in pigs

Summary.  This study was designed to assess the time course and nature of the vascular procoagulant response after 1.5-fold balloon overstretch injury of the coronary arteries in pigs. Arteries were excised for chromogenic assay of bound factor (F)Xa and thrombin at 24 h, 3 days, 1 week, or 2 weeks after injury. FXa at the site of injury remained elevated for 1 week (4.9 ± 5.9 µg cm⁻², n  = 10), compared with non-injured control arteries (0.4 ± 0.2 µg cm⁻², n  = 18, P  = 0.00025), while thrombin was increased only at 24 h. Tissue factor protein was abundant in non-injured coronaries (10 ± 6 ng µg⁻¹ total protein, n  = 9) and levels were unchanged by injury (13 ± 11 ng µg⁻¹, n  = 6) or 24-h administration of tissue factor pathway inhibitor (16 ± 6 ng µg⁻¹, n  = 6). Persistent tissue factor-mediated procoagulant activity may explain the need for prolonged anticoagulation to attenuate neointimal formation after balloon-induced coronary injury.     

Glycoprotein IIb/IIIa inhibition attenuates platelet-activating factor-induced platelet activation by reducing protein kinase C activity

Summary.  Glycoprotein (GP)IIb/IIIa inhibition may abolish activated leukocyte-induced platelet activation, in which leukocyte-released platelet-activating factor (PAF) is a major mediator. The present study thus investigated if and how GPIIb/IIIa inhibitors interfere with PAF-induced platelet activation. Platelet and leukocyte activation were monitored by flow cytometry and immunoblotting. GPIIb/IIIa inhibitors (c7E3, non-peptide SR121566, and MAb RFGP56) attenuated PAF-induced, but not adenosine diphosphate (ADP)- or thrombin receptor activating peptide ( TRAP)-induced platelet P-selectin expression in whole blood. GPIIb/IIIa blockade enhanced ADP- or TRAP-induced leukocyte CD11b expression, but not the response to PAF. GPIIb/IIIa blockade attenuated PAF-induced, but enhanced ADP- or TRAP-induced platelet–leukocyte aggregation. Under the present experimental conditions, thromboxane A₂ receptor antagonism did not significantly influence PAF-induced platelet activation, and GPIIb/IIIa inhibition did not interfere with calcium mobilization/influx in platelets. Protein kinase C (PKC) blockade inhibited PAF-induced platelet P-selectin expression, and PAF-induced PKC activity was reduced by GPIIb/IIIa inhibition. PAF (=1 µ m ) did not induce MEK 1/2 or ERK 1/2 phosphorylation, whilst thrombin induced marked responses, which were enhanced by GPIIb/IIIa blockade. Thus, GPIIb/IIIa inhibition attenuates PAF-induced platelet activation via inhibiting PKC activity. GPIIb/IIIa blockade enhances thrombin-induced platelet MEK 1/2 and ERK 1/2 activation, and augments ADP- and TRAP-induced leukocyte activation by enhancing platelet–leukocyte aggregation.     

Liposomal dexamethasone effectiveness in the treatment of hypersensitivity pneumonitis in mice

Abstract. The effects of daily intranasal instillation of liposomal dexamethasone and free dexamethasone phosphate were compared in a murine model of hypersensitivity pneumonitis induced by Saccharopolyspora rectivirgula (formally known as Micropolyspora faeni ). After 3 weeks of antigen and liposome instillations, lung response was evaluated by bronchoalveolar lavage cell counts, lung index and histopathology. Systemic absorption was evaluated by measuring plasma adrenocorticotropic hormone (ACTH) level. Free dexamethasone phosphate induced a dose-dependent response with the maximal effect reached at 1 mgkg^-1. At 0.1 mgkg^-1, liposomal dexamethasone had a greater effect than free dexamethasone phosphate on bronchoalveolar cells ml^-1: 3.01 × 10⁵± 0.35× 10⁵ compared to 4.70×10⁵± 0.34 × 10⁵, and lung index: 1.22 ±0.10 compared to 1.86 ± 0.07. Effect on histopathology was similar. Plasma ACTH levels (pg ml^-1) were: 75.1 ± 14.0 for animals receiving antigen and free dexamethasone phosphate (0.2 mg kg^-1), and 149.7 ± 12.0 for animals receiving antigen and liposomal dexamethasone (0.2 mg kg^-1). In conclusion, liposome-incorporated dexamethasone is efficient in the treatment of experimental hypersensitivity pneumonitis and, contrarily to free dexamethasone phosphate, does not inhibit ACTH secretion.     

Low-density lipoprotein-induced formation of nitric oxide by cultured vascular smooth muscle cells of the rat

There is evidence that low-density lipoprotein (LDL) plays a crucial role in atherogenesis. On the cellular level, LDL has been shown to activate a number of mechanisms involved in atherogenesis and vasoconstriction. Local immoderate vasoconstriction is physiologically antagonized by nitric oxide, which is released from the endothelium. To find out whether LDL also influences the synthesis of nitric oxide in vascular smooth muscle cells, both the conversion of arginine to citrulline and the production of nitrite were determined as a measure of nitric oxide formation. After incubation of rat vascular smooth muscle cells with native LDL (25 μg mL⁻¹) for 24 h, the production of both l -[¹⁴C]-citrulline [39 600 (3600) cpm mg⁻¹ cell protein] and nitric oxide [2.95 (0.56) μmol L⁻¹] were about twice and 1.5-fold the amount of the corresonding values in untreated cells (mean ± SD, P  < 0.05, n  = 4). Oxidized LDL was less effective than the native form. The presence of the arginine analogue N ^G-methyl- l -arginine reduced citrulline production dose-dependently but augmented DNA synthesis, both induced by LDL. In addition, the lipoprotein caused a 1.6-fold increase in cyclic GMP production following a 24-h incubation [control = 10.9 (3.8) pmol mg⁻¹ cell protein, P  = 0.016]. The results suggest that native LDL might partly impair its atherogenic potential on the vasculature by stimulating the production by smooth muscle cells of both nitric oxide and cyclic GMP.     

Plasma endothelin-1 levels during transient acute myocardial ischaemia in men: effects of coronary revascularization

The endothelium-derived peptide endothelin-1 (ET-1) was evaluated in 14 male patients [mean age 52.74 years (SEM 1.10)] affected by coronary artery disease during a bicycle electrocardiographic stress test and dipyridamole echocardiogram. Both tests were performed before and after coronary revascularization. Fourteen healthy male subjects served as controls [mean age 53.21 years (SEM 1.63)]. Baseline plasma endothelin-1 levels were higher ( P  < 0^.0001) in ischaemic patients [1^.81 pg mL⁻¹ (0^.15, n  = 14)] than in control subjects [0^.61 pg mL⁻¹ (0^.03, n  = 14)], but did not increase with exercise in both groups. Similar results were obtained with dipyridamole infusion. Endothelin-1 levels significantly decreased after coronary revascularization [before: mean 1^.81 pg mL⁻¹ (SEM 0^.15, n  = 14); after: mean 1^.16 pg mL⁻¹ (SEM 0^.11), P  < 0^.002], without changes in the peptide response to both tests. In conclusion, elevated plasma endothelin-1 concentrations were found in patients with stable angina compared with non-ischaemic subjects. No changes were observed during exercise or dipyridamole infusion in both groups. Coronary revascularization was followed by a significant decrease in plasma endothelin-1 levels.     

Inhibition of paracrine angiotensin-converting enzyme in vivo: effects on interstitial glucose and lactate concentrations in human skeletal muscle

Microdialysis was used to selectively assess the effect of the paracrine renin–angiotensin system (RAS) on interstitial glucose and lactate concentration profiles in skeletal muscle of healthy volunteers ( n  = 8) during basal and insulin-stimulated conditions. Paracrine RAS was selectively inhibited by local retrodialysis with enalaprilate. Under basal conditions, local administration of enalaprilate (2 μg mL⁻¹) increased interstitial dialysate glucose concentration from 0.71 ± 0.14 mmol L⁻¹ to 0.84 ± 0.14 mmol L⁻¹ and decreased the serum interstitial gradient (SIG_glu) compared with baseline ( P  < 0.02). Under clamp conditions, enalaprilate, even at the lowest concentration (0.02 μg mL⁻¹), increased interstitial dialysate glucose concentration from 0.77 ± 0.11 mmol L⁻¹ to 1.02 ± 0.09 mmol L⁻¹ and decreased SIG_glu compared with baseline ( P  < 0.01). Interstitial lactate concentrations slightly increased during basal as well as during clamp conditions ( P  < 0.05 vs. baseline). Selective inhibition of paracrine muscle angiotensin-converting enzyme (ACE) increases interstitial glucose and lactate concentrations and decreases SIG_glu in muscle by facilitating transcapillary glucose transport. This effect is more pronounced during hyperinsulinaemia and may be of clinical relevance in diabetic patients treated with therapeutic doses of enalapril.     

Vasoactive substances in early dumping syndrome: effects of dumping provocation with and without octreotide

In patients after gastric surgery, early dumping symptoms can be provoked by oral glucose challenge. Octreotide effectively prevents the occurrence of dumping symptoms. We have studied plasma renin activity (PRA), aldosterone and atrial natriuretic peptide (ANP) concentrations in nine patients with early dumping, 10 surgical control subjects and nine healthy control subjects after an oral glucose challenge preceded by either placebo or 25 μg of octreotide subcutaneously (s.c.). In the dumping group, basal PRA was signifi-cantly ( P  < 0.01) higher (3.9 ± 0.6 μg L⁻¹ h⁻¹) than in either surgical or healthy control subjects (1.1 ± 0.3 μg L⁻¹ h⁻¹ and 1.1 ± 0.2 μg L⁻¹ h⁻¹ respectively) and showed a significant rise after glucose ingestion to 5.4 ± 0.9 μg L⁻¹ h⁻¹ that did not occur in control subjects. Aldosterone concentration showed a concomitant rise. In dumping patients, plasma ANP decreased after glucose ingestion from 31 ± 6 ng L⁻¹ to 21 ± 5 ng L⁻¹ ( P  < 0.05). This decrease did not occur in control subjects. Early dumping is associated with an activation of the renin–aldosterone axis and a decrease in plasma ANP, reflecting a hypovolaemic state. Octreotide prevents the occurrence of these changes.     

Variability of platelet aggregate dispersal with glycoprotein IIb–IIIa antagonists eptifibatide and abciximab

Summary.  Background:  Utilization of glycoprotein IIb–IIIa (GPIIb–IIIa) inhibitors improves outcomes of patients with acute coronary syndromes (ACS), including those undergoing percutaneous coronary intervention (PCI). These results may be related to the ability of the inhibitors to destabilize coronary thrombi, reduce microembolization, and restore vessel patency. Objective:  To evaluate in vitro the ability of GPIIb–IIIa antagonists, abciximab and eptifibatide, to promote the disaggregation of platelet-rich thrombus. Methods: Antagonist-induced disaggregation was assayed in plasma by aggregometry, as well as in whole blood by point of care and capillary perfusion systems. Fibrinogen dissociation from the platelet surface was quantified by flow cytometry. Results:  Significant disaggregation of 5 μ m ADP-induced aggregates was observed after addition of either agent. The maximum extent and rate of disaggregation were significantly higher with eptifibatide than with abciximab. Both antagonists also dispersed 2 μg mL⁻¹ collagen-induced aggregates, again with eptifibatide having a greater effect. Eptifibatide, but not abciximab (up to 10 μg mL⁻¹), was efficient at dissociating aggregates to single platelets in whole blood and dispersing aggregates that had been aged for 30 min before treatment. Eptifibatide also reduced existing thrombus burden in the perfusion model under arterial flow conditions. A key mechanism of aggregate dispersal was antagonist-induced displacement of platelet-bound fibrinogen, which was greater with eptifibatide, a competitive inhibitor of fibrinogen binding, than with the noncompetitive inhibitor, abciximab. Conclusions:  These results suggest that drug concentration and residence time, along with thrombus extent and age, may be critical determinants in promoting timely recanalization.     

Glycaemic control and in vivo non-oxidative Maillard reaction: urinary excretion of pyrraline in diabetes patients

The presence of pyrraline in human urine has recently been described. Using reversed-phase high-performance liquid chromatography, we measured urinary pyrraline in 45 insulin-treated diabetic patients with preserved renal function and in 30 age- and sex-matched healthy subjects. The relationship between urinary pyrraline and metabolic control parameters in the diabetic population (glycaemia, fructosamine, haemoglobin Al_c, and 1-year mean haemoglobin A1_c) was evaluated. The mean urinary level of pyrraline in diabetic patients with poor glycaemic control (HbA_1c > 9.5%) was higher than that in healthy subjects (1.12 ± 0.35 vs. 0.75 ± 0.2 μmol mmol⁻¹ creatinine, P  < 0.04), whereas in patients with good to moderate glycaemic control (HbA1_c < 9.5) it was slightly but not significantly higher than in healthy subjects (0.80 ± 0.3 μmol mmol⁻¹ creatinine vs. 0.75 ± 0.2 μmol mmol⁻¹ creatinine). There is a significant correlation between urinary pyrraline level and glycaemia ( P  < 0.008), haemoglobin A1_c ( P  < 0.01) and 1-year mean haemoglobin A1_c values ( P  < 0.007), but not with fructosamine. The results of the present work prove, for the first time, that glycaemic status influences circulating levels of advanced Maillard reaction products.     

The role of tirofiban in acute coronary syndromes

Platelet activation and aggregation play an important and essential role in the formation of intracoronary thrombus in acute coronary syndromes (ACS). ACS still carries unacceptably high rates of morbidity and mortality despite intensive antianginal therapy and the wide use of aspirin and heparin. Two glycoprotein IIb/IIIa receptor inhibitors are now licensed for concomitant use with heparin and aspirin in ACS. Glycoprotein IIb/IIIa receptor inhibitors block the final step for platelet aggregation and fibrinogen binding, thus preventing thrombus formation. Tirofiban is a potent, synthetic, non-peptide and specific glycoprotein IIb/IIIa receptor inhibitor. In three major international trials involving over 7200 patients (PRISM, PRISM-PLUS and RESTORE), tirofiban was shown to be well tolerated and to reduce the risk of ischaemic complications in patients with unstable angina, non-Q-wave myocardial infarction and high-risk patients undergoing revascularisation when used in combination with aspirin and heparin. These and ongoing studies are discussed.     

Maintenance of in vitro properties of leukoreduced whole blood–derived pooled platelets after a 24-hour interruption of agitation

BACKGROUND: Continuous agitation during platelet concentrate (PC) storage is frequently interrupted during shipping. Studies have evaluated the effects of interrupted agitation in apheresis and single whole blood–derived PCs, but not PC pools. This study evaluated in vitro properties of pooled whole blood–derived platelets (PLTs) after a 24-hour interruption of agitation.
STUDY DESIGN AND METHODS: Eleven ABO-identical leukoreduced whole blood–derived PCs (Leukotrap RC-PL, Pall), pooled in a transfer container, were equally divided into each of two CLX-HP containers (Acrodose PL, Pall). One pool (test) was held in a shipping container unagitated for 24 hours between Day 2 and Day 3, while the other (control) was continuously agitated.
RESULTS: Ten pairs underwent in vitro assays after 5 and 7 days' storage. Pools contained a mean (±SD) of 5.0 × 10¹¹ ± 0.4 × 10¹¹ PLTs. Interrupting agitation for 24 hours reduced test pool pH versus control after 5 days' storage (6.77 ± 0.15 vs. 6.98 ± 0.06, p = 0.0005). Test and control pH differences were greater after 7 days' storage (6.17 ± 0.29 vs. 6.65 ± 0.14, p < 0.0001); 5 of 10 test pool pHs were less than 6.2 (vs. 0 of 10 controls). Other test pool key in vitro variables were reduced compared with controls after 5 days' storage, with greater differences after 7 days.
CONCLUSION: After 5 days' storage, pooled leukoreduced whole blood–derived-PCs in CLX-HP containers adequately maintained pH and other key in vitro variables after a 24-hour interruption of agitation. After 7 days' storage, 5 of 10 pools did not maintain a pH value of 6.2 or greater while matched continuously agitated units did.     

Characterization of a tissue factor/factor VIIa-dependent model of thrombosis in hypercholesterolemic rabbits

Summary.  Tissue factor (TF) expressed in arterial atherosclerotic plaque plays a key role in activating the extrinsic coagulation pathway and triggering acute coronary syndromes. In this study, we developed and characterized a TF–factor (F)VIIa-mediated thrombosis model in rabbits. Balloon catheter-induced endothelial denudation in the femoral artery and a 4-week high cholesterol diet produced a localized atherosclerotic plaque at the injured site. High levels of TF mRNA and TF protein antigen (152 ± 25 vs. 49 ± 12 pg mg⁻¹ protein in normal vessels) were detected in these atherosclerotic plaques. Plasma FVII coagulant activity (FVII:C) was significantly increased in the hypercholesterolemic rabbits (36 ± 1 s) compared with the normal rabbits (44 ± 1 s, P  < 0.0001). Plaque rupture was induced by balloon angioplasty, which resulted in thrombus formation in the injured vessel segment after a brief period of stasis. FVIIai, a specific TF–FVIIa inhibitor, was administered intravenously to rabbits before plaque rupture at 0.3 and 1.0 mg kg⁻¹. FVIIai dose-dependently reduced thrombus mass (14.7 ± 2.5 and 5.9 ± 2.2 mg, respectively, vs. 21.6 ± 1.9 mg in the control group). PD198961, a novel factor Xa inhibitor, and argatroban, a thrombin inhibitor, also dose-dependently inhibited thrombosis. These results indicate that thrombus formation in this model is initiated by the activation of TF–FVIIa pathway, which is attributed to TF expression in the atherosclerotic plaque and enhanced plasma FVII coagulant activity. This model may be useful for evaluating in vivo efficacy of new antithrombotic drugs, particularly TF–FVIIa inhibitors.