Prothrombin complex concentrate for emergency surgery in patients on oral Xa-inhibitors.

BACKGROUND: It is uncertain whether prothrombin complex concentrate (PCC) improves hemostasis in patients on treatment with oral factor Xa-inhibitors (XaI) who require emergency surgery. OBJECTIVES: To evaluate whether, in patients with therapeutic levels of oral XaI, preoperative PCC prevents excessive bleeding during and after emergency surgery and is not associated with thrombotic complications. METHODS: We conducted a prospective cohort study wherein a fixed 2000 IU dose of 4-factor PCC was given to patients taking oral XaI with plasma XaI levels of at least 75 ng/mL before the emergency surgery with an expected blood loss of at least 50 mL. Patients were followed for 30 days. The primary efficacy outcome was the incidence of normal or mildly abnormal surgical hemostasis, as assessed by the surgeon; primary safety outcome was the incidence of thromboembolic events within 7 days. RESULTS: We included 20 patients, 50% were female, on apixaban (75%) or rivaroxaban (25%) with median XaI level of 128 ng/mL (range 77-497). The median duration of surgery was 2h 42 min (15 min to 8h 17 min). Normal or mildly abnormal hemostasis was observed in 16 patients (80%), 2 patients had moderately abnormal and 2 had severely abnormal hemostasis, 1 each of those was considered due to local or technical factors. There were 4 deaths (20%) secondary to underlying disease and 1 incidental pulmonary embolism in a patient with cancer. CONCLUSION: A fixed dose PCC appears to control hemostasis in patients with therapeutic plasma levels of apixaban or rivaroxaban requiring emergency surgery.

Extended thromboprophylaxis following colorectal surgery in patients with inflammatory bowel disease: a comprehensive systematic clinical review.

AIM: Patients with inflammatory bowel disease (IBD) are at increased risk of postoperative venous thromboembolism (VTE) following major abdominal surgery. The pathogenesis is multifactorial and not fully understood. A combination of pathophysiology, patient and surgical risk factors increase the risk of postoperative VTE in these patients. Despite being at increased risk, IBD patients are not regularly prescribed extended pharmacological thromboprophylaxis following colorectal surgery. Currently, there is a paucity of evidence-based guidelines. Thus, the aim of this review is to evaluate the role of extended pharmacological thromboprophylaxis in IBD patients undergoing colorectal surgery. METHOD: A search of Ovid Medline, EMBASE and PubMed databases was performed. A qualitative analysis was performed using 10 clinical questions developed by colorectal surgeons and a thrombosis haematologist. The Newcastle-Ottawa Scale was utilized to assess the quality of evidence. RESULTS: A total of 1229 studies were identified, 38 of which met the final inclusion criteria (37 retrospective, one case-control). Rates of postoperative VTE ranged between 0.6% and 8.9%. Patient-specific risk factors for postoperative VTE included ulcerative colitis, increased age and obesity. Surgery-specific risk factors for postoperative VTE included open surgery, emergent surgery and ileostomy creation. Patients with IBD were more frequently at increased risk in the included studies for postoperative VTE than patients with colorectal cancer. The risk of bias assessment demonstrated low risk of bias in patient selection and comparability, with variable risk of bias in reported outcomes. CONCLUSION: There is a lack of evidence regarding the use of extended pharmacological thromboprophylaxis in patients with IBD following colorectal surgery. As these patients are at heightened risk of postoperative VTE, future study and consideration of the use of extended pharmacological thromboprophylaxis is warranted.

Thrombus stability explains the factor V Leiden paradox: a mouse model.

Humans carrying the factor V Leiden (FVL) variant have a fivefold increased risk for venous thrombosis. However, incidence of deep vein thrombosis (DVT) is proportionally greater than that of pulmonary embolism (PE) in these individuals. This is known as the FVL paradox. We hypothesized that the rate of initial DVT development is similar in FVL and noncarriers, but thrombi in FVL carriers are more stable and develop into a clinically significant DVT more often than in noncarriers. To test this, we induced thrombi in the femoral vein of wild-type (WT), heterozygous (F5L/+), and FVL homozygous (F5L/L) mice. Using intravital microscopy, thrombus size and embolization were visualized and emboli in the lungs were quantified. Compared with WT, femoral vein thrombi in F5L/+ and F5L/L mice were larger and embolized less. Total and large embolic events, the percentage of thrombus that embolized, and PE burden were significantly decreased in F5L/L mice. This suggests that in noncarriers (reflected by WT), a minor injury initially resulting in a small DVT tends to remain small and asymptomatic because of the embolization of the otherwise growing thrombus. Alternatively, the same insult in people with FVL (reflected by F5L/L) leads to thrombus growth as a result of less embolization, and thus symptomatic DVT development.

Reversal of dabigatran-associated major bleeding with activated prothrombin concentrate: A prospective cohort study.

The reversal of dabigatran-associated major bleeding can now be achieved with the antidote idarucizumab. We evaluated activated prothrombin complex concentrate (aPCC) as an alternative for this purpose. Patients treated with dabigatran and suffering a major bleed were treated as per existing hospital protocol with aPCC. They were subsequently recruited for a 30-day follow-up. Effectiveness was evaluated by the treating physician, using an Assessment Guide. Safety outcomes were arterial or venous thromboembolism or death. A comparison was also made with historic cases with dabigatran-associated major bleeds treated with supportive care, by matching 1:2 for type of bleed, age and sex. We aimed at 32 evaluable cases but the study was prematurely discontinued after 14 cases due to the availability of the approved antidote. The effectiveness of aPCC was assessed as Good in 9 (64%), moderate in 5 (36%) and poor in none. There were no thromboembolic events and one death. In the secondary adjudication of effectiveness, using the same criteria and by the same adjudicators as previously done for the historic cases, the outcome was graded for the current cases versus the historic cases as Good, Moderate, or Poor in 10 (71%) versus 16 (57%), 3 (21%) versus 4 (14%), and 1 (7%) versus 8 (29%), respectively. Although supportive care is sufficient to manage many patients with dabigatran-associated bleeding, aPCC might provide an additional benefit to control life-threatening bleeding in selected cases and does not appear to cause an excess of thromboembolic events.

Comparison of the effect of dabigatran and dalteparin on thrombus stability in a murine model of venous thromboembolism.

UNLABELLED: ESSENTIALS: Does thrombus stability alter the presentation of venous thromboembolism and do anticoagulants alter this? In a murine model, we imaged a femoral vein thrombus and quantified emboli in the pulmonary arteries. Dabigatran decreases thrombus stability via factor XIII increasing embolization and pulmonary emboli. This cautions against the unapproved use of dabigatran for acute initial treatment of deep vein thrombosis. BACKGROUND: Venous thromboembolism (VTE) is a collective term for deep vein thrombosis (DVT) and pulmonary embolism (PE). Thrombus instability possibly contributes to progression of DVT to PE, and direct thrombin inhibitors (DTIs) may alter this. AIM: To develop a model to assess thrombus stability and its link to PE burden, and identify whether DTIs, in contrast to low-molecular-weight heparin (LMWH), alter this correlation. METHODS: Twelve minutes after ferric chloride-induced thrombus formation in the femoral vein of female mice, saline, dalteparin (LMWH) or dabigatran (DTI) was administered. Thrombus size and embolic events breaking off from the thrombus were quantified before treatment and at 10-min intervals after treatment for 2 h using intravital videomicroscopy. Lungs were stained for the presence of PE. RESULTS: Thrombus size was similar over time and between treatment groups. Total and large embolic events and pulmonary emboli were highest after treatment with dabigatran. Variations in amounts of pulmonary embolic events were not attributed to variations in thrombus size. Large embolic events correlated with the number of emboli per lung slice independent of treatment. Embolization in factor XIII deficient (FXIII(-/-) ) saline-treated mice was greater than that in wild-type (WT) saline-treated mice, but was similar to WT dabigatran-treated mice. CONCLUSION: We have developed a mouse model of VTE that can quantify emboli and correlate this with PE burden. Consistent with clinical data, dabigatran, a DTI, acutely decreases thrombus stability and increases PE burden compared with LMWH or saline, which is a FXIII-dependent effect.

Rapid quantitative D-dimer to exclude pulmonary embolism: a prospective cohort management study.

UNLABELLED: ESSENTIALS: It is not known if D-dimer testing alone can safely exclude pulmonary embolism (PE). We studied the safety of using a quantitative latex agglutination D-dimer to exclude PE in 808 patients. 52% of patients with suspected PE had a negative D-dimer test and were followed for 3 months. The negative predictive value of D-dimer testing alone was 99.8%, suggesting it may safely exclude PE. BACKGROUND: Strategies are needed to exclude pulmonary embolism (PE) efficiently without the need for imaging tests. Although validated rules for clinical probability assessment can be combined with D-dimer testing to safely exclude PE, the rules can be complicated or partially subjective, which limits their use. OBJECTIVES: To determine if PE can be safely excluded in patients with a negative D-dimer without incorporating clinical probability assessment. PATIENTS/METHODS: We enrolled consecutive outpatients and inpatients with suspected PE from four tertiary care hospitals. All patients underwent D-dimer testing using the MDA D-dimer test, a quantitative latex agglutination assay. PE was excluded in patients with a D-dimer less than 750 µg FEU L(-1) without further testing. PATIENTS: with D-dimer levels of 750 µg FEU L(-1) or higher underwent standardized imaging tests for PE. All patients in whom PE was excluded had anticoagulant therapy withheld and were followed for 3 months for venous thromboembolism (VTE). Suspected events during follow-up were adjudicated centrally. RESULTS: Eight hundred and eight patients were enrolled, of whom 99 (12%) were diagnosed with VTE at presentation. Four hundred and twenty (52%) patients had a negative D-dimer level at presentation and were not treated with anticoagulants; of these, one had VTE during follow-up. The negative predictive value of D-dimer testing for PE was 99.8% (95% confidence interval, 98.7-99.9%). CONCLUSIONS: A negative latex agglutination D-dimer assay is seen in about one-half of patients with suspected PE and reliably excludes PE as a stand-alone test.

Clinical challenges in patients with cancer-associated thrombosis: Canadian expert consensus recommendations.

Venous thromboembolism is a common complication in cancer patients, and thromboembolism is the second most common cause of death after cancer progression. A number of clinical practice guidelines provide recommendations for the management of cancer-associated thrombosis. However, the guidelines lack recommendations covering commonly encountered clinical challenges (for example, thrombocytopenia, recurrent venous thromboembolism, etc.) for which little or no evidence exists. Accordingly, recommendations were developed to provide expert guidance to medical oncologists and other health care professionals caring for patients with cancer-associated thrombosis. The current expert consensus was developed by a team of 21 clinical experts. For each identified clinical challenge, the literature in medline, embase, and Evidence Based Medicine Reviews was systematically reviewed. The quality of the evidence was assessed, summarized, and graded. Consensus statements were generated, and the experts voted anonymously using a modified Delphi process on their level of agreement with the various statements. Statements were progressively revised through separate voting iterations and were then finalized. Clinicians using these recommendations and suggestions should tailor patient management according to the risks and benefits of the treatment options, patient values and preferences, and local cost and resource allocations.

Comparison of the effect of coagulation and platelet function impairments on various mouse bleeding models.

Haemostatic impairments are studied in vivo using one of several murine bleeding models. However it is not known whether these models are equally appropriate for assessing coagulation or platelet function defects. It was our study objective to assess the performance of arterial, venous and combined arterial and venous murine bleeding models towards impaired coagulation or platelet function. Unfractionated heparin (UFH) or αIIbß3inhibitory antibody (Leo.H4) were administered to mice, and their effects on bleeding in saphenous vein, artery, and tail tip transection models were quantified and correlated with their effects on plasma clotting and ADP-induced platelet aggregation, respectively. All models exhibited similar sensitivity with UFH (EC50 dose = 0.19, 0.13 and 0.07 U/g, respectively) (95% CI = 0.14 - 0.27, 0.08 - 0.20, and 0.03 - 0.16 U/g, respectively). Maximal inhibition of ex vivo plasma clotting could be achieved with UFH doses as low as 0.03 U/g. In contrast, the saphenous vein bleeding model was less sensitive to αIIbß3 inhibition (EC50 = 6.9 µg/ml) than tail transection or saphenous artery bleeding models (EC50 = 0.12 and 0.37 µg/ml, respectively) (95% CI = 2.4 - 20, 0.05 - 0.33, and 0.06 - 2.2 µg/ml, respectively). The EC50 of Leo.H4 for ADP-induced platelet aggregation in vitro (8.0 µg/ml) was at least 20-fold higher than that of the tail and arterial, but not the venous bleeding model. In conclusion, venous, arterial and tail bleeding models are similarly affected by impaired coagulation, while platelet function defects have a greater influence in models incorporating arterial injury.

Diannexin, an annexin A5 homodimer, binds phosphatidylserine with high affinity and is a potent inhibitor of platelet-mediated events during thrombus formation.

BACKGROUND: Shielding of procoagulant phosphatidylserine (PS) with annexin A5 attenuates thrombosis, but annexin A5 (35.7 kDa) is rapidly cleared from the circulation. In contrast, Diannexin, a 73.1 kDa homodimer of annexin A5, has an extended half-life. OBJECTIVES: To quantify the affinity of Diannexin for PS, examine its interaction with activated platelets and determine its effects on platelet-mediated events during thrombus formation. METHODS: The affinities of Diannexin and annexin A5 for PS-containing lipid bilayers were compared using surface plasmon resonance, and binding to activated platelets was assessed by flow cytometry. Calibrated automated thrombography and thromboelastography were employed to study the effects of Diannexin on thrombin generation and platelet-fibrin clot formation, respectively, whereas intravital videomicroscopy was used to examine its effect on platelet accumulation and activation after laser-induced injury to murine cremaster arterioles, and a tail tip bleeding model was used to explore its effects on hemostasis. RESULTS: Diannexin and annexin A5 bind PS with K(D) values of 0.6 and 5 nm, respectively, and both bind to the same subpopulation of PS-exposing platelets. Diannexin inhibited thrombin generation and platelet-fibrin clot formation in vitro at 10 nm (P<0.05-0.001 compared with control), and reduced platelet accumulation at 1 µg g(-1) (P<0.05) and activation at 0.25 µg g(-1) (P<0.001) in experimentally induced arterial thrombi in mice while increasing blood loss at 1 µg g(-1) (P<0.01). CONCLUSIONS: Diannexin binds to PS with high affinity and is a potent inhibitor of platelet-mediated events during thrombus formation.

Abnormal hemostasis in a knock-in mouse carrying a variant of factor IX with impaired binding to collagen type IV.

BACKGROUND: Factor IX binds to collagen type IV, but this binding has no known consequence. OBJECTIVES: To determine the effect of reduced binding of FIX to collagen IV. METHODS: We constructed and characterized 'knock-in' mice containing the mutation lysine 5 to alanine (K5A) in the Gla domain of their FIX. The K5A mutation dramatically reduced the affinity of FIX for collagen type IV, but had no measurable effect on platelet binding, phospholipid binding, or in vitro clotting activity. However, K5AFIX mice had a mild bleeding tendency, despite their in vitro clotting activity being normal. Hemostatic protection from delayed rebleeding was intermediate between wild-type and hemophilia B mice (which had no detectable clotting activity); moreover, survival of K5A FIX mice after nascent clot removal was dramatically improved as compared with hemophilia B mice. Importantly, there was no detectable difference between K5AFIX and wild-type mice in either a laser-induced thrombosis model or the chromogenic FIX activity assay. In contrast, after ferric chloride injury, which exposes collagen IV as well as other basement membrane proteins, intravital microscopy revealed that vessel occlusion was significantly slower in K5AFIX mice than in wild-type mice. CONCLUSIONS: Our results indicate that the FIX molecule with decreased affinity for collagen IV has altered hemostatic properties in vivo and that the binding of FIX to collagen IV probably plays a significant functional role in hemostasis.

New antithrombotic drugs.

Thrombosis, both venous and arterial, is a major cause of morbidity and mortality worldwide. Consequently, there is an ongoing search for new antithrombotic drugs, particularly novel antiplatelet agents and anticoagulants. A better understanding of the biochemical pathways involved in platelet activation and coagulation and of the links between these systems and the impact of thrombosis on inflammation has led to the identification of new targets for antithrombotic drugs. This paper focuses on these new targets and new antiplatelet drugs and anticoagulants and describes the major advances in the continuing search for more potent antithrombotic drugs that have limited effects on hemostasis.

Characterization of the Reaction of North Dakota Dry Bean Cultivars to Three Races of Colletotrichum lindemuthianum.

Colletotrichum lindemuthianum, causal agent of anthracnose, a serious disease of dry bean, had been prevalent mostly in the eastern United States and Michigan. In 2001, the disease was observed affecting several commercial bean fields in North Dakota. To assess the potential impact of this pathogen on the North Dakota dry bean industry, 30 of the most widely planted cultivars were inoculated with races 7, 73, and 89 of C. lindemuthianum. Race 73 is present in Michigan, and has been detected in recently North Dakota and Manitoba, while races 7 and 89 are currently present in Michigan and Ontario. All cultivars were spray inoculated with a suspension of 106 conidia/ml in the greenhouse and evaluated for disease reaction using a 0-to-9 scale 8 days after inoculation. Most kidney beans were susceptible to race 7, but resistant to races 73 and 89. 'Isles' and 'Drake' were the only kidney beans resistant to all three races. All pinto beans were susceptible to races 73 and 89 except 'Topaz', which was moderately resistant to race 73 but susceptible to race 89. Most pinto beans were resistant or moderately resistant to race 7. Navy bean cvs. Newport and Envoy were resistant to all three races; however, 'Norstar', the most widely planted navy bean cultivar, was susceptible to all three races. Research is being conducted to identify germplasm with resistance to these races. To reduce the possibility of spreading this pathogen to new fields in the region, the use of noncertified seed should be discouraged among producers, and fields used for seed production should be monitored very carefully.

First Report of Dry Bean Anthracnose (Colletotrichum lindemuthianum) Race 73 in North Dakota.

Dry bean (Phaseolus vulgaris L. cv. Pintoba) plants showing typical anthracnose symptoms were observed in three commercial fields in North Dakota (Towner, Steele, and Pembina counties) in July 2001. Disease incidence in all fields ranged from 5 to 20%. The fungus was isolated from leaves and pods on potato dextrose agar and identified as Colletotrichum lindemuthianum (Sacc. & Magnus) Lams.-Scrib. (3). Pathogenicity and race identification were determined on a set of 12 standard differentials (2). Three isolates, one from each county, were grown for 7 days in Mathur's medium. Spores were suspended in water and Tween 80 (0.1% vol/vol) and adjusted to 106 spores per ml. Thirty 2-week-old seedlings of each differential were inoculated with each isolate on the adaxial side of the primary leaves using a Paasche airbrush. Inoculated plants were incubated in moist chambers for 5 days at 20°C under 14 h of fluorescent light and then moved back to the greenhouse. Disease reaction was assessed 3 days later. Isolates of C. lindemuthianum races 7 and 73 obtained from J. Kelly (Michigan State University) were used as positive controls. Inoculations were repeated once. All three North Dakota isolates and the positive control for race 73 produced sporulating lesions on the differentials 'Michelite', 'Cornell 49242', and 'Mexico 222'. No lesions were observed in the other differentials. An unidentified anthracnose race retrieved from a single plant in 1982 constitutes the first report of the presence of anthracnose in North Dakota (4). In 1992, Michigan breeding materials infected with race 73 were planted in North Dakota (1); upon detection, the infected plants were destroyed and the fields quarantined. The epidemics observed in the 2001 season, developed in sites distant from the places where the Michigan materials were planted and have been associated with a single seed source. To our knowledge, the presence of anthracnose race 73 reported here constitutes the first report of anthracnose in commercial dry bean fields in North Dakota. References: (1) J. D. Kelly et al. Plant Dis. 78:892, 1994. (2) M. A. Pastor-Corrales. Phytopathology 81:694, 1991. (3) B. C. Sutton, The Coelomycetes, CAB International, Wallingford, Oxon, UK, 1980. (4) J. R. Venette and P. A. Donald. Bean Improv. Coop. 26:24, 1983.

Overwinter Survival of Bean Rust Urediniospores in North Dakota.

Bean rust (Uromyces appendiculatus) reached epidemic proportions in North Dakota and Minnesota from 1993 to 1996. Although U. appendiculatus is a macrocyclic autoecious rust, neither pycnia nor aecia have been observed in commercial dry bean (Phaseolus vulgaris) in North Dakota fields. The source of initial inoculum is not clearly understood. This study determined the potential for urediniospore survival overwinter. Uredia-bearing bean leaves from artificially inoculated greenhouse-grown plants were kept outside near a field from November to May from 1990 to 1996. Based on bioassays urediniospores survived overwinter, but viability declined over time. Overwinter survival indicates urediniospores may function as initial inoculum.

The endothelium and thrombosis.

Thrombosis reflects an imbalance between procoagulant and anticoagulant mechanisms. In some cases, thrombotic lesions are precipitated by gross changes in blood flow, vascular wall integrity, or systemic levels of coagulation factors. In other cases, thrombosis is induced by functional changes within the endothelium. Endothelial cells express a wide variety of factors that contribute to hemostasis, including procoagulants, anticoagulants, cell adhesion molecules, vasomotor substances, and cell survival signals. Because the endothelium displays a remarkable diversity of structure and function, the relative contribution of any one of these factors to the hemostatic balance varies between different vascular beds. In this review, we emphasize the heterogeneous nature of endothelial cell function. We then examine the role of endothelial diversity in modulating the phenotypic expression of thrombotic disorders.

Enhanced collagen-induced responses of platelets from rabbits with diet-induced hypercholesterolemia are due to increased sensitivity to TxA2. Response inhibition by chronic ethanol administration in hypercholesterolemia is due to reduced TxA2 formation.

The effects of dietary cholesterol and chronic administration of moderate amounts of ethanol on collagen-induced platelet responses were investigated. Three groups of rabbits were fed the following diets for 8 weeks: a normal chow diet, a cholesterol-enriched (0.25% wt/wt) chow diet, and a cholesterol-enriched chow diet plus 6% ethanol in the drinking water for the final week of the dietary period. Cholesterol feeding enhanced collagen-induced responses-aggregation, secretion of [14C]serotonin from prelabeled platelets, and thromboxane formation--of suspensions of washed platelets, and chronic ethanol treatment significantly reduced these enhanced responses. These effects are mediated by thromboxane A2 (TxA2) rather than ADP. Experiments with collagen-stimulated platelets in which feedback amplification of TxA2 was blocked with the prostaglandin H2/TxA2 receptor blocker BM 13.177 and experiments with aspirin-treated platelets stimulated with the stable TxA2 mimetic U46619 showed that cholesterol feeding enhanced platelet sensitivity to TxA2 rather than formation of TxA2 by platelets that had interacted with collagen. Without BM 13.177 or aspirin, TxA2 increased the amount of TxA2 formed by feedback amplification. In contrast, decreased responsiveness to collagen by platelets from cholesterol-fed rabbits given ethanol was due to inhibition of TxA2 formation rather than reduced sensitivity to TxA2. Platelets from cholesterol-fed rabbits given ethanol did not develop tolerance to the acute inhibitory effects of ethanol. Our results indicate that administration of moderate amounts of ethanol to cholesterol-fed rabbits inhibits enhanced collagen-induced responses of platelets by a TxA2-dependent pathway that involves reduction of TxA2 formation rather than reduction of platelet responses to TxA2.

Acute in vitro effects of ethanol on responses of platelets from cholesterol-fed and Watanabe heritable hyperlipidemic rabbits.

The effects of ethanol on platelets from rabbits with two different types of hypercholesterolemia, diet-induced and genetically determined, were investigated. There were no differences between the hypercholesterolemic groups and their controls in the extent of (primary) ADP-induced aggregation of washed platelets, and this aggregation was not inhibited by ethanol. Platelets from cholesterol-fed rabbits were more sensitive to aggregation and secretion induced by collagen, whereas platelets from Watanabe heritable hyperlipidemic (WHHL) rabbits were less sensitive. Ethanol inhibited collagen-induced responses of platelets from both hypercholesterolemic groups, but the extent of inhibition of aggregation was not different compared with controls. Because ethanol did not affect U46619-induced responses of aspirin-treated platelets, ethanol does not inhibit aggregation and secretion stimulated by collagen via an effect on thromboxane A2 (TxA2)-induced responses. Platelets from cholesterol-fed rabbits were more sensitive to thrombin even when TxA2 formation was blocked by aspirin, and inhibition of aggregation by ethanol was less in cholesterol-fed rabbits than in controls. However, neither the extent of thrombin-induced responses nor the inhibitory effect of ethanol was different in platelets from WHHL rabbits compared with controls. Thus, different etiologies of hypercholesterolemia produce different changes in platelet function, and ethanol has different effects on the platelets from cholesterol-fed rabbits compared with the platelets from WHHL rabbits. The inhibitory effect of ethanol on the thrombin-induced aggregation of platelets from cholesterol-fed rabbits is attenuated compared with controls, and this finding contrasts with the reported greater inhibitory effect of ethanol on platelets enriched with saturated fats.

Platelet function in Watanabe heritable hyperlipidemic rabbits. Decreased sensitivity to thromboxane A2.

The characteristics of platelets from seven 5-7-month-old homozygous Watanabe heritable hyperlipidemic (WHHL) rabbits (plasma cholesterol, 13.9 +/- 1.7 mM, mean +/- SEM) were compared with those of platelets from normocholesterolemic age/weight- and sex-matched control rabbits (plasma cholesterol, 2.2 +/- 0.3 mM). Whole-blood platelet count and platelet size and protein content were not different in the two groups of rabbits, and the platelets from the WHHL rabbits were not enriched in cholesterol as indicated by identical mean cholesterol:phospholipid molar ratios (C/P). Responses of washed platelets stimulated with various agonists were studied to determine the effects of the genetically determined hypercholesterolemia on the various pathways of platelet aggregation in the absence of plasma components. In platelets from WHHL rabbits compared with controls, aggregation induced by ADP (0.5-5 microM) did not differ; collagen-induced (0.25-1.5 micrograms/ml) responses (aggregation, secretion of carbon-14-labeled serotonin from the amine storage granules of prelabeled platelets, and thromboxane A2 [TxA2] formation) were significantly less extensive; with aspirin-treated platelets, aggregation and secretion of granule contents induced by the TxA2 mimetic U46619 (0.25-1 microM) were significantly less extensive; and thrombin-induced (0.005-0.1 unit/ml) responses of untreated platelets (aggregation, secretion of granule contents, and TxA2 formation) or aspirin-treated platelets (aggregation and secretion of granule contents) did not differ. These observations are in direct contrast with previous studies of platelets from rabbits with diet-induced hypercholesterolemia, in which responses to TxA2 and thrombin were enhanced. Platelets from WHHL rabbits are hyposensitive to aggregation induced by TxA2.

Platelet hypersensitivity in cholesterol-fed rabbits: enhancement of thromboxane A2-dependent and thrombin-induced, thromboxane A2-independent platelet responses.

Hypercholesterolemia (mean plasma cholesterol: 15 mM) was induced in rabbits by the feeding of a chow diet enriched with a low amount (0.25%, w/w) of cholesterol only. Platelet size and protein content decreased significantly, but the whole blood platelet count did not change. The platelets became enriched in cholesterol, as indicated by a significant increase in the cholesterol:phospholipid molar (C/P) ratio. Specific responses of washed platelets stimulated with various agonists were studied to determine the effects of hypercholesterolemia on the various pathways of platelet aggregation in the absence of plasma components. In platelets from hypercholesterolemic rabbits compared with controls: aggregation induced by ADP was not altered; collagen-induced responses (aggregation, secretion of [14C]serotonin from prelabelled platelets, thromboxane A2 (TXA2) formation, mobilization of [3H]arachidonate from prelabelled platelets) were enhanced; with aspirin-treated platelets, aggregation induced by the TXA2 mimetic U46619 was enhanced: and thrombin-induced responses of both untreated platelets (aggregation, secretion of granule contents, TXA2 formation) and aspirin-treated platelets (aggregation) were enhanced. Thus, platelets from cholesterol-fed rabbits not only form more TXA2, but they aggregate more extensively when stimulated by its mimetic. In addition, it has not been previously recognized that these platelets are also hypersensitive to thrombin-induced aggregation that is independent of TXA2.