Canine Immune-mediated Hemolytic Anemia Research Articles

Clinical presentation, outcome and prognostic factors in dogs with immune-mediated haemolytic anaemia: a retrospective single-centre study of 104 cases in Ireland (2002–2020)

BackgroundImmune-mediated haemolytic anaemia (IMHA) has a high mortality rate within the first weeks to months of diagnosis. Identifying dogs at increased risk of death may help guide decision-making for owners and veterinarians. Prior studies have identified several but inconsistent prognostic factors. The objectives of the study were to describe the clinical presentation and outcome of canine immune-mediated haemolytic anaemia in Ireland and to assess for independent factors associated with survival including long-term survival. Medical records from a single centre were reviewed between 2002 and 2020 to identify dogs with immune-mediated haemolytic anaemia using the American College of Veterinary Internal Medicine (ACVIM) consensus statement algorithm. Survival analysis was performed using univariable Cox proportional hazards regression models with Breslow method for ties to identify prognostic factors.ResultsOne hundred and four cases were included. The diagnosis of immune-mediated haemolytic anaemia was classified as definitive, supportive and suspicious in 42 (40%), 50 (48%), and 12 dogs (12%) respectively. Twenty-two dogs (21%) were diagnosed with associative IMHA and 82 dogs were diagnosed with non-associative IMHA (79%). 65% of the cases received more than one immunosuppressive medication during the course of treatment. The mortality rate at one and three months was 16% (95% confidence interval [CI] 9–26) and 31% (95% CI 21–43) respectively. Excluding dogs that died within three months, the median survival time was 2664 days. The relapse rate during the follow-up period was 7%. Survival did not improve over the course of the study period. Thrombocytopenia and hyperbilirubinaemia were identified as negative prognostic indicators (Hazard ratio 2.2 and 2.5, 95% CI 1.1–4.1 and 1.1–5.6, respectively).ConclusionsExcluding dogs that died within three months, the outcome was good in dogs with non-associative immune-mediated haemolytic anaemia in Ireland. The relapse rate was low regardless of the presence of associative causes. Thrombocytopenia and hyperbilirubinaemia were the only independent negative prognostic factors. The one-month and three-month mortality rates were similar compared to prior studies and survival did not improve over time during the study period: the mortality rate of canine immune-mediated haemolytic anaemia remains high in the acute phase.

Immune-Mediated Hemolytic Anemia and Clinically Suspected Acute Pancreatitis in Dogs, a Pilot Study

Acute pancreatitis can be a complication of massive hemolysis, above all when intravascular in nature. Therefore, the aim of this study was to investigate the association between canine immune mediated hemolytic anemia (IMHA) and clinically suspected acute pancreatitis (CSAP) and the role of calculated free plasma hemoglobin (Hbfp) in CSAP occurrence/development. In this cohort study the records of 95 dogs with IMHA and 95 sick dogs with pathologies other than IMHA were compared for CSAP occurrence/development. At presentation, 12/95 dogs with IMHA met criteria for CSAP, while only 3/95 sick control dogs met these criteria (χ2 =1.58, P = .008). Within 7 days of hospitalization 9 additional dogs with IMHA had developed CSAP. The Hbfp was calculated and compared for dogs with IMHA that had/developed CSAP and for those without CSAP. In dogs with IMHA, a calculated Hbfp concentration ≥ 0.08 g/dL resulted in an increased relative risk (RR) of having/developing CSAP (RR = 2.54, 95% CI, 1.51-4.29; P = .003). No significant effect on short-term prognosis in dogs with IMHA was found between those having/developing CSAP and those without CSAP. This study showed that dogs with IMHA have an increased risk of having CSAP and Hbfp concentration may be involved in the pathogenesis of acute pancreatitis.

Preliminary Study on Treatment Outcomes and Prednisolone Tapering after Marine Lipid Extract EAB-277 Supplementation in Dogs with Immune-Mediated Hemolytic Anemia.

Immune-mediated hemolytic anemia (IMHA) is a common autoimmune disorder in dogs with a high fatality rate and it remains a therapeutic challenge. The marine lipid extract, EAB-277, is a natural anti-inflammatory nutraceutical product. However, the effects of EAB-277 in IMHA dogs has rarely been investigated. The objective of this study is to assess the clinical effects of EAB-277 and prednisolone dose-tapering for supplemental therapy in IMHA dogs. Prednisolone was given to 18 anemic IMHA dogs according to a standard regimen. Six dogs were supplementally treated with EAB-277 for 28 days and the remaining twelve dogs were a control group of untreated supplementations. The results demonstrate that the supplement group showed slightly better survival rates (66.7 ± 19.2%) than the control group (16.7 ± 0.7%), but the difference was not statistically significant (p = 0.408). When compared to pre-therapy, the supplement group's blood profiles improved (p < 0.05). The EAB-277 treated group showed a moderate decrease in the incidence rate (4.20 times) of prednisolone tapering compared to the control group. The dosage reduction of prednisolone in supplement group was more than that in the control group (p < 0.0001). Our results suggest that EAB-277 supplementation may enhance clinical outcomes and lessen prednisolone dose-tapering in canine IMHA therapy.

Utility and prognostic significance of leukocyte ratios in dogs withPrimary Immune-Mediated Hemolytic Anemia.

Canine immune-mediated hemolytic anemia (IMHA) is a life-threatening condition that is commonly associated with neutrophilia and monocytosis. Leukocyte ratios have been found to have prognostic value in humans and animals affected by a range of inflammatory, infectious, and neoplastic disorders. We hypothesized that in primary IMHA, neutrophil to lymphocyte (NLR), neutrophil to monocyte (NMR), band neutrophil to segmented neutrophil (BNR) and monocyte to lymphocyte (MLR) ratios would be higher in dogs that did not survive to discharge. Medical records of dogs diagnosed with IMHA at two veterinary teaching hospitals were retrospectively reviewed. Twenty-three of the 72 included dogs did not survive to discharge. NLR, NMR, BNR and MLR ratios were compared between dogs that survived to discharge and dogs that died or were euthanized. None of the ratios were significantly different between survivors and non-survivors (P = 0.14-0.99). Area under the Receiver Operating Characteristic (ROC) curve for prediction of non-survival ranged from 0.5 (95% confidence interval 0.38-0.62) for MLR to 0.61 (0.49-0.72) for NMR and was not significantly different from 0.5 for any ratio (P = 0.29-0.99). After exclusion of 31 dogs that received one or both immunosuppressive medications and blood transfusion before presentation, the area under the ROC curve for prediction of survival was significantly different from 0.5 for MLR (0.78, P = 0.01) and NMR (0.78, P = 0.0002). This study suggests that lower MLR and higher NMR may predict poorer prognosis in untreated dogs with IMHA.

Centrifugal therapeutic plasma exchange in dogs with immune-mediated hemolytic anemia (2016-2018): 7 cases.

To describe the technique of centrifugal therapeutic plasma exchange (cTPE) in dogs diagnosed with immune-mediated hemolytic anemia (IMHA) and summarize the outcome of the procedure. Retrospective review of cTPE performed at North Carolina State University from 2016 to 2018, through a search of the institutional database for cTPE and IMHA. University teaching hospital. Seven dogs with confirmed IMHA were presented to a university teaching hospital ICU for cTPE. Six dogs were not responsive to standard medical management with immunosuppressive agents, while 1 dog presented before immunosuppressive agents were begun. All dogs underwent multiple cTPE procedures using 1 of 2 commercially available apheresis systems. At presentation, the median HCT was 0.15 L/L (15.7%) (range, 0.10-0.19 L/L [10.3%-19%]) and the median total serum bilirubin was 32.5 mmol/L (1.9mg/dl) (range, 15.4-597 mmol/L [0.9-34.9mg/dl]). The median number of transfusions before cTPE was 1 (range, 1-4), with a median total of infused RBCs of 12.9ml/kg (range, 8.8-37ml/kg). cTPE with an exchange of ≥4 times total plasma volumes was used to decrease the level of circulating autoreactive antibodies. The median total plasma volumes exchanged was 4.5 times (range, 2.5-6.5 times) over 2-4 procedures. Anticoagulation was performed using a combination of systemic heparinization and regional citrate in all dogs. Six of 7 dogs (85.7%) were discharged from the hospital and were alive 90 days after discharge. One dog (14%) did not respond to cTPE (∼6.5 times total plasma volume exchanged) and was euthanized. cTPE is a feasible and relatively safe bridging treatment option for the management of canine IMHA.

Retrospective evaluation of systemic hypertension in dogs with nonassociative (primary) immune-mediated hemolytic anemia (December 2016 to April 2019): 26 cases.

To report the prevalence of arterial hypertension in a population of dogs with nonassociative immune-mediated hemolytic anemia (IMHA) on presentation and during hospitalization. To determine the relationships of systolic blood pressure (SBP) with mortality and a prognostic indicator, the canine hemolytic anemia objective score. Retrospective observational study (December 2016 to April 2019). University teaching hospital. Twenty-six clinical dogs presenting to the ICU with nonassociative (primary) IMHA and a control group of 23 clinical dogs with idiopathic epilepsy hospitalized in the ICU for seizure treatment or monitoring. None. Hypertension was defined as SBP≥160mm Hg and severe hypertension as SBP≥180mm Hg. Mean SBP was significantly increased in IMHA dogs (161mm Hg, SD=21) compared to ICU control dogs (138mm Hg, SD=14; P<0.005). Hypertension was present in 13 of 26 (50.0%) dogs across the period of hospitalization and was severe in three of 26 (11.5%). During at least 1 day of hospitalization, 18 of 26 (69.2%) dogs were hypertensive and eight of 26 (34.6%) were severely hypertensive. Hypertension was not associated with short-term mortality or canine hemolytic anemia objective score. In this retrospective study, hypertension was more prevalent in dogs with nonassociative IMHA than a control population of ICU-hospitalized dogs. An association between autoimmune conditions and hypertension has been previously reported in people but not within a canine population. Hypertension in dogs may have an inflammatory or autoimmune etiology. SBP should be monitored closely in canine IMHA, in case antihypertensive treatment is required.

Pharmacokinetics, pharmacodynamics and safety evaluation of 5,5’-methylenebis(2-acetoxybenzoic acid) in dogs following intravenous administration

Complement-mediated intravascular hemolysis occurs in canine immune-mediated hemolytic anemia (IMHA). Complement inhibitors might enhance treatment of this disease. Dimers of acetylsalicylic acid such as 5,5’-methylenebis(2-acetoxybenzoic acid) (DAS) have been reported to inhibit complement. This study aimed to characterize the pharmacokinetics and safety profile of a single 3 mg/kg IV dose of DAS in 6 healthy mixed-breed dogs. Serum concentrations of DAS and its primary metabolites were measured by liquid chromatography-tandem mass spectrometry at baseline and at 5, 10 and 30 min, and 1, 2, 4, 6, 8, 12, 18 and 24 h post-administration. Additional blood samples were collected 7 and 14 days after drug administration. Complete blood counts, serum chemistry panels, C-reactive protein measurements, coagulation testing and cytokine analyses were used for safety monitoring. Following IV administration of 3 mg/kg DAS, the estimated mean maximum plasma concentration was 54,709 ng/mL. Pharmacokinetic modeling suggested that DAS was eliminated with a half-life value of 8.1 h, equivalent to a clearance of 6.93 L/hr kg and a volume of distribution of 56 mL/kg. Plasma concentrations of the metabolites were measured rapidly (within 15−60 min for M1 and M2 respectively). Overall, the relative exposure to M1 and M2 suggest significant biotransformation of DAS occurred, but DAS was the most abundant circulating species. No adverse clinical reactions were noted following DAS administration and safety studies suggested DAS caused no inflammatory response or coagulation disturbance. Further clinical evaluation of DAS is warranted.

Nursing the canine immunemediated haemolytic anaemia patient part 2: supportive care and nursing

Patients with immune-mediated haemolytic anaemia (IMHA) require substantial nursing care considerations, and can be very unwell on presentation. The registered veterinary nurse (RVN) plays an important role, with comprehensive supportive care of vital importance. Patients with IMHA have the potential for many complications, therefore it is important for the RVN to have knowledge of the condition, its treatment options, and how they may impact the patient's needs. Alongside vigilant nursing, and frequent communication with the clinician, the RVN's impact on these patients can be vast, making them extremely rewarding cases to care for.

Canine immune-mediated haemolytic anaemia part 1: presentation, diagnosis and treatment

Immune-mediated haemolytic anaemia (IMHA) is one of the most prevalent immune-mediated diseases in canines, and it carries a guarded prognosis because of the substantial rates of morbidity and mortality. Comprehensive supportive nursing care is critical in patients with IMHA.This first article discusses the presentation, diagnosis and treatment options in canine IMHA patients. A subsequent article will follow focusing on how the condition, and its treatment options, impact the supportive nursing requirements that should therefore be considered, as well as possible complications that may arise. Through good nursing care, as well as a knowledge and understanding of the requirements of these patients, RVNs have a vital role in influencing these patients' outcomes.

Does vaccination increase the risk of immune-mediated haemolytic anaemia in dogs?

Bottom line You admitted Scout, a four-year-old female neutered English springer spaniel, yesterday as she was jaundiced, anaemic and collapsed. Her haematocrit was 13 per cent, with prominent spherocytosis on...

Pharmacokinetics of human recombinant C1-esterase inhibitor and development of anti-drug antibodies in healthy dogs

Complement-mediated intravascular hemolysis occurs in canine immune-mediated hemolytic anemia (IMHA). Complement inhibitors such as recombinant C1 esterase inhibitor (rC1-INH) might prevent this process and alter the disease course. This study aimed to characterize the pharmacokinetics of a single 500 IU IV dose of rC1-INH in 8 healthy beagle dogs, evaluate the dogs for any adverse effects of drug administration, and determine whether rC1-INH administration induces anti-drug antibody formation. Serum rC1-INH concentrations were measured using a commercial functional ELISA at baseline and at 10, 20, 40, 60, 80, 100, 120, 240, 360, 480, 600, 720, 960, and 1440 min post drug administration. Complete blood counts were conducted at baseline, 720 and 1440 min. Western blot analysis, using rC1-INH as the target antigen was used to detect anti-drug antibodies in 14-day serum samples. No adverse clinical reactions were noted following rC1-INH administration. Pharmacokinetic modelling suggested that the peak C1-INH concentration achieved is 0.21 IU/mL and that C1-INH concentration is significantly greater than baseline for 100 min following injection. A robust antibody response was detected which suggests that rC1-INH should not be re-administered after an initial course. Clinical trials of rC1-INH in dogs with intravascular IMHA are now warranted.

How to…

This month Simon Tappin, Editor of Companion, discusses diagnosis and treatment of canine immune-mediated haemolytic anaemia.

Thromboelastographic assessment of the contribution of platelets and clotting proteases to the hypercoagulable state of dogs with immune-mediated hemolytic anemia.

Hypercoagulability is a well-known feature of canine immune-mediated hemolytic anemia (IMHA) and is believed to increase the risk of thrombosis. This study was undertaken to differentiate the relative contribution of platelets and clotting proteases to this hypercoagulability using thromboelastography (TEG). Retrospective observational study. Thromboelastograms from 27 dogs with IMHA were retrospectively evaluated. Standard TEG parameters (R, K, α, MA), the G value, and the novel parameter delta (Δ) were determined. Hypercoagulability was attributed to the platelet component of hemostasis when there was an increased G value with a normal Δ value. Nineteen of 27 dogs (70.4%) had ≥ 2 TEG variables suggestive of hypercoagulability, 18 (66.7%) had a hypercoagulable G value, and 11 (40.7%) had a hypercoagulable Δ value. Ten of 27 (37%) samples met the criteria for platelet hypercoagulability. Our report documents the derivation and application of the Δ value to differentiate enzymatic from platelet hypercoagulability. Further studies are required to validate the use of these TEG variables in this manner. The hypercoagulable tendency in dogs with IMHA is complex and multifactorial, and in some dogs this hypercoagulability may be attributed primarily to platelet hyper reactivity. Our findings may support the use of anti-platelet drugs in some dogs with IMHA.

Comparison of 4 Direct Coombs' Test Methods with Polyclonal Antiglobulins in Anemic and Nonanemic Dogs for In‐Clinic or Laboratory Use

BackgroundDifficulties with the direct antiglobulin test (DAT) and its apparent lack of sensitivity and specificity for immune‐mediated hemolytic anemia (IMHA) in dogs have raised skepticism regarding its diagnostic value.ObjectiveTo compare different DATs and other hematologic parameters in dogs.AnimalsAnticoagulated blood samples from 59 nonanemic and 46 anemic dogs (± IMHA) from a research colony and veterinary clinics.MethodsProspective observational study: Immunochromatographic strip, gel microcolumn, and capillary techniques were compared with standard microtiter DAT using 2 polyvalent antiglobulins. Spherocytosis, autoagglutination, osmotic fragility, and clinical data were assessed.ResultsBlood samples from all 59 nonanemic dogs were DAT‐. Among 46 anemic dogs, 33 were suspected of IMHA, but only 20 were DAT+. Old and new DAT methods yielded comparable and consistent results even after storage of chilled blood samples for 1 week. Spherocytosis and autoagglutination (that did not persist after washing) were noted in 15 and 16 DAT+ dogs, respectively. The other 26 anemic dogs, including 21 previously transfused dogs and 4 with autoagglutination, tested DAT‐ by the other methods. Osmotic fragility was increased in 70% (19/27) of anemic and all 15 DAT+ dogs tested. Limited follow‐up testing revealed DAT+ results for 3–70 days.Conclusions and Clinical ImportanceThe novel strip and capillary DAT methods are promising adjunct in‐clinic tools. Despite prior immunosuppressive treatment and presence of autoagglutination, the DAT was positive in anemic dogs with IMHA. Transfusion did not cause false DAT+ results. Our results support DAT as a cornerstone in the diagnosis of canine IMHA.

The use of the rapid osmotic fragility test as an additional test to diagnose canine immune-mediated haemolytic anaemia

BackgroundDiagnosing canine immune-mediated haemolytic anaemia (IMHA) is often challenging because all currently available tests have their limitations. Dogs with IMHA often have an increased erythrocyte osmotic fragility (OF), a characteristic that is sometimes used in the diagnosis of IMHA. Since the classic osmotic fragility test (COFT) is time-consuming and requires specialized equipment, an easy and less labour-intensive rapid osmotic fragility test (ROFT) has been used in some countries, but its diagnostic value has not yet been investigated.This study aimed to evaluate erythrocyte osmotic fragility in dogs with and without IMHA, to compare results of the classic (COFT) and rapid (ROFT) test and to assess the value of the ROFT as diagnostic test for canine IMHA.Nineteen dogs with IMHA (group 1a), 21 anaemic dogs without IMHA (group 1b), 8 dogs with microcytosis (group 2), 13 hyperlipemic dogs (group 3), 10 dogs with lymphoma (group 4), 8 dogs with an infection (group 5) and 13 healthy dogs (group 6) were included.In all dogs, blood smear examination, in-saline auto-agglutination test, Coombs’ test, COFT and ROFT were performed. In the COFT, OF5, OF50 and OF90 were defined as the NaCl concentrations at which respectively 5, 50 and 90% of erythrocytes were haemolysed.ResultsCompared with healthy dogs, OF5 and OF50 were significantly higher in group 1a (P < 0.001) and OF5 was significantly higher in group 3 (P = 0.0266). The ROFT was positive in 17 dogs with IMHA, 10 hyperlipemic dogs, one anaemic dog without IMHA and one healthy dog.ConclusionsOsmotic fragility was increased in the majority of dogs with IMHA and in dogs with hyperlipidemia, but not in dogs with microcytosis, lymphoma or an infection. Although more detailed information was obtained about the osmotic fragility by using the COFT, the COFT and ROFT gave similar results. The ROFT does not require specialized equipment, is rapid and easy to perform and can be used easily in daily practice. Although, the ROFT cannot replace other diagnostic tests, it may be a valuable additional tool to diagnose canine IMHA.

Lack of evidence of a beneficial effect of azathioprine in dogs treated with prednisolone for idiopathic immune-mediated hemolytic anemia: a retrospective cohort study

BackgroundAzathioprine is used as an immunosuppressant in canine immune-mediated hemolytic anemia (IMHA), but this potentially toxic and carcinogenic drug has not been proven to be beneficial. The aim of this study was to determine the difference in outcome and survival of dogs with idiopathic IMHA treated with a protocol that included azathioprine and prednisolone versus a protocol that included prednisolone alone.ResultsThe study included 222 dogs with a hematocrit lower than 0.30 L/L and either a positive Coombs' test or spherocytosis and no evidence of diseases that could trigger IMHA. The clinical and laboratory data at the time of diagnosis and the response to therapy and survival were compared in dogs treated according to the prednisolone and azathioprine protocol (AP protocol; n = 149) and dogs treated according to the prednisolone protocol (P protocol; n = 73). At study entry, the two groups were comparable, except that thrombocyte counts were significantly lower and clinical signs had been present significantly longer in the AP protocol group. No significant difference in survival was found between the two groups: the 1-year survival was 64% (95% CI 54 - 77%) in the P protocol group and 69% (95% CI 59-80%) in the AP protocol group, respectively.ConclusionsAzathioprine would appear not to be beneficial as standard treatment for all cases of IMHA; however, a blinded, randomized clinical trial is needed to establish whether outcome is different with the two treatment protocols.

Haemoplasma infection is not a common cause of canine immune-mediated haemolytic anaemia in the UK

The aim of this study was to investigate whether the two canine haemoplasma species, Mycoplasma haemocanis and "Candidatus Mycoplasma haematoparvum," are commonly associated with immune-mediated haemolytic anaemia (IMHA) in UK dogs. Three groups of dogs were recruited to the study: anaemic dogs with primary IMHA (n=37); anaemic dogs not meeting the inclusion criteria for primary IMHA (n=77) and non-anaemic dogs (n=113). DNA was extracted from 100 μl of blood and subjected to real-time quantitative polymerase chain reaction (qPCR) assays for both species of Mycoplasma. Each assay incorporated co-amplification of canine glyceraldehyde-3-phosphate dehydrogenase (GAPDH) as an endogenous internal control. Canine GAPDH was successfully amplified by qPCR from all 227 canine blood samples but none contained M. haemocanis or "Candidatus M. haematoparvum" DNA. Haemoplasma infection is uncommon in dogs in the UK and no evidence was found that these organisms act as triggers for IMHA.

Evaluation of platelet activation in canine immune-mediated haemolytic anaemia

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. 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. There was a strong correlation between platelet activation and severe thrombocytopenia, with heightened platelet activation being observed predominantly in severely thrombocytopenic dogs. 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.

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Use of human immunoglobulin in addition to glucocorticoids for the initial treatment of dogs with immune‐mediated hemolytic anemia

To determine the utility of human intravenous immunoglobulin (hIVIG) for the initial treatment of canine immune-mediated hemolytic anemia (IMHA). Blinded, randomized, clinical trial. Veterinary teaching hospital. Twenty-eight, client-owned dogs with primary IMHA. At enrollment, after diagnosis of IMHA, dogs were randomly assigned to receive either hIVIG or placebo, in a blinded fashion. For the next 14 days, all dogs received glucocorticoids as the sole immunosuppressant agent. All dogs received low-molecular-weight heparin as an anticoagulant. D-dimer concentrations were evaluated at the beginning and end of the study protocol to monitor for thromboembolic complications. Twenty-five of 28 dogs (89%) were discharged from the hospital. Thirteen of those received hIVIG and 12 received placebo. Twenty-four dogs (86%) were alive 14 days after enrollment, and of these 13 received hIVIG and 11 received placebo. D-dimer concentrations were elevated in 86% of all dogs at the time of diagnosis. For initial treatment of dogs with IMHA, the addition of hIVIG to corticosteroid treatment did not improve initial response, nor did it shorten hospitalization.