Monoclonal antibody-induced inhibition of platelet function: effects on hemostasis and vascular graft thrombosis in baboons

The usefulness of antiplatelet agents in vascular graft recipients remains controversial because of uncertainties regarding drug mechanisms of action and dose-duration effects. In our study, a well-characterized murine monoclonal antibody (LJ-CP8, 10 mg/kg) was infused into baboons to assess the hemostatic consequences and antithrombotic effectiveness of blocking the platelet glycoprotein IIb-IIIa receptor for fibrinogen and other adhesive glycoproteins. Five treated animals and six control animals were evaluated with serial measurements of platelet count, bleeding time, and platelet aggregation ex vivo (in response to adenosine diphosphate and collagen). Indium 111-labeled platelet deposition onto femoral vascular grafts (4 mm inner diameter Gore-Tex) implanted immediately after antibody infusion was measured by quantitative gamma camera imaging. Although the antibody did not alter circulating platelet counts, bleeding times were immediately prolonged to more than 30 minutes (vs. 4.8 +/- 0.4 minutes pretreatment) with only partial normalization by 48 hours (8.3 +/- 1.0 minutes, p less than 0.05). Platelet aggregation in response to both collagen and adenosine diphosphate was abolished immediately and remained impaired for 48 hours. Despite the profound inhibition of platelet function, graft platelet deposition was equivalent postoperatively in both the treated and untreated groups (p greater than 0.5), averaging approximately 5 x 10(9) platelets per graft. Graft-associated indium 111-labeled platelet activity increased over 48 hours and was not reduced by the antibody treatment (p greater than 0.5 at all times). All grafts were removed at 8 days; only one graft from a treated animal was found patent.(ABSTRACT TRUNCATED AT 250 WORDS)     

Aspirin-independent antithrombotic effects of acutely attached cultured endothelial cells on endarterectomized arteries

We have compared the acute antithrombotic effects of aspirin-treated versus normal endothelial cell (EC) coverage of endarterectomized baboon aortic segments (EAS) incorporated into chronic exteriorized arteriovenous shunts in baboons. Human ECs grown in culture were incubated in control medium or medium containing aspirin (100 mumols/ml) and then attached at saturation density by incubating EC suspensions (6 x 10(5) cells/100 microliters) within EAS for 20 minutes. Nonendarterectomized aortic segments and untreated EAS served as negative and positive controls, respectively. The inhibitory effect of aspirin treatment on EC production of prostacyclin was confirmed by radioimmunoassay of its stable metabolic breakdown product, 6-keto-prostaglandin F1 alpha in supernatant medium. Thrombus formation in vivo was measured as the accumulation of indium 111-labeled platelets on endarterectomy sites in real time by scintillation camera imaging. 111In-labeled platelets were deposited rapidly, reaching a plateau by 60 minutes of 4.40 +/- 0.89 x 10(9) platelets/cm, compared with 111In-labled platelet deposition on nonendarterectomized segments of 0.89 +/- 0.26 x 10(9) platelets/cm (p = 0.008). Coverage of EAS with normal cultured ECs significantly reduced platelet deposition on EAS (1.05 +/- 0.45 x 10(9) platelets/cm; p = 0.009 at 1 hour compared with EAS not incubated with ECs). Aspirin-treated ECs also produced a marked reduction in platelet disposition (0.71 +/- 0.24 x 10 platelets/cm; p = 0.007 compared with EAS without ECs) that was equivalent to the effect of non-aspirin-treated ECs (p greater than 0.5). Scanning and transmission electron microscopy confirmed the antithrombotic effects of attached ECs. We conclude that endarterectomy of normal arteries produces a highly thrombogenic surface and the thrombogenicity is abolished by acutely attaching cultured human ECs.     

The hemostatic effect of transfusing fresh whole blood versus platelet concentrates after cardiac operations

The major cause of nonsurgical bleeding after cardiopulmonary bypass is delayed recovery of platelet count and function. Recovery of platelet count and function was compared in 27 patients who were randomized preoperatively to receive after cardiopulmonary bypass either 1 unit of fresh whole blood (15 patients) or 10 units of platelet concentrates (12 patients). Platelet count, bleeding time, platelet aggregation (adenosine diphosphate, collagen, epinephrine, and ristocetin) and platelet thromboxane formation were abnormal after cardiopulmonary bypass in all the patients. The increase of platelet count after 1 unit of fresh whole blood (from 115 +/- 32 X 10(9)/L to 148.5 +/- 36 X 10(9)/L) was similar to that achieved by 4 units of platelets (from 140 +/- 61 X 10(9)/L to 171 +/- 60 X 10(9)/L). The increase was doubled after 10 platelet units (from 140 +/- 61 X 10(9)/L to 209 +/- 55 X 10(9)/L). Bleeding time returned to normal values after fresh whole blood or after 8 platelet units. However, platelet thromboxane formation was higher after 1 unit of fresh whole blood than after 10 platelet units (95 +/- 25 versus 46 +/- 35 ng/ml, p less than 0.05), as was platelet aggregation response to collagen and epinephrine. The 24-hour blood loss was smaller in the fresh whole blood group (560 +/- 420 ml versus 770 +/- 360 ml), although the difference was not statistically significant. The results suggest that the hemostatic effect of 1 unit fresh whole blood after cardiopulmonary bypass is at least equal, if not superior, to the effect of 10 units of platelets.     

Loss of platelet fibrinogen receptors during clinical cardiopulmonary bypass

In 10 patients, cardiopulmonary bypass decreased the number of fibrinogen binding sites from 31,730 +/- 12,802 per platelet to 18,590 +/- 9,644 per platelet. Bypass also decreased the amount of the platelet membrane glycoprotein IIIa, which is part of the fibrinogen receptor complex, from 17.1 +/- 3.6 ng/10(9) platelets to 12.9 +/- 4.7. The fibrinogen binding constant did not change. Platelet sensitivity to adenosine diphosphate did not change; however, template bleeding times increased from 5.2 +/- 1.5 minutes before bypass to 8.5 +/- 2.3 minutes after bypass. Analysis of detergent washings from the perfusion circuit after bypass in five patients indicated that platelet material remains attached to the surface as membrane fragments and degranulated platelets. These data further elucidate the mechanism of platelet loss and dysfunction during cardiopulmonary bypass and highlight the importance of platelet membrane fibrinogen receptors and surface adsorbed fibrinogen in this process.     

Effect of recombinant human erythropoietin (rHuEpo) on the hemostatic system in chronic hemodialysis patients.

Hemostatic measurements were undertaken in eight chronic hemodialysis uremic patients on recombinant human erythropoietin (rHuEpo). Same measurements were repeated in another seven patients in whom anemia was corrected by the transfusion of red blood cells. The correction of the anemia by rHuEpo therapy was accompanied by 1. correction of the prolonged Simplate Bleeding Time (BT) to normal less than 10.0, minutes after 16 weeks of rHuEpo treatment; 2. significant increases in the levels of fibrinogen, clotting FVIII:C, vWF:antigen, vWF:ristocetin cofactor and platelet count; 3. enhanced aggregation responses to ADP, adrenaline, arachidonic acid, collagen and ristocetin. There was no significant fluctuation in other coagulation parameters PT, APTT, TT, reptilase time and antithrombin III and plasma fibrinogen. In patients who were treated with RBC transfusion and despite the correction of the anemia, the bleeding time shortened significantly but not corrected, mean BT before and after RBC transfusion was 17.1 +/- 1.4 and 11.6 +/- 1.9 minutes respectively. Besides there was significant elevation of vWF:Ricofactor levels but not FVIII:C, vWF:Ag or platelet count. Platelet aggregation responses to ADP remained unchanged. It is concluded that significant elevations of FVIII:related activities, plasma fibrinogen, improved platelet aggregability and correction of the BT are salient hemostatic changes that follow rHuEpo therapy in uremic patients.     

Preformed confluent endothelial cell monolayers prevent early platelet deposition on vascular prostheses in baboons

We assessed the capacity of preformed confluent endothelial cell (EC) monolayers on small-caliber prosthetic grafts to prevent early platelet deposition in a baboon model. Cultured human umbilical vein ECs were attached to expanded polytetrafluoroethylene (Gore-Tex, 4 mm inner diameter, 3 cm length) precoated with type I collagen and perfused in vitro for 2 hours at 15 ml/min with serum-containing culture medium to achieve cell spreading into confluent monolayers. Cell numbers were quantified by deoxyribonucleic acid assay or isotopic counting of indium 111-labeled ECs. Saturation density for cell attachment was 3.55 +/- 0.29 x 10(5) cells per square centimeter of graft. After 1 hour of in vitro perfusion at 100 ml/min, 92.8% +/- 1.8% of cells remained attached and the flow surface was morphologically confluent. When grafts were inserted as extension segments into arteriovenous silicone rubber (Silastic) shunts in baboons, thereby exposing the endothelialized grafts to native flowing blood (100 ml/min) for 1 hour, the EC monolayers remained confluent with 81.05% +/- 5.88% of the cells attached. Indium 111-labeled platelet deposition onto grafts was quantified by dynamic scintillation camera imaging. Platelet deposition on 10 endothelialized grafts was markedly reduced (0.16 +/- 0.04 x 10(9) platelets per graft) compared with 10 untreated control grafts (1.84 +/- 0.59 x 10(9) platelets, p less than 0.02), eight grafts with early attached unspread ECs (2.38 +/- 0.66 x 10(9) platelets, p less than 0.005), and 11 grafts treated with collagen alone (5.93 +/- 0.72 x 10(9) platelets, p less than 0.002).(ABSTRACT TRUNCATED AT 250 WORDS)     

The effects of heparin and extracorporeal circulation on platelet counts and platelet microaggregation during cardiopulmonary bypass

BACKGROUND: Cardiopulmonary bypass is associated with platelet activation and reduced platelet counts. Platelet activation may artifactually lower platelet counts by causing aggregation. In vivo platelet activation may increase existent platelet microaggregation ex vivo. We studied platelet counts and existent platelet microaggregation at different stages of cardiopulmonary bypass. METHODS: Twenty-one patients were studied before and after heparinization (300 U. kg(-1)) and at the end of cardiopulmonary bypass. Unaggregated (or single) platelets were counted in hirudin-anticoagulated blood, and total platelets were counted in ethylenediaminetetraacetic acid-anticoagulated blood. RESULTS: The total platelet count, 198 +/- 61 x 10(9). L(-1), was unaffected by heparin and stayed at 197 +/- 60 x 10(9). L(-1) (P =.7) but fell during extracorporeal circulation; the hemodilution-corrected count was 163 +/- 52 x 10(9). L(-1) (P =.0004). Heparinization reduced the unaggregated platelet count from (mean +/- 1 SD) 178 +/- 62 x 10(9). L(-1) to 155 +/- 60 x 10(9). L(-1) (P =.0001). Extracorporeal circulation had little additional effect. The hemodilution-corrected count was 142 +/- 48 x 10(9). L(-1) (P =.6). CONCLUSIONS: Heparinization caused platelet activation and increased existent platelet microaggregation ex vivo. During extracorporeal circulation, there was a reduction in total platelets that was greater than could be explained by hemodilution alone, but the unaggregated platelet count did not change significantly when corrected for hemodilution. Furthermore, the increased platelet microaggregation observed after heparinization was no longer evident after this loss. These findings suggest that during extracorporeal circulation, the platelets that formed into microaggregates after heparinization were lost from the circulation in preference to single platelets.     

Quantitative study of platelet deposition on expanded polytetrafluoroethylene grafts using 111indium-labeled platelets

The count of platelets deposited on expanded polytetrafluoroethylene grafts (2 cm long, 3 mm internal diameter, implanted into thoracic aortae of 59 rabbits) was measured using 111Indium-labeled autologous platelets. The changes due to the time courses after implantation, the effect of aspirin and the differences of the platelet deposition on the thrombus and thrombus free surface were studied. In non-treated grafts, platelet deposition on the thrombus free surface was significantly and repeatedly decreased on the 3rd (42.3 +/- 16.6 X 10(4)/mm2, mean +/- S.D.), 9th (17.7 +/- 5.3 X 10(4)/mm2), and 30th (6.8 +/- 3.2 X 10(4) mm2) days after the implantation (p less than 0.05). In treated grafts, platelet deposition on the thrombus free surface was significantly inhibited by aspirin on the 3rd day (20.5 +/- 8.4 X 10(4)/mm2) after implantation (p less than 0.03), while significant inhibition of platelet deposition by aspirin was not noted on the 9th (13.4 +/- 3.8 X 10(4)/mm2) and 30th (13.1 +/- 4.5 X 10(4)/mm2) days after the implantation. Platelet deposition on the thrombus was not inhibited by aspirin. (3600 +/- 2400 X 10(4)/mm2, non-treated grafts. 3500 +/- 520 X 10(4)/mm2, treated grafts). It is concluded that administration of aspirin is necessary immediately after implantation, and its effect is not noted at the time when the deposition of the platelet decreased, and the effect of aspirin cannot be expected in tissues where easy platelet aggregation occurs.     

Iloprost infusion decreases skeletal muscle ischemia-reperfusion injury.

Iloprost (a long-acting prostacyclin analog) has been demonstrated to decrease cardiac muscle infarct size after ischemia-reperfusion injury. We investigated the ability of iloprost to decrease skeletal muscle injury and platelet sequestration after ischemia-reperfusion injury in a canine bilateral isolated gracilis muscle model. Anesthesized animals (n = 13) were subjected to 6 hours of gracilis muscle ischemia and 1 hour of reperfusion. Fifteen minutes before muscle reperfusion, the animals were infused with radium 111-labeled autogenous platelets. Experimental animals (n = 7) received a continuous preischemic intravenous infusion of iloprost (0.45 microgram/kg/hr) and two 0.45 microgram/kg intravenous injections of iloprost (10 minutes before the ischemic interval and 10 minutes before reperfusion). Muscle injury was measured with triphenyltetrazolium chloride histochemical staining. Platelet sequestration within ischemic muscle specimens was determined by measuring indium 111 activity in a gamma counter. Iloprost infusion decreased muscle infarct size from 57.0% +/- 12.6% in control animals to 15.8% +/- 4.4% in experimental animals (p less than 0.05). Platelet uptake in experimental and control muscle was 1.2 +/- 0.21 x 10(7) and 2.17 +/- 0.48 x 10(7) platelets/gm ischemic muscle, respectively (p = 0.1). Although platelet sequestration was not altered significantly in this experiment, a reduction in skeletal muscle injury was confirmed. Further investigation on the mechanisms of action of iloprost in chronic and acute skeletal muscle ischemia is warranted.     

Trapidil decreases the aggregation of platelets from heart transplant recipients ex vivo

Heart transplant recipients show platelet hyperaggregability, which may be related to the incidence of graft vasculopathy. We investigated whether trapidil can inhibit the aggregation of platelets from these patients. Platelet count, mean platelet volume (MPV), and adenosine diphosphate (ADP)-induced platelet aggregation were determined in 18 heart transplant recipients and 12 healthy subjects. Additionally, platelet-rich plasma from the patients was incubated with trapidil or with saline, prior to measuring ADP-induced aggregation. The MPV was significantly greater in patients compared to controls (9.4+/-1.1 vs 8.5+/-0.7 fL; P=.01), and ADP-induced platelet aggregation was significantly increased in patients compared to controls (81.2%+/-13.1% vs 69.6%+/-16.2%; P=.04, respectively). The trapidil-treated samples showed significantly decreased platelet aggregation compared to the control samples (24.2%+/-12.6% vs 66.7%+/-11.7%; P<.001). Platelets from heart transplant recipients showed an increased MPV and increased ADP-induced aggregation. Trapidil effectively reduced the ADP-induced aggregation ex vivo.     

Persistent platelet activation by passivated grafts.

Vascular grafts in canines exhibit similar healing patterns to humans in that the graft surface forms a pseudointima over time but endothelializes only near the anastomotic sites. Thus the pseudointima at the midportion of the graft may represent a nidus for persistent platelet activation. The purpose of this investigation was to examine the effect of the maturing graft surface on platelet activation. Long Dacron subcutaneous carotid to aorta grafts (50 cm x 8 mm) were placed in nine dogs. Blood samples were obtained by direct graft puncture, at the proximal and distal ends of the graft, at 1, 24, 48, 72 hours, 1, 2, 3, 4 weeks, and monthly thereafter for 8 months. Seven sham dogs had subcutaneous grafts implanted without arterial anastomoses, and blood samples were drawn from the femoral artery. Platelet counts were determined with a platelet counter. Platelet aggregation and release of adenosine triphosphate was determined with a whole blood aggregometer by use of arachidonic acid, collagen, and adenosine diphosphate as agonists. No difference was found in platelet aggregation to collagen or adenosine diphosphate stimulation across the graft, but platelets released significantly less adenosine triphosphate to collagen and adenosine diphosphate stimulation distally versus proximally. In the graft dogs a decrease in systemic platelet counts of 50% occurred from the preoperative level which persisted over 8 months (p less than 0.01). Also less response occurred to collagen and ADP stimulated platelet aggregation in the graft animals than the sham animals during the first month of study. These data suggest that significant platelet-graft interactions occur even after the graft has formed a mature pseudointima.     

Increased platelet aggregation and production of platelet-derived microparticles after surgery for upper gastrointestinal malignancy

BACKGROUND: Since platelet function is known to play a major role in arterial thrombosis, we investigated postsurgery alterations in platelet function that might predispose patients with upper gastrointestinal malignancy to postoperative thrombotic complications. SUBJECTS AND METHODS: Shear-induced platelet aggregation (SIPA) in platelet-rich plasma was measured in 23 patients who elected to undergo abdominal surgery. Measurement was done by cone-plate viscometer under low shear stress (12 dyn/cm(2)), a physiological condition, and under high shear stress (108 dyn/cm(2)), a pathological condition that simulates in vivo conditions such as those in stenotic arteries. Platelet microparticle (PMP) formation was analyzed by flow cytometry. Plasma von Willebrand factor (vWF) was also measured. RESULTS: SIPA under high shear stress was significantly enhanced from 44.0 +/- 13.4% preoperatively to 69.5 +/- 15.8% on postoperative day (POD) 1, and it returned to preoperative levels on POD 14. PMP formation under high shear stress was enhanced before surgery (140.8 +/- 38.7%) compared to that under a static condition, and the enhancement was further augmented on POD 1 (219.1 +/- 49.3%). The enhancement of SIPA and PMP formation had no association with disease stage. vWF levels increased significantly on POD 1. Exogenous vWF augmented SIPA and PMP formation under high shear stress, and this augmentation was inhibited by anti-vWF antibody. CONCLUSIONS: Because PMPs are highly procoagulant, increased SIPA and PMP formation induced by surgical intervention possibly contribute to thrombotic complications. Blockage of platelet interaction with vWF may prevent arterial thrombus formation perioperatively.     

Activated neutrophils and platelet microaggregates impede blood filterability through microchannels during simulated extracorporeal circulation

BACKGROUND: Neutrophil sequestration and platelet microaggregates in organ capillaries have been implicated in the inflammatory response associated with cardiopulmonary bypass. We examined the filterability of neutrophils and platelet microaggregates through silicon microchannels during simulated extracorporeal circulation. We hypothesize that blood contact with artificial surfaces over time decreases the ability of neutrophils, platelets, and their aggregates to pass through microchannels. METHODS: Fresh human blood from donors (n = 9) was recirculated for 120 minutes in a simulated extracorporeal circuit. Blood samples were obtained from a donor at 0, 30, 60, and 120 minutes of recirculation. The microchannel transit time and the flow behavior of blood cells were evaluated by a silicon microchannel array flow analyzer. CD11b, L-selectin, and F-actin of neutrophils were measured by flow cytometry. Neutrophil and platelet counts and platelet aggregation to adenosine diphosphate were measured. RESULTS: The microchannel transit time was prolonged during recirculation, reaching 185.9% +/- 25.6% of baseline at 120 minutes. The video microscope showed that neutrophils and platelet microaggregates plugged the microchannels. CD11b, L-selectin, and F-actin levels changed significantly by 120 minutes. Platelet counts decreased and platelet aggregability was attenuated. CONCLUSIONS: Simulated extracorporeal circulation caused a progressive loss in the ability of neutrophils, platelets, and their aggregates to pass through the microchannels independent of neutrophil adhesion molecule expression.     

Platelet protection by aprotinin in cardiopulmonary bypass: electron microscopic study.

To evaluate the functional integrity of platelets in patients administered the proteinase inhibitor aprotinin during cardiopulmonary bypass, 20 patients undergoing a complicated and prolonged open heart operation were studied. They were randomized to receive either a high dose of aprotinin (total dose, 6 to 7 x 10(6) KIU) before and during cardiopulmonary bypass (10 patients) or a placebo (10 patients). Blood samples were collected preoperatively, at the termination of bypass, and 90 minutes thereafter to assess platelet count and aggregation on extracellular matrix, which was studied by scanning electron microscopy. On a scale of 1 to 4, mean preoperative platelet aggregation grades were similar in both groups (3.5 +/- 0.5). Postoperatively, at the termination of cardiopulmonary bypass and 90 minutes thereafter, all 10 patients treated with aprotinin revealed normal, unchanged platelet aggregation (grade, 3.5 +/- 0.5), whereas all placebo-treated patients showed severely disturbed aggregation (grade, 1.4 +/- 0.5) (p less than 0.001). The platelet count was similar in both groups before and after operation (preoperatively, 182 +/- 75 x 10(9)/L and 146 +/- 30 x 10(9)/L, and postoperatively, 87 +/- 13 x 10(9)/L and 80 +/- 27 x 10(9)/L for the aprotinin and placebo groups, respectively). Total 24-hour postoperative bleeding and blood requirement were significantly lower in the aprotinin group (371 +/- 84 mL and 2 +/- 0.7 units, respectively) compared with the placebo group (608 +/- 28 mL and 3.4 +/- 1.3 units, respectively) (p less than 0.01). These results demonstrate that improved postoperative hemostasis is directly related to the complete preservation of platelet function achieved by the protective properties of aprotinin.     

The hemodynamics of thrombus formation in arteries

Alterations in arterial blood flow are thought to predispose to thrombus formation, but the exact relationships have not been fully elucidated. The effect of varying blood flows on the accumulation of thrombotic material within arteries was investigated, with use of shear rate as an index of flow across the luminal surface. Partially denuded rabbit aortas were perfused with fresh nonanticoagulated human blood for 3 minutes, with an in vitro recirculating apparatus, Indium 111-labeled platelets, and fibrinogen I 125. Shear rates ranged from zero to 1500 sec-1, correlating with the hemodynamics of various segments of the human arterial tree. A significant correlation was observed between shear rate and platelet deposition, ranging from 5.2 +/- 2.8 x 10(6) platelets/cm2 of vessel surface area at zero shear to a maximum of 64.7 +/- 8.3 x 10(6) platelets/cm2 at a shear rate of 1500 sec-1 (F = 5.01, p less than 0.05). Fibrin deposition paralleled that of platelets, ranging from 28.2 +/- 7.6 micrograms/cm2 at zero shear to 354.1 +/- 62.7 micrograms/cm2 at a shear rate of 1500 sec-1 (F = 5.91, p less than 0.05). These results suggest that shear rate is a most important determinant of platelet and fibrin deposition on altered arterial surfaces.     

Efficacy of FK633, an ultra-short acting glycoprotein IIb/IIIa antagonist on platelet preservation during and after cardiopulmonary bypass.

OBJECTIVE: Temporary pharmacologic inhibition of platelet function during and after cardiopulmonary bypass (CPB) (platelet anesthesia) is an attractive strategy for preserving platelets during CPB. We examined the efficacy of FK633, an ultra-short acting glycoprotein IIb/IIIa antagonist. METHODS: The study was carried out in six mongrel dogs that received an intravenous bolus of 0.1 mg/kg of FK633 at the time of administration of heparin (group F), and six control dogs (group C). All animals underwent 60 min of normothermic CPB followed by a 2-h observation period. Blood samples for platelet count, platelet aggregation to adenosine diphosphate and parameters concerning the coagulation system were obtained at eight time points. Hemodynamics, bleeding time, and postoperative blood loss were assessed serially. Scanning electron micrograph of the oxygenator's membrane was investigated. RESULTS: FK633 significantly protected platelet number (group F, 59+/-10% versus group C, 38+/-15% of the pre-CPB value; P < 0.01), and inhibited platelet aggregation to adenosine diphosphate (group F, 13+/-12% versus group C, 35+/-9% of the pre-CPB value; P < 0.01) during CPB. Postoperative blood loss did not significantly differ between the two groups, but there was a tendency of less bleeding in group F (group F, 73+/-23 ml versus group C, 111+/-44 ml; P = 0.09). In group F, scanning electron micrograph of the oxygenator's membrane showed that its surface was free from platelets. There were no significant differences between the groups in hemodynamics. CONCLUSIONS: An ultra-short acting glycoprotein IIb/IIIa antagonist, FK633, is effective in preventing both platelet aggregation and thrombocytopenia during CPB, and may be effective for minimizing postoperative bleeding.     

Platelet aggregation in traumatic spinal cord injury.

Study design: Collagen-induced platelet aggregation and platelet count of ten paraplegic patients (four females, six males, aged 16 - 42 years) with traumatic spinal cord injury (SCI) (posttraumatic 12 - 48 weeks) and of ten age-matched healthy volunteers (control group; five females, five males, aged 18 - 37 years) were investigated. Objectives: Investigation of platelet aggregation in the whole blood of the patients with SCI. Setting: Ankara/Turkey. Methods: Platelet aggregation was evaluated by impedance technique using Chrono Log Model 560 WB aggregometer in whole blood. Platelet count was determined by Medonic Cell Analyser 610. Results: Maximal intensity of collagen-induced platelet aggregation of the patients was 18.50+/-8.28 ohm (mean+/-SD) and of the controls was 7.60+/-4.25 ohm. Maximal rate of collagen-induced aggregation of platelets from the patients was 3.98+/-1.59 ohm/min, maximal rate of aggregation of platelets from the controls was 1.57+/-1.01 ohm/min. Platelet counts of the patients and controls were 290 500+/-50 357/mm3 and 273 000+/-48 343/mm3 respectively. It was determined that both maximal rate (P<0.001) and maximal intensity (P<0.01) of collagen-induced platelet aggregation of the patients were significantly higher than those of the controls. There was no significant difference between the two groups in respect to platelet counts. Conclusion: Collagen-induced platelet aggregation of patients with traumatic SCI 12 - 48 weeks after the trauma was significantly higher than that of the controls. Our results indicate that increased tendency of platelet aggregation, which is probably induced by free radicals, may have a great impact on the late thromboembolic complications reported in patients with traumatic SCI.     

Effects of class I heparin binding growth factor and fibronectin on platelet adhesion and aggregation.

Fibronectin and heparin binding growth factor-type 1 have been affixed to vascular graft surfaces to enhance the attachment and the proliferation of transplanted endothelial cells, respectively. The current study examines the effect of fibronectin and heparin binding growth factor-type 1 on platelet adhesion and activation in vivo and on platelet aggregation in vitro. Expanded polytetrafluoroethylene prostheses (5 cm x 4 mm internal diameter) were treated either with fibronectin (n = 9), fibronectin/heparin/heparin binding growth factor-type 1/heparin (n = 12), or neither (n = 13) and were interposed into canine aortoiliac systems bilaterally. Autogenous radiolabeled (Indium 111 oxine, 650 microCi) platelets were injected intravenously before reestablishment of circulation. Perfusion was maintained for 30 minutes, and prostheses were removed with segments of native aorta and distal iliac arteries bilaterally. Specimens were examined for thrombus-free surface area, by gamma well counting for adherent radiolabeled platelets, and by light microscopy and transmission and scanning electron microscopic techniques. Results showed that both the fibronectin and fibronectin/heparin/heparin binding growth factor-type 1/heparin pretreated prostheses contained significantly greater numbers of platelets and adherent radioactivity than did control graft segments when normalized to their ipsilateral iliac arteries. Fibronectin/heparin/heparin binding growth factor-type 1/heparin pretreated prostheses contained 27 +/- 16 times more radioactivity per square millimeter than ipsilateral iliac arteries, fibronectin pretreated prostheses had 13 +/- 8 times more radioactivity per square millimeter than ipsilateral iliac arteries, and untreated expanded polytetrafluoroethylene had 4 +/- 3 times more radioactivity per square millimeter than ipsilateral iliac arteries. Fibronectin/heparin/heparin binding growth factor-type 1/heparin was more radioactive than fibronectin alone (p = 0.056). Histologic evaluation and thrombus-free surface area determinations corroborated the gamma well counting data. Platelet aggregation in vitro was not activated by either fibronectin (1 to 100 micrograms/100 microliters) or heparin binding growth factor-type 1 (25 to 2500 ng/100 microliters). These data suggest that fibronectin and heparin binding growth factor-type 1 promote platelet adhesion not aggregation.     

Anti-platelet effect of aspirin is substantially reduced after administration of heparin during carotid endarterectomy

OBJECTIVES: Aspirin therapy is usually continued throughout the perioperative period to reduce the risk for thromboembolic stroke and myocardial infarction after carotid endarterectomy (CEA). Aspirin irreversibly binds cyclooxygenase-1, thereby reducing platelet aggregation for the lifetime of each platelet. However, recent research from this unit has shown that aggregation in response to arachidonic acid increases significantly, but transiently, during CEA, which suggests that the anti-platelet effect of aspirin is temporarily reversed. The purpose of the current study was to determine when this phenomenon occurs and to identify the possible mechanisms involved. METHODS: Platelet aggregation was measured in platelet-rich plasma from 41 patients undergoing CEA who were stabilized with 150 mg of aspirin daily. Blood was taken at 8 time points: before anesthesia, after anesthesia, before heparinization, 3 minutes after heparinization, 3 minutes after shunt insertion, 10 minutes after flow restoration, 4 hours postoperatively, and 24 hours postoperatively. Platelet aggregation was also measured at similar times in a group of 18 patients undergoing peripheral angioplasty without general anesthesia. RESULTS: All patient platelets were effectively inhibited by aspirin at the start of the operation. There was a significant intraoperative increase in platelet response to arachidonic acid in both groups of patients, which occurred within 3 minutes of administration of unfractionated heparin. In the CEA group this resulted in a greater than 10-fold increase in mean aggregation, to 5 mmol/L of arachidonic acid (5 mmol/L), rising from 3.9% +/- 2.2% preoperatively to 45.1% +/- 29.3% after administration of heparin ( P <.0001). This increased aggregation persisted into the early postoperative period, but by 24 hours post operation aggregation had returned to near preoperative values. Aggregation in response to other platelet agonists (adenosine diphosphate, thrombin receptor agonist peptide) showed only a small increase at the same time, which could be accounted for by a parallel increase in the level of spontaneous aggregation. CONCLUSION: Administration of heparin significantly increases platelet aggregation in response to arachidonic acid, despite adequate inhibition by aspirin administered preoperatively. This apparent reversal in anti-platelet activity persisted into the immediate early postoperative period, and could explain why a small proportion of patients are at increased risk for acute cardiovascular events after major vascular surgery, despite aspirin therapy.     

Effects of the sulphonylurea drugs gliclazide and glibenclamide on blood glucose control and platelet function.

Platelet aggregation and adhesion are commonly increased in diabetes mellitus. These abnormalities may in part be responsible for the increased incidence of vascular disease in diabetics. We have investigated the effects of diet, diet plus glibenclamide, and diet plus gliclazide on plasma glucose control and platelet function in 10 newly diagnosed maturity-onset diabetics who had not previously been treated. Before treatment, the mean postprandial plasma glucose value was 13,4 +/- 0,8 mmol/l, which fell insignificantly on dietary treatment, to 12,2 +/- 1,0 mmol/l (P greater than 0,05). Both glibenclamide and gliclazide, when added to the diet, significantly lowered mean plasma glucose values to 9,3 +/- 0,8 mmol/l and 7,8 +/- 0,8 mmol/l respectively (P less than 0,05). Platelet aggregation in response to 1 mumol adenosine diphosphate (ADP) was increased in the diet period, whereas aggregation in response to 10 mumol and 100 mumol was normal. This suggests an increased sensitivity of the platelets to ADP in diabetic patients. The addition of both glibenclamide and gliclazide reduced the magnitude of the response to within the normal range. Platelet aggregation in response to 10 mumol adrenaline and 750 micrograms/ml collagen was significantly reduced by glibenclamide (P less than 0,05). We conclude that sulphonylurea therapy appears to reduce the increased platelet aggregation which occurs in diabetics. This may play a role in the prevention of vascular disease.