Potentiation of platelet responses in vitro by feline infectious peritonitis virus

The effect of feline infectious peritonitis virus (FIPV) on platelet aggregation and 14C-serotonin release induced by threshold levels of four agonists (adenosine diphosphate [ADP], collagen, arachidonic acid, and epinephrine) was examined in vitro in ten specific-pathogen-free cats. Purified suspensions of FIPV added to stirred platelet suspensions (virus to platelet ratio equal to 1:320) 1 minute prior to the addition of agonist potentiated the ADP-induced aggregation response by greater than 100% in seven cats. Platelet 14C-serotonin release was increased by greater than 100% in four cats. Collagen-induced platelet aggregation was enhanced in ten cats while collagen-induced 14C-serotonin release was enhanced in eight cats. Potentiation of arachidonic acid-induced platelet aggregation was observed in three cats, two of which demonstrated enhanced platelet 14C-serotonin release. Although epinephrine-induced platelet aggregation was enhanced in five cats, the samples displayed only fine microaggregates. Enhanced 14C-serotonin release from platelets in response to epinephrine was not demonstrated. Interaction with the outer platelet membrane and internalization of viral particles within the surface-connected open canalicular system were demonstrated by electron microscopy within 5 minutes of the addition of virus to platelet suspensions with or without added agonists. Decreasing the virus concentration by ten- or one hundred-fold abolished the potentiating effect observed previously, while increasing the concentration tenfold resulted in direct platelet activation in the absence of agonist.(ABSTRACT TRUNCATED AT 250 WORDS)     

Vortex mixing of feline blood to disaggregate platelet clumps

Abstract: Platelet clumping is a common cause of erroneous platelet counts in cats. Mixing of blood with a vortex mixer was evaluated as a method to disaggregate platelet clumps in feline blood and thus obtain accurate platelet counts. Whole blood samples from 42 cats with platelet clumping and 10 control cats without platelet clumping were mixed for 1 minute at the maximal setting using a standard vortex mixer. Blood smears (for subjective assessment of the type and amount of platelet clumping), platelet counts, and total leukocyte counts were evaluated before and after mixing. Vortex treatment of blood samples with platelet clumps caused an increased platelet count in all but 1 sample. Although most samples had strong increases in platelet counts after mixing, only a minority of samples (5 of 42) appeared to have all platelet clumps dispersed. Of 39 feline blood samples with platelet counts initially <200×10⁹ cells/L, 23 counts increased to >200×10⁹ cells/L and 34 counts increased to >100×10⁹ cells/L. Overall, mixing gave inconsistent and partial improvement in platelet counts. Total leukocyte counts were not significantly affected by vortex mixing. Vortex mixing of 10 feline blood samples without platelet clumping had no consistent effect on platelet or WBC counts. In conclusion, vortex mixing of feline blood does not appear to be a consistent means of correcting the problem of feline platelet clumping.     

Estimation of platelet counts on feline blood smears

Blood samples were obtained from 50 cats admitted for hematologic evaluation at the Royal (Dick) School of Veterinary Studies. Manual platelet counts were done using a hemacytometer, and the average number of platelets per oil immersion field (1,000X magnification) was determined on stained blood smears. A hemacytometer count was not obtained for one sample because of a failure in erythrocyte lysing. In nine samples, obvious platelet clumps in the blood smear prevented accurate determination of the number of platelets per oil immersion field. Hemacytometer counts on these nine samples ranged from 260-587 X 10 (3) platelets/microliter, suggesting that platelet clumps on a blood smear were usually associated with adequate platelet numbers. Simple regression analysis of hemacytometer counts and the average umber of platelets per oil immersion field for the remaining 40 samples yielded correlation coefficients (r) of 0.776 on untransformed data, and 0.892 on log10-transformed data. Each platelet per oil immersion field represented a circulating platelet count of approximately 20 X 10(3)/microliter, similar to conversion factors reported for dogs and human beings. It was concluded that estimation of platelet number on stained blood smears is a simple and quick method that appears to be reliable over a wide range of platelet counts in cats.     

Neutrophil and platelet activation in equine recurrent airway obstruction is associated with increased neutrophil CD13 expression, but not platelet CD41/61 and CD62P or neutrophil–platelet aggregate formation

Recurrent airway obstruction (RAO) in mature horses is characterized by reversible airway obstruction and neutrophilic inflammation; there is also functional activation of circulating platelets and neutrophils. This study was undertaken to determine if changes in activation marker expression and heterotypic aggregate formation can be used as an indicator of this increased functional responsiveness.In vitro conditions for flow cytometric measurement of CD13, CD41/61 and CD62P expression on activated cells and heterotypic aggregate formation were established. Values were then compared before and after antigen challenge of RAO and healthy horses. Platelet adhesion to serum-coated plastic was measured as a functional marker of platelet activation.In vitro activation resulted in increased expression of neutrophil CD13 and platelet CD41/61 and CD62P. Activation of both cell types caused a significant increase in neutrophil–platelet aggregates.In horses with RAO, but not controls, there was a significant increase in the percentage of CD13 positive neutrophils at 10 h and 24 h and in the mean fluorescence intensity at 10 h. This was accompanied at 24 h by an increased mean platelet side scatter and thrombin-stimulated platelet adhesion.In conclusion, CD13 expression can be used as an indicator of equine neutrophil activation both in vitro and in vivo. Equine platelet activation in vitro can be detected by measuring CD41/61 or CD62P expression, and PAF-activated platelets and neutrophils form aggregates. However, despite evidence of circulating platelet activation, neither a change in expression of platelet activation markers, nor heterotypic aggregate aggregate formation could be detected.     

Mean platelet component as an indicator of platelet activation in foals and adult horses

BACKGROUND: Mean platelet component (MPC) is a new platelet variable, measured by modern commercial complete blood count analyzers, that is reduced during platelet activation in humans and small animals. HYPOTHESIS: MPC decreases in horses with clinical conditions that cause platelet activation and disseminated intravascular coagulation (DIC). ANIMALS: We obtained 418 CBCs from 100 sick and 20 healthy neonates and 178 sick and 45 sound adult horses. Sick neonates were classified into septic and nonseptic, and DIC and non-DIC groups. Adults were grouped by diagnoses (systemic inflammatory disorders, gastrointestinal problems, and thrombocytopenia). METHODS: MPC together with platelet count, mean platelet volume, platelet distribution width, and platelet component distribution width were measured with a commercial analyzer and compared between the different disease and control groups in neonates and in adults. RESULTS: MPC values were significantly lower in the septic and nonseptic neonates (24.0 +/- 3.5 g/dL and 26.6 +/- 2.6 g/dL, respectively) than in the control group (28.1 +/- 1.7 g/dL). Neonates with DIC had the lowest MPC values (23.8 +/- 6.3 g/dL). MPC values in adult horses were significantly lower in the inflammatory (23.5 +/- 4.7 g/dL), gastrointestinal obstruction (23.0 +/- 5.0 g/dL), enteritis (23.6 +/- 4.6 g/dL), ischemic (23.9 +/- 5.1 g/dL), and thrombocytopenia (20.2 +/- 5.7 g/dL) groups when compared with control horses (26.2 +/- 3.5 g/dL). Other platelet variables were not different between the control and the disease groups. CONCLUSION AND CLINICAL IMPORTANCE: MPC might be a useful variable for quickly and easily detecting platelet activation in sick neonates and adult horses.     

Enhanced platelet reactivity in cats experimentally infected with feline infectious peritonitis virus

Platelet function was evaluated in six specific-pathogen-free cats prior to and following intraperitoneal inoculation with feline infectious peritonitis virus (FIPV). By 4 days post-inoculation, platelet samples from five of six cats responded with irreversible platelet aggregation to threshold concentrations of adenosine diphosphate (ADP). This was accompanied by enhanced platelet 14C-serotonin release (greater than 10%) in two cats. Compared to one of six baseline samples, five of five post-inoculation samples exhibited microaggregate formation in response to 20 microM epinephrine. Enhanced platelet 14C-serotonin release did not accompany these responses. Enhanced platelet responses to ADP and epinephrine were also observed on day 11 post-inoculation and day 16 (when one cat died) or 21 (the end of the study). Platelet 14C-serotonin release in response to 20 microM epinephrine increased markedly in three of five cats on day 21. Enhanced collagen-induced platelet responses were not demonstrated. Although the mechanism for the enhanced platelet responses observed on day 4 was unknown, a direct effect on the virus on platelets, mononuclear inflammatory cells, and endothelial cells must be considered.     

Effects of aspirin and propranolol on feline platelet aggregation

Three groups of cats were given aspirin in single dosages of 5, 10, and 25 mg/kg. A 4th group was given 2 dosages of 25 mg/kg, 3 days between doses. The degree of inhibition of platelet function and the duration of antagonism to platelet aggregation were studied. The dosage of 25 mg/kg (single and repeated) consistently inhibited platelet function, and the effects lasted 3 to 5 days. Additive drug effect was not noted in group 4, since the 2nd dosage antagonized platelet function to a degree and duration similar to the effects of the 1st. The dosage of aspirin recommended for antiplatelet activity in the average-sized cat (3 kg) equals approximately 1 pediatric aspirin (90 mg) given twice a week. Propranolol, given to cats at dosages of 2.5 to 15 mg had no effect on platelet aggregation.     

Prevalence of low automated platelet counts in cats: comparison with prevalence of thrombocytopenia based on blood smear estimation

Abstract: True thrombocytopenia is uncommon in cats; however, low platelet counts frequently are found using automated cell counters. Although this discrepancy is a well known problem, the prevalence of low automated platelet counts in feline blood samples has not been documented. We retrospectively compared the prevalence of low automated platelet counts with low blood smear-estimated platelet counts in feline blood samples. Results of blood sample analysis from 359 cats during a 1-year period at the University of Glasgow Veterinary Haematology Laboratory were examined. Smear estimates of platelet number were done in those cases in which records did not indicate adequate platelet numbers. Platelet counts obtained with an impedance counter (Minos Vet, Abx Hematologie) were <200×10⁹ cells/L in 256 samples (71%) and <50×10⁹ cells/L in 43 samples (12%). However, based on estimation of platelet numbers from blood smears, only 11 samples (3.1%) had platelet counts of <200×10⁹ cells/L and 9 samples (2.5%) had counts of <50×10⁹ cells/L. Four cats with thrombocytopenia estimated by blood smear evaluation had clinical signs of a bleeding disorder. Disorders associated with thrombocytopenia included neoplasia, cytotoxic chemotherapy, and infectious diseases. There was no evidence that delay due to mailing of samples was associated with lower automated platelet counts than would have been obtained on the day of sampling. The high prevalence of apparent thrombocytopenia in automated platelet counts was attributed to a combination of platelet aggregation and the impedance method of cell differentiation by size. Vigilance and careful examination of blood smears is required to identify the few cats with true thrombocytopenia.     

Evaluation of platelet additive solution for prolonging storage of functional canine platelet concentrate

Objective

To assess storage lesion development, platelet function, and bacterial growth in canine platelet concentrates (PCs) stored in a platelet additive solution (PAS) or a plasma control at 4°C for 21 days.

Design

Prospective, ex vivo, experimental controlled study.

Setting

University veterinary teaching hospital.

Animals

Ten units of canine PCs collected from blood bank donations.

Interventions

The PCs were separated into 2 bags, 1 containing 100% plasma and the other containing 35% plasma and 65% of a PAS (Plasma-Lyte A), and stored at 4°C for 21 days. At days 0, 7, 14, and 21, PCs were analyzed for the presence of swirling, aggregate formation, platelet counts, platelet indices, glucose, lactate, lactate dehydrogenase, Pvco ₂, Pvo ₂, aggregation via light aggregometry, activation percentages using flow cytometry, and bacterial growth.

Measurements and main results

Cold-stored PCs in both PAS and plasma control maintained mean pH >6.8 and mean lactate <9.0 mmol/L over 21 days, with no difference in glucose utilization. Swirl was maintained in both solutions for most days (76/80 combined total samples), with no difference in aggregate formation between solutions. The Pvco ₂ was higher in plasma on all days (P < 0.001), with no difference in Pvo ₂. Platelet indices did not reflect significant storage lesion development in either solution. Lactate dehydrogenase did not differ between solutions but did increase from day 7 to day 21. Mean maximal aggregation percentage was reduced overall but with no significant difference between solutions. The only observed difference in mean activation percentage between solutions was in PAS on day 7, which was significantly higher than plasma (P < 0.05). No bacterial growth occurred during storage.

Conclusions

Cold storage in PAS and plasma allowed PCs to be stored for up to 21 days with minimal storage lesion development, maintenance of platelet function, limited platelet activation, and no bacterial growth within stored bags.     

Evaluation of platelet count and its association with plateletcrit, mean platelet volume, and platelet size distribution width in a canine model of endotoxemia

BACKGROUND: Platelets are of great importance in the pathogenesis of endotoxemia. Although thrombocytopenia is used as a diagnostic sign of endotoxemia, changes in values for platelet indices (plateletcrit [PCT], mean platelet volume [MPV], and platelet size distribution width [PDW]) in response to endotoxin are still unknown. OBJECTIVE: The aim of this study was to evaluate platelet count and its relations with platelet indices in a canine model of endotoxemia. Methods: Twenty dogs were divided into 2 groups of 10 each, and treated intravenously with Escherichia coli endotoxin (1 mg/kg) or vehicle. Venous blood samples were collected before treatment (0 hour) and 0.5, 1, 2, 4, 6, 8, 12, and 24 hours after treatment. Platelet counts and indices were determined on a CELL-DYN hematology analyzer. RESULTS: The platelet count and PCT decreased by a mean of 73% and 93%, respectively (P<.001), at 0.5 hour, and remained 70% and 85% lower than baseline values (P<.001) for 24 hours after endotoxin injection. MPV and PDW increased by a mean of 28% and 45%, respectively (P<.01), at 0.5 hour, and remained increased by 7% and 16% over baseline values for 24 hours (P<.01-.001). Platelet count correlated positively with PCT (P<.001), but correlated negatively with MPV (P<.001) and PDW (P<.01). CONCLUSIONS: Changes in platelet count and its association with platelet indices may reflect changes in platelet production and reactivity. Platelet indices have potential value in the diagnosis and monitoring of dogs and humans with endotoxemia.     

Evaluation of platelet function screening tests to detect platelet procoagulant deficiency in dogs with Scott syndrome

Background: Clinical diagnosis of platelet dysfunction is complex and technically challenging. The wide repertoire of platelet responses requires a test panel to assess different parameters of platelet reactivity. While “global” hemostasis analyzers and whole blood assays have potential for testing platelet function, their ability to evaluate platelet procoagulant activity is ill‐defined. Objectives: The aim of this study was to determine whether platelet procoagulant deficiency, the pathophysiologic defect of Scott syndrome, could be detected in point‐of‐care and whole blood assays. Methods: Study subjects were 4 Scott syndrome‐affected German Shepherds and 8 control dogs. We evaluated 2 point‐of‐care instruments: the platelet function analyzer (PFA‐100) and thromboelastograph (TEG). TEG analysis was performed on recalcified citrated whole blood with and without tissue‐factor activation. A whole blood flow cytometric assay was configured to detect thrombin‐induced platelet P‐selectin expression and platelet‐derived microparticle release. Cytometric samples were analyzed after 1 hour and 1 day of storage. Results: We found no significant differences between Scott and control dogs in PFA‐100 COL/ADP closure times or in any TEG parameter in tissue‐factor–activated samples. In nonactivated samples, mean clotting time (K) and time to maximal rate of thrombus generation were significantly prolonged in Scott dogs; however, values overlapped with those of control dogs. Cytometric analysis of samples from Scott dogs showed significantly diminished platelet‐derived microparticle release. Samples from all dogs reanalyzed after 1 day of storage had nonspecific increases in basal P‐selectin expression and vesiculation. Conclusion: A whole blood cytometric assay to detect stimulated platelet microparticle release can be used to screen for Scott syndrome. However, platelet activation artifacts preclude overnight storage for next‐day analysis.     

Evaluation of methods of platelet counting in the cat

Platelet counts were performed in 50 cats presented for diagnostic investigation. For each cat, counts were obtained using a manual haemocytometer method and compared with counts obtained by estimation from a stained blood smear, a QBC VetAutoread analyser, a Zynocyte VS/2000 analyser, impedance automated counts on a Baker System using both EDTA and citrated anticoagulated blood, and use of a Zynostain modified counting chamber kit. None of the methods gave high correlation with the haemocytometer counts. The blood smear estimation of platelet counts had the highest correlation (r = 0.776) and was the only method to have reasonable values for both sensitivity and specificity. With the impedance automated counts, citrated anticoagulated blood had marginally higher correlation than EDTA anticoagulated blood, and the time between blood sampling and platelet count determination had no effect on the count obtained. When in-house analyser or impedance automated platelet counts are abnormal or not consistent with clinical findings, the authors recommend that a manual platelet count using either haemocytometry or examination of a blood smear is performed.     

Characteristics, diagnosis, and treatment of inherited platelet disorders in mammals

Inherited intrinsic platelet disorders have been identified in dogs, cattle, horses, and cats as well as other animals. The prevalence of mutations in some breeds is high, making these disorders potentially as common as von Willebrand disease in certain breed lineages.     

Triggers for prophylactic use of platelet transfusions and optimal platelet dosing in thrombocytopenic dogs and cats.

Prophylactic platelet transfusions are frequently given to human patients with hypoproliferative thrombocytopenia. For several decades, the most common transfusion trigger was 20,000/microL, but the trend is now to use 10,000/microL in the absence of other risk factors for bleeding. This trigger seems to reduce the number of transfusions without increasing the risk of severe bleeding. Most studies involved in establishing platelet transfusion policies have involved patients with acute leukemia, with fewer studies involving patients undergoing hematopoietic stem cell transplantation or aggressive chemotherapy for other cancers and patients with aplastic anemia. In the presence of other risk factors for spontaneous bleeding, 20,000/microL is still considered an appropriate trigger. The trigger for prophylactic transfusion before surgery has not undergone the same recent scrutiny as has the trigger for spontaneous bleeding. The recommendation remains to raise the platelet count to 50,000 to 100,000/microL if possible, although it is recognized that surgery and other invasive procedures have been performed at lower platelet counts without major bleeding. Prophylactic transfusion is not used in disorders of platelet consumption and destruction to prevent spontaneous bleeding but is used before surgery. Because of the comparative lack of experience with platelet transfusion in veterinary medicine, it is difficult to make generalizations for dogs and cats. Using the guidelines established for therapeutic and prophylactic transfusion of human patients is a reasonable starting point, however. A therapeutic transfusion policy is suggested in the veterinary setting provided that the patient can be closely observed for critical bleeding and a prompt transfusion can be given. This policy should ultimately reduce the overall number of platelet transfusions given to hospital patients. If an animal cannot be closely observed or the ability to transfuse on demand is limited, prophylactic transfusion is recommended. The triggers for initiating a platelet transfusion in dogs are extrapolated from human data; these values are lower by 50% for cats. Because of the imprecision of platelet counting at low values, platelet counts must always be interpreted in conjunction with clinical signs of hemorrhage. If platelet-rich plasma or platelet concentrate is available, a dose of 1 platelet unit per 10 kg is recommended, although resources may dictate a smaller dose. This will raise the recipient platelet count by a maximum of about 40,000/microL. Assuming a trigger of 10,000/microL, a transfusion will probably be required approximately every 3 days. It must be remembered that the frequency of platelet transfusions may be greater in the presence of factors accelerating platelet loss or destruction. If fresh whole blood is used, a rule of thumb is to transfuse 10 mL/kg, which will raise the recipient platelet count by a maximum of approximately 10,000/microL. Daily transfusions or transfusions every other day will probably be required.     

Effects of the glycoprotein IIb/IIIa antagonist abciximab on thrombus formation and platelet function in cats with arterial injury

An established model of arterial injury was used to evaluate the ability of the glycoprotein (GP) IIb/IIIa antagonist abciximab to inhibit platelet function and deter thrombus formation in cats. The study was a blinded evaluation, with control cats receiving aspirin and placebo and treatment cats receiving aspirin plus abciximab. The group treated with aspirin plus abciximab had significantly (P<.05) less thrombus formation based on a smaller number and frequency of cyclic flow reductions and reduced luminal thrombus area than cats treated with aspirin plus placebo. Abciximab administered in addition to aspirin also resulted in significant (P<.05) inhibition of platelet function based on mucosal bleeding time and change in bleeding time from baseline.     

A comparison of platelet parameters in EDTA- and citrate-anticoagulated blood in dogs

BACKGROUND: Platelet aggregates are a common artifact in canine blood. Aggregates may affect the accuracy of platelet counts, with important consequences for patient care. OBJECTIVES: The purpose of this study was to determine if platelet counts in dogs were more accurate if blood was collected into citrate instead of EDTA as an anticoagulant. METHODS: Blood was collected from 50 dogs with neoplasia admitted to the oncology service at Cornell University. EDTA and citrate Vacutainer tubes were filled with blood in random order. Platelet counts and parameters (mean platelet volume [MPV], platelet distribution width [PDW], mean platelet component concentration [MPC], platelet component distribution width [PCDW], and automated platelet clump count [APCC]) were determined using an optical-based hematology analyzer (ADVIA 120). Blood smears from each anticoagulated sample were scored visually for platelet aggregates. RESULTS: The median platelet count was significantly lower (median decrease, 27 x 10(9)/L) in citrate-anticoagulated blood compared with EDTA-anticoagulated blood. This was attributed to platelet activation and aggregation: significantly more aggregates were seen in smears of citrate- than of EDTA-anticoagulated blood. Aggregates were typically small and not detected by the analyzer. Also, the MPV and MPC (or density) were significantly higher (median increase, 3 fL) and lower (median decrease, 33 g/L) in citrate-anticoagulated samples, respectively. CONCLUSIONS: Platelets aggregate, likely from activation, when blood from dogs with neoplasia is anticoagulated with citrate for hematology testing, resulting in lower platelet counts. Citrate also yields inaccurate results for MPV and MPC, likely because of inadequate sphering of platelets. Thus, we recommend that citrate not be used as an anticoagulant when accurate platelet counts are desired in dogs.     

Evaluation of platelet activation in canine immune-mediated haemolytic anaemia

Objectives : To establish whether heightened platelet activation is a common feature of canine immune-mediated haemolytic anaemia, and to evaluate the hypothesis that platelet activation plays a role in the pathogenesis of thromboembolism. Methods : Using whole-blood flow-cytometric analysis, the proportion of activated platelets and platelet-leucocyte aggregates in blood samples from 14 dogs with immune-mediated haemolytic anaemia and 14 healthy dogs was calculated. General linear models with binomial errors were used to compare groups. Results from the immune-mediated haemolytic anaemia-affected dogs were then correlated with established risk factors for thromboembolism in canine immune-mediated haemolytic anaemia, D-dimer concentration and antithrombin activity. Results : There was a strong correlation between platelet activation and severe thrombocytopenia, with heightened platelet activation being observed predominantly in severely thrombocytopenic dogs. Clinical Significance : Dogs with immune-mediated haemolytic anaemia, particularly those with concurrent severe thrombocytopenia, are likely to have heightened platelet activation, which may play a role in the pathogenesis of thromboembolism.     

Evaluation of platelet aggregation using a point-of-care instrument in retired racing Greyhounds

Veterinarians involved in Greyhound rescue have anecdotally observed that 10-15% of Greyhounds bleed profusely after simple surgical procedures. In most patients, platelet counts and hemostasis profiles are normal; therefore, it is possible that these dogs have platelet dysfunction. The PFA-100 is a novel point-of-care platelet function analyzer that has recently been evaluated as a rapid method to assess platelet function in dogs. The objectives of this study were to characterize platelet function in a group of healthy Greyhounds by means of the PFA-100. Blood samples were collected from the jugular vein from 30 healthy Greyhounds. CBC, biochemical profile, PFA-100 assay with collagen/epinephrine (COL-EPI) and collagen/ adenosindiphosphate (COL-ADP), plasma von Willebrand factor antigen concentration (vWF:Ag), and vWF collagen-binding assay (vWF:CBA) were performed. PFA-100 closure times (CTs) with COL/ADP ranged from 63 to 92 seconds (mean +/- SD, 74.7 +/- 7.9 seconds) and with COL/EPI from 87 to 238 seconds (138 +/- 41 seconds); vWF: Ag ranged from 22 to 120% (87.52 +/- 25.5%) and vWF: CBA ranged from 36 to 102% (77.4 +/- 17.3%); and platelet counts ranged from 147 to 265 x 10(9)/L (194.6 +/- 31.64 x 10(9)/L). Greyhound CTs were significantly shorter than CTs in a mixed population of 50 healthy non-Greyhound dogs, in which the COL/ADP CTs ranged from 61 to 172 seconds (mean +/- SD, 87 +/- 21.6 seconds), and the COL/ EPI CTs ranged from 81 to 300 seconds (mean +/- SD, 183 +/- 67.6 seconds; P = 0.005 for COL/ADP CT; P = 0.001 for COL/ EPI CT). Also, platelet counts were significantly lower (P = 0.001) and packed cell volume was significantly higher (P = 0.001) in the Greyhound than in the non-Greyhound group. The PFA-100 is a reproducible method that can be used in the clinical setting to assess platelet function in Greyhounds; however, normal CTs in healthy Greyhounds are shorter than in other breeds. The results obtained in this study will be used to screen for abnormal platelet function in Greyhounds with postoperative bleeding.     

Platelet function in clinically healthy cats and cats with hypertrophic cardiomyopathy: analysis using the Platelet Function Analyzer‐100

Background: There is currently no simple analytical tool for the evaluation of hypercoagulability in cats. The Platelet Function Analyzer‐100^® (PFA‐100; Dade Behring Inc., Deerfield, IL, USA) is a bench‐top machine that evaluates platelet function by measuring closure time (CT) in citrated whole blood under high shear conditions. We hypothesized that cats with hypertrophic cardiomyopathy (HCM) have up‐regulated platelet function, which shortens their CT and increases their risk for thromboembolic events. Objectives: The goals of this study were to: (1) establish a feline reference interval for CT using the PFA‐100, (2) measure CT in blood from cats with HCM, and (3) determine if there is a measurable difference between the CT of healthy cats compared with cats with HCM. Methods: Citrated blood samples from 42 clinically healthy cats and 30 cats with HCM were analyzed according to manufacturer's specifications. CT was measured in triplicate and the mean value was used for analysis. Transformed data were compared between clinically healthy cats and cats with HCM using a Student's t‐test, and among cats with mild, moderate, or severe HCM using ANOVA. Results: The median CT of clinically healthy cats was 64 seconds (range 43–176 seconds). The median CT of cats with HCM was 74 seconds (range 48–197 seconds). There was no significant difference in CT between cats with HCM and clinically healthy cats. There also were no significant differences in cats with mild, moderate, or severe HCM. Conclusions: A feline reference interval for PFA‐100 CT will be useful in future studies of platelet function in cats. Cats with HCM do not have shorter CTs when compared with clinically healthy cats.     

Assessment of platelet function in horses: ultrastructure, flow cytometry, and perfusion techniques

We studied equine platelet function and activation using ultrastructural examination, flow cytometry, and perfusion. The main aim of the study was to evaluate hemostatic mechanisms in horses using these techniques. Ultrastructural observations were done on resting and activated platelets. Flow cytometry was used to evaluate binding of antibodies to major platelet glycoproteins (GPIIb-IIIa, GPIV, and GPIb) and activation-dependent antigens (P-selectin and lysosomal integral membrane protein [LIMP]). Perfusion techniques were used to evaluate the interaction between platelets and damaged subendothelium. Aggregation experiments were done to identify the best agonists for flow cytometry. Ultrastructural observations confirmed that equine platelets lack a developed open canalicular system and that release of granule contents occurs by fusion of adjacent granule membranes that ultimately connect with external membranes. Flow cytometry identified a 2-fold increase in binding of antibodies against GPIIb-IIIa and GPIV after activation. Binding of antibodies against P-selectin and LIMP increased from 2.12 and 1.74% to 15.5 and 11.6%, respectively, in response to thrombin and to 21.86 and 10.50%, respectively, in response to collagen. Annexin V binding increased moderately after activation. Perfusion experiments with citrated blood indicated that equine platelets react more strongly to subendothelium than do human platelets. When blood was anticoagulated with low molecular weight heparin, a marked impairment of platelet interactions was observed. In conclusion, although some differences were observed between human and equine platelet function, some techniques currently used to assess human platelet function may be useful to assess equine platelets.