Hereditary thrombocytopenia with platelet sialic acid deficiency and mutations in the GNE genes.

BACKGROUND: The inherited macrothrombocytopenias are rare disorders and the underlying cause can be identified in many cases but in some, this can remain enigmatic. Platelet transfusions are often administered during hemorrhagic events. METHODS: A patient with previously unexplained inherited macrothrombocytopenia with a platelet count between 3-20 × 109 /L is described in which studies were performed using exome sequencing (ES) and platelet flow cytometry. RESULTS: Both the hemoglobin and white cell counts were normal. ES revealed two suspicious variants, one likely pathogenic and one a variant of uncertain significance, in the UDP-N-acetylglucosamine 2-epimerase/N-acetylmannosamine kinase (GNE) gene, and flow cytometry showed diminished expression of surface platelet sialic acid (about 5%) but normal red cell sialic acid. The Thrombopoietin (TPO) level was low, and the patient responded to TPO-mimetic treatment with an increase in the platelet count. CONCLUSION: Two variants in the GNE gene were able to be upgraded to pathogenic with apparently restricted expression to the megakaryocyte lineage. Platelet transfusion may be avoided in these patients with TPO-mimetic treatment.

Discordance in tests used to detect inhibition of the P2Y12 receptor in patients undergoing interventional neuroradiology procedures.

BACKGROUND: Clopidogrel is an inhibitor of the P2Y12 platelet receptor but testing to demonstrate a drug effect is controversial since there are often discordant results between different tests methods. METHODS: Samples from patients taking clopidogrel prior to intracranial flow-diversion procedures were tested using light transmission aggregometry (LTA), whole blood impedance aggregometry (WBIA) and the VerifyNow device (VND). Samples were classified as concordant if all test results were either responsive (inhibition) or resistant. Discordant results were separated using the VND into those with a responsive versus a resistant test result. RESULTS: Samples from 96 patients were studied. Concordance for all three tests was seen in 53/96 (55%) of samples, of which 41 (43%) were responsive and 12 (12%) were resistant. Discordance was observed in 43 samples (45%), 37 (28%) of which were caused by responsive VND and either a resistant WBIA or LTA and 6 (7%) of which were caused by a resistant VND but a responsive test result using either WBIA or LTA. These two discordant groups differed in both platelet count and hematocrit, but no such difference was present between the two concordant groups. CONCLUSION: Discordance in P2Y12 inhibition testing may be partly explained by sample platelet count and hematocrit.

In vitro comparison of CPD whole blood with conventional blood components.

INTRODUCTION: Resuscitation of severely injured trauma patients is commonly performed using red blood cells in additive solution supplemented with plasma and platelet concentrates. There is an increasing interest in the use of low anti-A titer Group O whole blood (LTOWB) in the early management of the resuscitation. It is unclear whether clinical outcome is improved using this approach. METHODS: Expired units of CPD-LTOWB were studied on Day 22 and expired units of thawed plasma on Day 6 and Day 7. LTOWB was assessed for hemoglobin content, clotting factor levels and platelet numbers and function using thromboelastography (TEG) and impedance aggregation. Assays of fibrinogen and FV, FVIII, FVII and FX were performed on the expired plasma. The LTOWB hemoglobin was compared to red cells in additive solution (AS-RBCs) and the clotting factor levels to those of expired thawed plasma. Platelet function was compared to fresh whole blood samples from healthy subjects. RESULTS: LTOWB contained slightly more hemoglobin than the AS-RBCs (Medians, 66 v 59 G), and the plasma content of fibrinogen was similar. Other clotting factors were reduced by approximately 15% except for FVIII which was 30% less. Both TEG and impedance aggregometry showed evidence of residual platelet function despite the prolonged period of refrigerator storage. CONCLUSION: LTOWB contains higher hemoglobin and adequate clotting factors, and residual platelet function is demonstrated indicating that this product would be expected to be at least equivalent to a single unit of each of the conventional components commonly used in trauma resuscitation.

Remote Dispensing of Emergency Release Red Blood Cells.

OBJECTIVES: Patients with acute bleeding are frequently transfused with emergency release (ER) group O RBCs. This practice has been reported to be safe with a low rate of acute hemolytic transfusion reactions (AHRs). METHODS: Records of patients who received ER RBCs over a 30-month period were examined at our hospitals. During this period, satellite refrigerators were on site in the emergency department (ED), which were electronically connected to the blood bank (electronically connected satellite refrigerator [ECSR]). Nurses accessing the refrigerator were required to give patient identification information, when known, prior to removal of the ER RBCs, allowing technologists the opportunity to check for previous serologic records and communicate directly with the ED if a serologic incompatibility was potentially present. RESULTS: In total, 935 patients were transfused with 1,847 units of ER RBCs. Thirty of these patients had a current (22/30) or historic (8/30) antibody. In 15 cases, incompatible RBCs were interdicted. In six cases, the transfusion was considered urgent, and an AHR occurred in four of these six (overall 0.4%), including one fatal AHR due to anti-KEL1. CONCLUSIONS: Use of KEL1-negative RBCs and ECSR merits consideration as approaches to mitigate the occurrence of ER RBC-associated AHRs.

Influenza-associated thrombotic thrombocytopenic purpura: A report of two cases and a brief review of the literature.

BACKGROUND AND OBJECTIVES: Thrombotic thrombocytopenic purpura (TTP) is often preceded by a recent history of an acute infection and influenza is the most implicated virus. MATERIALS AND METHODS: We identified two cases of TTP, which were preceded by influenza between 2010 and 2021. In one patient, we epitope mapped the binding specificity of antibodies using an overlapping peptide approach of the stalk protein of Influenza B and the cysteine-rich spacer domain (CRSD) of ADAMTS13. A literature search was performed for reports of influenza-associated TTP over the period 1980-2021. RESULTS: Two patients were identified in which TTP was preceded by influenza, one Influenza A and the other Influenza B. Epitope mapping of the latter's plasma identified target epitopes in both the stalk protein of Influenza B and CRSD of ADAMTS13. The literature review revealed only seven case reports, all but one from Europe or Asia and associated with Influenza A. Severe ADAMTS13 deficiency was demonstrated in only four cases. CONCLUSION: We report the first small case series of influenza-associated TTP. Moreover, it is the first case implicating Influenza B and a mechanism favouring polyclonal B-cell proliferation rather than molecular mimicry as the stimulus to form anti-ADAMTS13 auto-antibodies is suggested.

Laboratory Methods in the Assessment of Hereditary Hemostatic Disorders.

In patients presenting with a suspect hereditary bleeding disorder a detailed bleeding history is first obtained. Testing proceeds in a tiered manner with platelet count, platelet morphology, platelet histogram, PFA-100, fibrinogen, prothrombin time, and activated partial thromboplastin time. More detailed testing includes von Willebrand factor, individual clotting factor assays, and platelet function testing. Next, testing for a dysfibrinogenemia, FXIII, or a fibrinolytic defect is considered. Hemostatic abnormality is not demonstrated in a fraction of patients. An approach to management in these patients, such as desmopressin or antifibrinolytic therapy, may be required and empiric use of blood component therapy is discouraged.

Red Cell Exchange as Adjunctive Therapy for Babesiosis: Is it Really Effective?

Human babesiosis is a parasitic disease prevalent in the Northeastern and Midwestern United States (US). Treatment with antibiotics is the standard of care but red cell exchange (RCE) has been used as an adjunctive treatment in more severe disease. Data for the efficacy of RCE in the treatment of babesiosis has been based on case reports and case series. An English language literature search was conducted for cases of babesiosis treated with RCE since 1980 and relevant laboratory and clinical outcome data were extracted. Similar data were obtained on severe cases of babesiosis referred for RCE in our hospitals in the time period 2000 to 2020. Fifty reports including forty-one individual case reports and nine case series were retrieved. There were 108 patients that underwent RCE with an overall mortality rate of 20%. Some patients had more than one RCE. The patients varied in the level of anemia and evidence of compromise of renal or pulmonary function. The pre-RCE level of parasitemia varied between 1.7% to 85% with the vast majority >10%. The post-RCE level of parasitemia varied between 1% to 10%. Since 2000, 32 patients were referred for RCE in our hospitals and RCE was performed on 23 of 32. There were more patients treated with RCE in the second decade as compared to the first decade, 19 versus 4 respectively. The overall mortality was 22% similar to the national data. Comparing the cohort treated with RCE to the 9 patients who were treated only with antibiotics, there were similar levels of parasitemia and laboratory parameters. The overall number of days needed to achieve a parasite count <1% was similar between the two cohorts and mortality for the antibiotics only cohort was 0%. More than 40 years after the first reported case of RCE in severe babesiosis it cannot be concluded that this adjunctive therapy favorably influences the clinical outcome. Since there is largely equipoise, a registry of severe patients treated with or without RCE could identify a benefit or otherwise.

Causes of an elevated international normalized ratio in the intensive care unit and the implications for plasma transfusion.

BACKGROUND: The presence of an elevated international normalized ratio (INR) is common in patients in the intensive care unit (ICU), but the cause rarely determined. These patients are at risk to receive prophylactic plasma prior to invasive procedures. STUDY DESIGN AND METHODS: Samples from patients with an INR of 1.5 or greater were frozen and subsequently thawed and assayed for procoagulant and anticoagulant clotting factors and anti-Xa to determine the likely cause of the INR. Samples showing a low FVII, FX, PC, and PS were categorized as a vitamin K deficiency pattern. Samples showing a low FV, low or normal fibrinogen, and high FVIII were categorized as a liver disease pattern. Samples showing an anti-Xa >0.01 IU/ml were assayed for anti-Xa DOACs. Samples which could not be categorized were grouped as equivocal. RESULTS: A total of 48 samples were obtained over a 6-month period. Nineteen showed a Vitamin K deficiency pattern, 17 a liver disease pattern, 7 showed an anti-Xa DOAC and 5 were equivocal. High FVIII and D-dimers and reduced levels of the anticoagulant proteins were present in the majority of the samples. FVII levels correlated inversely with the INR (r = -0. 81), as did FX (r = -0.67) but not FV (r = -0.04) nor fibrinogen (r = -0.15). CONCLUSION: Transfusion of plasma to reverse an elevated INR in the ICU should be discouraged since such a practice is either avoidable by the use of vitamin K or inappropriate in the case of liver disease or an anti-Xa DOAC.

Heavy parasitemia in babesiosis treated without adjunctive red cell exchange.

Babesiosis is a tick-borne infectious disease caused by the protozoa Babesia but transplacental, and transfusion transmission may occur. While most infections are asymptomatic, rarely, it can present with a severe, life-threatening illness. Treatment is primarily with antibiotics, but red cell exchange (RCE) has been used in more severe cases which are characterized by high-grade parasitemia, evidence of severe hemolysis and or multi-organ failure. A threshold parasite level of 10% has arbitrarily been applied as an indication for RCE; however, this threshold is not evidence-based. We report on three cases of severe babesiosis in which we considered the use of RCE on the basis of a parasite level greater than 10%, but the procedure was not performed. We deferred RCE on account of the good clinical state of the patient and the absence of end-organ failure. All patients were followed daily until discharge. Two of these patients had been splenectomized, and each received a single unit of red blood cells during the hospitalization. The third patient had a long history of refractory lymphoma and was pancytopenic requiring multiple transfusions during the years before the diagnosis of babesiosis. She had transfusion-transmitted babesiosis from a red blood cell transfused 46 days prior to diagnosis. All three patients responded well to antibiotics, and none expired. This small case series suggests that requests for RCE solely on the basis of an arbitrary level of parasitemia should be questioned and the clinical state and evidence of end-organ failure considered in the decision to perform RCE.

Adjunctive treatment of clinically severe babesiosis with red blood cell exchange: a case series of nineteen patients.

BACKGROUND: Infection with the protozoan parasite Babesia, the causative agent of babesiosis, can result in asymptomatic to life-threatening illness. Severe cases of babesiosis are characterized by high levels of parasitemia (>4%-10%) and commonly treated with adjunctive red blood cell exchange (RCE) in addition to antimicrobial therapy. The efficacy of RCE in this context is unknown. STUDY DESIGN AND METHODS: Blood bank records were examined for requests for RCE during a 10-year period from 2007 to 2017. Relevant clinical and laboratory variables were extracted from medical records from presentation to 35 days after RCE and analyzed in univariate and multivariate models. RESULTS: Nineteen cases of babesiosis were identified in which RCE was performed. The median age of patients was 77 years, 74% of whom were male. A total of 37% of patients were asplenic. RCE was performed on average 1.3 days after presentation, with procedural urgency driven mainly by the level of parasitemia. Mean pre- and post-RCE levels of parasitemia were 12.9 and 3.4%, respectively, resulting in a mean percent reduction in parasitemia of 75%. Preprocedural parasitemia (p = 0.047) and age (p = 0.028) were both significant predictors of postprocedural hospital length of stay (post-RCE LOS). Neither postprocedural parasitemia (p = 0.12) nor percent reduction in parasitemia (p = 0.72) correlated with post-RCE LOS. Four patients died, none of whom were asplenic. Mortality was not correlated with hematologic, parasitologic, or clinical variables analyzed. CONCLUSIONS: Reduction in the level of parasitemia is the only known benefit of RCE in severe babesiosis.

Refractoriness to red blood cell transfusion therapy due to hypersplenism.

BACKGROUND: A 55-year-old male presented with myelodysplastic/myeloproliferative neoplasm and severe splenomegaly. The patient is blood group O, D+ with a negative indirect antiglobulin test. Transfusion of 5 units of red blood cells (RBCs) increased the hemoglobin (Hb) level from 6.7 to 7.2 g/dL. No active bleeding or hemolysis was evident. The patient was readmitted 1 week later with a Hb level of 3.3 g/dL. An additional 6 units of RBCs showed only an increase from 3.3 to 3.5 g/dL. Partial splenic embolization was performed, which resulted in a stabilization of the Hb level at approximately 7 g/dL. Because of this, total splenectomy was performed, which resulted in a gradual increase in Hb level to approximately 13 g/dL. The patient remains transfusion independent 160 days postsplenectomy. RESULTS: RBC transfusion increases Hb concentration by 1 g/dL per unit in a typical adult. This increase is attenuated in the presence of ongoing hemolysis or active blood loss. Occasionally, a low-RBC-volume unit transfused to a recipient with a large intravascular blood volume may show an unexpectedly small increase. In rare situations, however, the etiology of a greatly attenuated response is more perplexing. The pattern of Hb concentration posttransfusion was suggestive of splenic sequestration in our patient. CONCLUSION: Severe refractoriness to RBC transfusion attributable to severe hypersplenism is a rare event. Our case suggests that splenic artery embolization may be a useful initial approach in individual cases and a potential predictor of the utility of a subsequent surgical splenectomy.

False-negative compatible antiglobulin crossmatches in samples with alloantibodies to cognate red blood cell antigens.

BACKGROUND: Patient samples showing a positive indirect antiglobulin test are further tested to identify alloantibody specificity using a panel of phenotypically characterized group O reagent red blood cells (RBCs). Donor RBCs phenotypically negative for the antibody specificity are then serologically crossmatched using an antiglobulin reagent. This latter test is performed to identify any incompatibility due to the presence of undetected minor blood group antibodies and considered an important step in patient safety. STUDY DESIGN AND METHODS: Samples with well-characterized alloantibodies were intentionally crossmatched against donor RBCs expressing the cognate antigen. In a separate set of specimens, the alloantibody was titered and crossmatched against both heterozygous and homozygous cells. RESULTS: Thirty-five samples containing 10 common alloantibodies crossmatched against 240 ABO-compatible donor cells phenotypically positive for the cognate antigen showed compatible crossmatches in 89 of 240 (37%). Antibody titering of 12 alloantibodies showed that a titer of 2 or more was required for incompatibility of all homozygous cells and a titer of 8 or more for incompatibility of all heterozygous cells. CONCLUSION: The antiglobulin crossmatch has a high failure rate (false-negatives) related to antibody titer and donor cell zygosity and is not reliable in interdicting incompatibility due to minor blood group antibodies.

Coagulation assays in a case of apixaban overdose.

: Intentional overdose of apixaban is rare and minimal data exist regarding the usefulness of routinely available laboratory tests to predict drug levels. A 50-year-old man was admitted after ingestion of 200-mg apixaban. Serial blood samples were obtained over a 54-h period for assessment of the fall-off in drug levels using the prothrombin time/international normalized ratio and anti-Xa assays and compared with an apixaban-specific chromogenic assay. The prothrombin time/international normalized ratio and anti-Xa assays correlated with the apixaban level when the drug was in the supratherapeutic range (>130 ng/ml) but not in the typical therapeutic trough to peak levels. Apixaban levels are best assessed by a specific anti-Xa test using optimized chromogenic substrates and specific calibrators. A standard anti-Xa test can be a useful surrogate when drug levels are high but use of a specific threshold level for discharge purposes requires caution.

Hemolysis in In-Date RBC Concentrates.

OBJECTIVES: Hemolysis is one of the most prominent changes that occur during the liquid storage of RBCs in additive solution (AS), but most studies have measured hemolysis only on day 42. METHODS: Prestorage leukoreduced RBCs in AS-1 and AS-3 were studied, one group on day 42 and a second group between day 0 and day 40. Each product was sampled for direct measurement of supernatant hemoglobin and hematocrit. RESULTS: Ninety day 42 and 218 day 7 to day 39 RBCs showed a mean ± SD supernatant hemoglobin of 75 ± 100 vs 25.5 ± 16 mg/dL respectively (P < .01). Supernatant hemoglobin correlated weakly with storage age (r = 0.2, P < .01) but more strongly with hematocrit (r = 0.4, P < .01). CONCLUSIONS: There are minimal differences in supernatant hemoglobin until the final days of liquid storage when some high hematocrit RBCs show excessive hemolysis.