Primary Immune-Mediated Thrombocytopenia in Cats
Feline primary immune-mediated thrombocytopenia (pIMT) is a rare condition, and only a few cases have been described in veterinary literature. Five cats with severe thrombocytopenia most likely due to pIMT are described. A flow cytometry platelet-bound antibody test was positive in all cats; underlying diseases or triggering factors causing thrombocytopenia were not detected. Three cats were transfused with blood type-compatible fresh whole blood; one cat received Oxyglobin as well. All cats were treated with prednisolone; one cat received chlorambucil in addition. Four cats responded to treatment and were discharged from the hospital. One cat was euthanized due to dyspnea. Primary immune-mediated thrombocytopenia is rarely diagnosed in cats, but it is important as a differential diagnosis in cats presented with surface bleeding.
Introduction
Thrombocytopenia is a common laboratory finding in cats.1 However, low platelet counts are often incorrect due to technical problems using automated cell counters.2,3 An increased destruction of platelets (immunemediated thrombocytopenia [IMT]), reduced production within the bone marrow, increased utilization (due to disseminated intravascular coagulation [DIC]), sequestration in the spleen (due to inflammatory or infiltrative disorders), or a combination of these pathomechanisms can cause thrombocytopenia. 4,5 Underlying diseases or triggers that can cause thrombocytopenia in cats include: viral infections (e.g., feline infectious peritonitis [FIP], feline leukemia virus [FeLV], feline immunodeficiency virus [FIV], panleukopenia); bacterial infections (e.g., pneumonia, cystitis, pyelonephritis, pyometra); aseptic inflammation (e.g., fat-tissue necrosis, trauma, pancreatitis, hepatitis, feline lower urinary tract disease); neoplasms (e.g., lymphoma, leukemia, hemangiosarcoma); primary bone marrow disorders (e.g., aplastic anemia, myelodysplasia); drugs (e.g., propylthiouracil, methimazole, griseofulvin, albendazole, chloramphenicol); and IMT.1,6–16
Immune-mediated thrombocytopenia is a disease in which antibodies bind to the surface of platelets, resulting in premature platelet destruction by macrophages. It occurs as a primary (pIMT) or idiopathic disease (the cause of which is unknown) and more commonly as a secondary form. Various diseases may trigger the production of antibodies and lead to an increased destruction of platelets by the mononuclear phagocytic system. 8–10,14,17,18 Primary IMT is a frequent cause of severe thrombocytopenia in dogs. However, only a few cases have been described in cats thus far.1,13,14,16,19,20 Whether pIMT actually occurs less frequently in cats or whether it is only diagnosed less frequently due to a reduced tendency for bleeding in cats is unclarified.
The diagnosis of pIMT is based on a low platelet count, exclusion of underlying diseases or triggering factors, the response to immunosuppressive therapy, and the presence of platelet-bound antibodies. In previous literature, the platelet-factor-3 test and immunofluorescence have been used to detect platelet-bound or antimegakaryocytic antibodies in cats.1,13,21,22 Direct detection of platelet-bound antibodies is a method that has been successfully used in specialized laboratories in dogs; however, little information is available describing this method in cats.14,23 Recently, a flow cytometric assay has been used to detect platelet-bound antibodies in cats.14
The objectives of this study were to describe five cats with pIMT and to review the literature regarding feline pIMT.
Materials and Methods
Medical records were reviewed of five cats with pIMT that were presented to the Small Animal Clinic, Free University of Berlin, between January 2000 and August 2008. Data collected from the medical records included signalment, history, clinical signs, results of laboratory examinations and of diagnostic imaging, treatment, and outcome. The diagnosis was based on a low platelet count, exclusion of underlying diseases or triggering factors, the presence of platelet-bound antibodies, and the response to immunosuppressive therapy.
A complete blood count, manual differential cell counts, biochemical analysis, and a coagulation profile were performed using standard laboratory methods.a,b The platelets were counted microscopically using Thrombo Plus tubesc and a Neubauer counting chamber to confirm the low platelet counts. All blood samples were macroscopically and microscopically evaluated for erythrocytic agglutination. In case of spontaneous erythrocytic agglutination, the red blood cells (RBC) were washed three times with physiological saline solution, and the macroscopic and microscopic evaluations for RBC agglutination were repeated. For reticulocyte counting, brilliant cresyl blue smears were prepared. Serological tests (FIV antibodies [n=5], FeLV antigen [n=5]) and a polymerase chain reaction (PCR) for Hemoplasma spp. (n=5) and Anaplasma (A.) phagocytophilum (n=1) were performed in external laboratories. The direct Coombs’ test (n=3)d and the platelet-bound antibody test (n=5)e were performed at the Institute of Immunology, University of Veterinary Medicine, Hannover, Germany, as previously described.14 Along with each cat’s blood sample (approximately 1 to 1.5 mL of potassium-ethylenediaminetetraacetic acid [K-EDTA] blood), a control sample (K-EDTA blood of a healthy cat) was sent overnight without cooling and was analyzed simultaneously. Platelet-bound antibody testing was negative in five control cats.14
Initially the cats were treated with an intravenous injection of a fast-acting glucocorticoid (methylprednisolone 10 mg/kg once) or with oral prednisolone (1 to 2 mg/kg q 12 hours). If the platelet counts were in the reference range, the dose of prednisolone was reduced gradually approximately every 2 to 3 weeks by 15% to 25% of the former dose. In one case, the immunosuppressive drug chlorambucil (0.02 mg/kg for 1 week; then 0.01 mg/kg q 24 hours per os [PO]) was added to the treatment. Furthermore, treatment consisted of the administration of H2-receptor antagonists (ranitidine 1 mg/kg q 12 hours PO), infusions (lactated Ringer’s solution supplemented with potassium), and antibiotics (doxycycline 5 mg/kg q 12 hours PO or enrofloxacin 5 mg/kg q 24 hours PO). In the case of bleeding and severe anemia or if a rapid further hematocrit (HCT) decrease was expected because of hemorrhage, the cats were transfused with blood type-compatible fresh whole blood. One cat received an Oxyglobinf transfusion in addition.24,25
The platelet count was measured daily at the beginning of therapy, then every 2 to 3 days. Once the number of platelets was stable (>100 × 109/L), rechecks were carried out at regular intervals.
Results
History and Clinical Signs
Signalments of the five cats with suspected pIMT are shown in Table 1. All cats had been vaccinated 6 to 12 months before admission. Three cats were strictly kept indoors; one cat (case no. 5) was allowed on the balcony; and one cat (case no. 2) was intermittently outdoors. All cats but one were presented with spontaneous surface bleeding or hematoma formation. During clinical examination, hemorrhages and petechiae were detected in all five cats [Table 1; Figures 1, 2]. One cat (case no. 3) developed episcleral bleeding during the time of hospitalization (on day 2).
Diagnostic Imaging
Conspicuous radiographic and ultrasonographic findings were a mild splenomegaly in three cats (case nos. 1, 2, 5) [Figure 3] and an enlarged cardiac silhouette in one cat (case no. 5).
Laboratory Analyses
The initial manual platelet count revealed severe thrombocytopenia in all cats. Initially, four cats were anemic [Table 2]. One cat (case no. 3) developed anemia with an HCT of 28% within 24 hours, because of acute bleeding. The anemia was regenerative in three of five cats (case nos. 2, 4, 5) at initial presentation; on day 3, the reticulocyte count in case no. 1 was 293,000/μL. In two cats (case nos. 1, 3), a mild erythrocytic agglutination was present, which did not persist after washing the erythrocytes with physiological saline.
The blood smear evaluations that were performed in three cats (case nos. 1, 4, 5) revealed anisocytosis and polychromasia. In one cat (case no. 4), schistocytes and an increased number of nucleated RBCs (11/100 white blood cells [WBCs]) were detected. The manual differential count revealed lymphocytosis and mild monocytosis in one cat (case no. 5) and a mild lymphopenia in another cat (case no. 4). Mild eosinophilia was present in case no. 5. The activated partial thromboplastin time (aPTT) was significantly prolonged in two cats (case nos. 4, 5), and the prothrombin time (PT) was slightly prolonged in one cat (case no. 4). In case no. 4, DIC was suspected as a result of thrombocytopenia, PT and aPTT prolongation, and a few schistocytes on the blood smear. Biochemical analysis revealed a mild hypoproteinemia in one cat (case no. 1), most likely due to blood loss. The increased urea in this cat might have been a result of swallowing blood. Case no. 3 had a hyperglobulinemia. A mild hyperbilirubinemia was present in two cats (case nos. 4, 5). Case no. 4 had an increase of the plasma alanine transaminase and aspartate transaminase (AST) activities; AST was also slightly increased in case nos. 1 and 5.
Serological or PCR tests, respectively, for infectious diseases (e.g., FIV, FeLV, Hemoplasma spp., A. phagocytophilum) were negative in all cats. A direct Coombs’ test was negative in all three cats tested. The platelet-bound antibody test was positive in all cases. In one cat (case no. 2), this test was repeated after 14 days and 3 months. While the first repeat test was still positive, the result was negative after 3 months.
Treatment
Three cats received an intravenous injection of a fast-acting glucocorticoid on the day of initial presentation. All five cats were treated with oral prednisolone starting on day 1 or 2. Since the platelet count in one cat (case no. 5) did not increase within the first 7 days, the immunosuppressive drug chlorambucil was added to the treatment regimen.
All cats received antibiotic treatment (doxycycline [n=2], enrofloxacin [n=3]), and four cats were infused with electrolyte solutions. Ranitidine was administered in all cats; in addition, one cat (case no. 5) received sucralfate after developing melena (a gastrointestinal ulcer was suspected). Because of acute bleeding and severe anemia, three cats (case nos. 1, 3, 4) were transfused with compatible fresh whole blood up to two times (4 to 7.5 mL/kg body weight per transfusion) within the first 2 days of hospitalization. One cat (case no. 4) received Oxyglobinf in addition, as no blood was available at that time. For one cat (case no. 5), a blood transfusion was considered, but a compatible donor (blood type B) was not available.
Course of Disease
Three cats (case nos. 1, 2, 3) improved within 3 days, and an increase of the platelet counts >50 × 103/μL was detected. Five to 12 days after treatment with glucocorticoids, the platelet counts were >140 × 103/μL in these three cats. In two cats (case nos. 4, 5), the platelet counts remained <20 × 103/μL in the first 7 days, despite immunosuppressive treatment with glucocorticoids. The immunosuppressive drug chlorambucil was added to the treatment regimen of case no. 5 from day 7 onward (0.2 mg/kg for 7 days, then 0.1 mg/kg). On day 9, a decrease in HCT to 19% was noted. The cat had vomited twice and had passed very dark feces (melena); for that reason, gastrointestinal ulcer formation from high-dose glucocorticoid therapy was suspected. The prednisolone dosage was decreased to 0.5 mg/kg, and sucralfate was administered in addition to ranitidine. No further gastrointestinal bleeding was noted. The platelet count increased from initial 5 × 103/μL to 17 × 103/μL on day 10, and to 695 × 103/μL on day 13. On day 36, the platelets had dropped to 46 × 103/μL. The prednisolone dosage was increased (1 mg/kg q 24 hours), and chlorambucil was administered at a dosage of 0.2 mg/kg. Due to a decrease of the WBC counts on day 42, the chlorambucil dose was reduced (0.1 mg/kg). On day 49, the platelet count was 90 × 103/μL; a further increase to 150 × 103/μL was noted on day 60.
Case no. 4 developed dyspnea on day 5, which deteriorated progressively until day 7. A follow-up radiographic examination of the thorax and an echocardiography were unremarkable. On day 8, the cat was euthanized because of severe dyspnea. A pulmonary thromboembolism was suspected.
Four cats were discharged from the hospital and were observed over a period of 12 to 120 days. Follow-up examinations were performed either by the Small Animal Clinic at the Free University of Berlin or the referring veterinarian. Case no. 1 was monitored for 2 months. On day 21 (platelet count 178 × 103/μL), the cat developed signs of an upper respiratory tract infection. The cat was treated with immune serum (Feliserin),g enrofloxacin (5 mg/kg q 24 hours PO), and oxytetracycline eye ointment for 7 days; the prednisolone dose was reduced to 0.75 mg/kg q 12 hours. Clinical symptoms were resolved on day 28, and the manual platelet count was 325 × 103/μL. Prednisolone was reduced to 0.5 mg/kg q 12 hours. The follow-up after 2 months was performed by the referring veterinarian, and the platelet count was 408 × 103/μL at that time. The dosage of prednisolone was 0.5 mg/kg q 48 hours. Case no. 2 was observed over a period of 3 months (manual platelet count after 3 months: 172 × 103/μL), and the test for platelet-bound antibodies was repeated with a negative result. One cat (case no. 3) could only be followed up until day 12 (manual platelet count: 240 × 103/μL). Case no. 5 was presented for follow-up consultations every 2 to 4 weeks. At the time of writing (4 months after initial presentation), the platelet counts were in the reference range, and no further relapse was noted (prednisolone dosage: 0.25 and 0.5 mg/kg on alternate days; chlorambucil dosage: 0.05 mg/kg).
Discussion
Five cats suffering from pIMT were presented with spontaneous surface bleeding or hematoma formation (bleeding from mouth or dental alveoli [n=2], epistaxis [n=1], petechiae on pinna [n=2], petechiae and ecchymoses of the abdomen and hind limb [n=1], and hematoma in facial region and oral mucosa [n=1]). The few cases with suspected pIMT described in the literature were presented with epistaxis (n=3), hematuria (n=2), petechiae on the ears (n=1), bleeding from multiple sites (n=1), or hemoptysis (n=1). The clinical examinations of these cats revealed petechiae and ecchymotic hemorrhages on the ventral abdomen (n=4), the ears (n=2), tongue (n=1), and mucous membranes (n=1).1,13,16,19,20
Spontaneous bleeding due to thrombocytopenia has been described less frequently in cats than in dogs. In one study, only five of the nine cats with platelet counts <10 × 103/μL were bleeding.1 In another study, seven of 63 cats suffering from thrombocytopenia displayed spontaneous hemorrhage; their platelet counts ranged from 10 to 57 × 103/μL.15 Spontaneous bleeding is observed rarely with platelet counts >20 to 40 × 103/μL if there is no concomitant thrombo-, vasculo-, or coagulopathy. Whether bleeding occurs or not is dependent on the number of platelets and their age and functionality. One cat had a platelet count >46 × 103/μL, which usually does not result in spontaneous bleeding. However, in cats with IMT, a platelet dysfunction in addition to thrombocytopenia was described, which might increase the bleeding tendency.26 Cats have a very efficient hemostatic system, and they seem to tolerate low platelet counts more readily than other species. Feline platelets are more easily stimulated by aggregation inductors (such as collagen, adenosine diphosphate, or thrombin) than those of dogs. The dense granules of cats were shown to contain three or one-and-a-half times the amount of serotonin compared to dense granules of humans or dogs, respectively. Serotonin is a potent vasoconstrictor and induces primary aggregation of platelets. In addition, serotonin will potentiate the aggregation effect of other agonists.27,28
According to various studies, no adequate correlation exists between automated and manual platelet counts in cats.2,15 This is because of the tendency for feline platelets to aggregate and the difficulties in differentiating between the similar size of some feline platelets and RBCs.2,28 Thus, a manual count immediately after blood withdrawal or an evaluation of the blood smear should always be undertaken to confirm a low platelet count.3,15,29 However, the particular tendency for feline platelets to aggregate cannot be avoided even with manual counting.
Case reports on primary IMT in cats are few.1,13,14,16,19,20 For detection of antimegakaryocytic antibodies, direct and indirect immunofluorescence tests have been described.13,19,22 Joshi et al (1979) used the platelet-factor-3 test, which is an indirect method to detect platelet-bound antibodies in the serum.19 Direct methods for detection of platelet-bound antibodies are much more reliable. In the cases examined here, flow cytometry was used for direct platelet-bound antibody testing.14 A disadvantage of all methods (including flow cytometry) is that positive test results do not allow for distinguishing between primary and secondary IMT. Platelet-bound antibodies may be antiplatelet autoantibodies, but they could also represent “secondary” antibodies. The latter include immune complexes (which are bound to platelet Fc-receptors), antibodies against platelet antigens (which are formed by modification of the antigen [e.g., caused by disease]), or antibodies that bind to antigens (which were adsorbed by the surface of thrombocytes [e.g., tumor antigens, drug metabolites]).30 In a study of 42 cats displaying thrombocytopenia, a direct platelet-bound antibody test using flow cytometry was performed. Nineteen cats were tested positive. In only two of these cats (i.e., those with pIMT and Evans syndrome), an underlying disease was not found. The other 17 cats suffered from viral and bacterial infections (e.g., FIP, FeLV, FIV, pyelonephritis), inflammatory diseases (e.g., fat-tissue necrosis, hepatitis), neoplasias (e.g., lymphoma, leukemia), or hyperthyroidism (i.e., secondary IMT).14
In the five cases of this study, underlying diseases or triggering factors were excluded. None of the cats had been vaccinated within the last few months. Diagnostic imaging studies as well as infectious disease screening did not reveal underlying diseases either; nevertheless, infectious diseases cannot be excluded completely. Coagulation parameters were within the reference range in four cats, and therefore DIC was unlikely. However, DIC was suspected in one cat displaying a prolonged aPTT and PT. An explanation of the DIC in this cat might have been severe blood loss and hypovolemia, followed by hypoxia and shock. All cats displayed mild to severe anemia that was attributed to blood loss. However, IMT can occur in association with immune-mediated hemolytic anemia (Evans syndrome). Evans syndrome has been described rarely in cats.14,19,22 The fact that one of the cats suffered from Evans syndrome cannot be completely excluded. However, three cats had no agglutination of erythrocytes; two cats had only mild, nonpersistent agglutination; and three cats (one with and two without agglutination) had a negative direct Coombs’ test. Two cats had a mild hyperbilirubinemia. Since hepatic or posthepatic causes seemed unlikely in these cats, the hyperbilirubinemia was presumed to be caused by the breakdown of hemoglobin originating from subcutaneous bleeding and hematomas.
In this study, ultrasonography revealed splenomegaly with homogeneous parenchyma in three cats. Similar to other immune-mediated diseases, enlarged spleens may be related to lymphoreticular hyperplasia or may be caused by extramedullary hematopoiesis.
Therapy of IMT is based on the use of immunosuppressive drugs. In the cases examined here, three cats displaying severe bleeding initially received fast-acting glucocorticoids intravenously, since oral administration was not possible. Subcutaneous injections are to be avoided in animals suffering from IMT. The effect of glucocorticoids is based on various mechanisms. In addition to their antiinflammatory effect, they cause a reduction of circulating lymphocytes, reduce the number of Fc-receptors on the surfaces of macrophages, and stabilize lysosomal membranes.31,32 While thrombopoietin is mainly responsible for enhanced production and division of megakaryocytes, a stimulating effect of glucocorticoids on platelet production and their release from megakaryocytes was suggested in dogs and humans.33 In humans and dogs, a decrease in platelet-bound antibodies and antibodies against platelets in the serum due to glucocorticoid therapy has been detected.26,34
All of the five cats presented here received an H2-receptor antagonist to prevent gastrointestinal ulcers. In three cats suffering from pIMT, an increase in platelet counts was observed a few days after initiation of therapy. Monitoring of the animals at regular intervals during IMT treatment is important. Tasker et al (1999) described a cat suffering from IMT, in which neither oral therapy of prednisolone (3 mg/kg q 24 hours) given by itself nor in combination with azathioprine (0.3 mg/kg q 24 hours) or vincristine (0.4 mg/m2) proved effective.13 An increase in platelet count was observed only after changing to dexamethasone (0.64 mg/kg q 24 hours PO) in combination with azathioprine.
Bianco et al (2008) described four cases of presumed pIMT. One cat showed platelet counts within the reference range a few days after oral administration of prednisolone (1 mg/kg q 12 hours). Two cats responded well to prednisolone therapy (1 mg/kg q 12 hours) initially, but they developed thrombocytopenia again after the dosage was reduced by 25%. One of these cats responded well to oral administration of cyclosporine (5 mg/kg q 12 hours), and the other cat improved with intravenous administration of dexamethasone (1 mg/kg q 24 hours). The fourth cat received dexamethasone (0.3 mg/kg q 12 hours) intravenously and a single intravenous administration of vincristine (0.02 mg/kg). However, this cat did not respond to therapy and was euthanized. Two of these four cats developed transient diabetes mellitus, and one cat had recurrent bacterial urinary tract infections due to long-term corticosteroid therapy.16 Garon et al (1999) described one cat that improved only with a combination therapy of prednisolone (2 mg/kg q 12 hours) and cyclosporine (5 mg/kg q 12 hours). Neither single administration of prednisolone nor a combination with vincristine (0.025 mg/kg) caused an increase in platelet count.20
In our study, three of five cats responded to prednisolone therapy. The use of chlorambucil has not been described in cats with IMT thus far. Since only few cases with pIMT have been described in previous literature, it is not possible to make a statement about the advantage of an immunosuppressive treatment regimen in comparison to others. Based on the few available data, prednisolone seems to be a potent treatment. However, side effects such as gastrointestinal ulcers or transient diabetes mellitus are documented in association with the use of glucocorticoids.
Cats displaying severe thrombocytopenia have to be monitored very carefully with regard to bleeding. To minimize the risk of bleeding, cats should be kept inside a cage and fed with soft food. Careful handling and avoidance of subcutaneous or intramuscular injections are important. Moreover, drugs that interfere with platelet production (e.g., nonsteroidal antiinflammatory agents) should not be administered or should be discontinued.5
Three cats displaying acute bleeding received blood type-compatible fresh whole blood to compensate for their anemia and to supply platelets. A transfusion of platelets is very effective if the platelets have a normal survival time. Transfusion is less effective if there is an increased utilization or sequestration of platelets or in cases of IMT, since the transfused platelets are destroyed as quickly as the body’s own platelets. Nonetheless, administration of platelets can be indicated as a life-saving measure in cases of acute bleeding from severe thrombocytopenia or when surgery is necessary. In cats, platelets are usually administered as a fresh whole blood transfusion, since manufacturing of platelet-rich plasma is very difficult because of the small amounts of blood that can be withdrawn.25 A volume of 10 mL/kg fresh blood results in an approximate increase of 10 × 103/μL platelets in the recipient.35 However, this can be enough to stop life-threatening hemorrhage.
In this study, antibiotics were administered in addition to an immunosuppressive therapy. Sick animals that receive glucocorticoid treatment are more prone to develop secondary infections. Moreover, thrombocytopenias have been described in relation to hemoplasma infections.36 Since it took several days for the results of the PCR to come back in the cases described here, antibiotic treatment was started at admission.
Relapse rates of 40% and a mortality rate of 10% have been described in dogs suffering from pIMT.23,37 Little is known regarding these rates in cats, since case studies are few. Of the 13 cats with presumed pIMT examined here and in previous case reports, 11 were released from the hospital (mortality rate 15%).1,13,16,19
Conclusion
Primary IMT is rarely diagnosed in cats, but it should be considered as an important differential diagnosis in cats presented with surface bleeding. The initial laboratory analyses should include a manual platelet count to verify thrombocytopenia, and underlying diseases and triggering factors should be carefully excluded before suspecting pIMT. Direct-flow cytometry to detect platelet-bound antibodies in dogs and cats is a method that has been successfully used in specialized laboratories, but it is rarely available. Comparing different immunosuppressive treatment regimens is not possible because of the few feline cases described in the veterinary literature; however, prednisolone seems to be an efficient treatment.
Activated partial thromboplastin time (aPTT), Pathromtin; Dade Behring, Marburg, Germany
Prothrombin time (PT), Hepato Quick; Diagnostica Stago, Asnieres, France
Sarstedt, Nümbrecht, Germany
Dianova, Hamburg, Germany
FACScan; Becton Dickinson, NJ 07417
Oxyglobin; Biopure Corporation, Firma Albrecht, Germany
Feliserin PRC; IDT Biologica GmbH, Dessau-Rosslau, Germany
Citation: Journal of the American Animal Hospital Association 46, 1; 10.5326/0460012
Citation: Journal of the American Animal Hospital Association 46, 1; 10.5326/0460012
Citation: Journal of the American Animal Hospital Association 46, 1; 10.5326/0460012
Cat (case no. 1) with hemorrhage due to primary immune-mediated thrombocytopenia.
Cat (case no. 5) with primary immune-mediated thrombocytopenia with swollen facial region and oral mucosa due to hemorrhage.
Laterolateral abdominal radiograph of a cat (case no. 1) with primary immune-mediated thrombocytopenia and mild splenomegaly.
Contributor Notes
