American Journal of Emergency Medicine 31 (2013) 1157.e5-1157.e7

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Case Report

EDTA-dependent pseudothrombocytopenia complicated by eosinophilic pneumonia?,??

Abstract

EDTA-dependent pseudothrombocytopenia (EDTA-PTCP) is a phenomenon that occurs in vitro when EDTA reacts with harvested blood. EDTA-dependent pseudothrombocytopenia usually does not indicate thrombocytopenia in vivo. Here, we report the first case of EDTA-PTCP complicated by eosinophilic pneumonia. A 70-year-old man with rectal cancer was admitted to the hospital for a Liver abscess and rectal cancer. At the time of admission, his platelet count was 20000/uL, but a peripheral blood smear showed platelet aggregation and the platelet count for a kanamycin-added EDTA blood sample was 180000/uL. The patient’s respiratory status worsened after treatment for the liver abscess and rectal cancer. The patient’s bronchoalveolar lavage contained 45% eosinophils, and a diagnosis of acute eosino- philic pneumonia was made. In recent studies, the occurrence of eosinophilic disease has been shown in idiopathic thrombocytopenic purpura. EDTA-dependent pseudothrombocytopenia is an in vitro phenomenon, although platelet activation that results in eosinophil invasion may occur in severe cases.

EDTA-dependent pseudothrombocytopenia (EDTA-PTCP) is a type of thrombocytopenia that occurs in vitro because of a reaction between EDTA and harvested blood, and it does not harm the patient. During blood sampling for complete blood count evaluation, EDTA is often used as an anticoagulant, and an automated blood-cell counter is used to assess the amount of circulating platelets. In some cases, EDTA causes agglutination of platelets. When a blood sample is analyzed using an automated blood cell counter, platelets cannot be accurately counted, and the results do not accurately reflect a patient’s platelet profile. For patients with EDTA-PTCP, citric acid or heparin can be used as an anticoagulant for blood samples to avoid errors in platelet counts, caused by agglutination. However, it should be noted that, in some cases, agglutination can also occur when these agents are used [1-3]. The incidence of EDTA-PTCP is estimated to be 0.09% to 0.11% [4], and this rate increases to 1.9% in patients admitted to the hospital [5]. Thus far, the mechanism underlying the development of EDTA-PTCP is not fully understood [6].

Recent studies reported cases of idiopathic thrombocytopenic purpura (ITP) complicated with eosinophilic disorders [7,8]. Idio-

? Author’s contributions: NO, KN, KT, TF, SN, and YN contributed to patient management. NO, KN, and RI performed study screenings independently. NO, KN, and RI contributed to writing and reviewing the report. Written consent to publish the case report was obtained from the guardians of the patient.

?? Consent: Written informed consent was obtained from the patient for

publication of this case report and any accompanying images. A copy of the written consent is available for review by the Editor-in-Chief of this journal.

pathic thrombocytopenic purpura is similar to EDTA-PTCP in that platelet agglutination occurs in vitro. In ITP, an antigen-antibody reaction results from the expression of antigen on glycoprotein (GP) IIb/IIIa [9,10]. Here, we here report a case of a 70-year-old man who was diagnosed with EDTA-PTCP complicated by eosinophilic pneu- monia. To our knowledge, this is the first report of EDTA-PTCP complicated by eosinophilic pneumonia.

A 70-year-old man was admitted to the emergency unit with a 1- month history of Loss of appetite, fatigue, and drowsiness. He had a history of chronic hepatitis B. His family history was unremarkable, and he had no previous blood transfusions or a history of smoking. At the time of admission, his blood pressure was 98/65 mm Hg, heart rate was 96 beats per minute, core body temperature was 36.9?C, and Glasgow Coma Scale score was E3V5M6. Physical examination showed percussion tenderness of the right upper hypochondriac region. Laboratory blood tests reported an increased inflammatory response, with White blood cell counts of 12000/uL, a C-reactive protein level of

18.3 mg/dL, and slightly increased hepatobiliary enzyme levels, with

glutamate oxalate transaminase level of 45 IU/L and glutamate pyruvate transaminase level of 89 IU/L. He also had an abnormally low platelet count of 20000/uL. Coagulation test results showed a slightly prolonged prothrombin time-international normalized ratio (1.47). Abdominal ultrasonography and contrast computed tomo- graphic (CT) scan showed a large liver abscess as well as evidence of rectal cancer. Treatment for sepsis and disseminated intravascular coagulation as a result of the liver abscess was initiated.

After transfusion of 10 platelet units, the liver abscess was drained. Streptococcus sanguis was identified in the abscess and blood cultures. Thus, the patient was administered antibiotic treatment with piperacillin plus tazobactam at a dose of 4.5 g every 8 hours. Platelet aggregation was identified in the peripheral blood smear performed on day 2, and consequently, the earlier platelet count was re- evaluated. At our hospital, 20 mg/mL kanamycin is added to the blood samples of patients suspected of having EDTA-PTCP because the addition of aminoglycoside antibiotics results in the disaggregation of platelets and prevents thrombocyte agglutination within 30 minutes [11]. In this case, kanamycin was added to the EDTA blood collection tube, and an actual platelet count of 180000/uL was revealed (the platelet count for EDTA alone was 34000/uL) (Table). Based on this result, the patient was diagnosed with EDTA-PTCP. Treatment with antibiotics was ended on day 53, and the patient’s condition clearly improved. On day 68, high anterior resection and transverse colostomy were performed for treatment of rectal cancer (adenocar- cinoma). The patient did not receive intraoperative blood transfusion or antibiotics after surgery, and his postoperative course was good.

On day 83, the patient developed a fever, and on day 86, his respiratory status worsened, and he was ventilated in the intensive

0735-6757/$ - see front matter (C) 2013

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Table

Changes observed in the blood samples after the addition of anticoagulant

			30		60	120
EDTA	WBC (/uL)	12500		12500	12700	12300
	Platelet (/uL)	34000		11000	10000	7000
EDTA plus kanamycin WBC (/uL)		12500		12300	12300	12200
Platelet (/uL)		185000		190000	189000	201000

Anticoagulant Blood tests (/min) 0

Abbreviation: WBC, white blood cell.

care unit. A radiographic image showed bilateral lung infiltrative shadow, particularly in the middle portion of each lung (Fig. 1). Thoracal CT images showed infiltration in the right upper, middle, and dorsal portions of the left upper lung lobes, and pleural fluid could be seen on either side of the lungs (Fig. 2). The patient was then started on antibiotic therapy for nosocomial pneumonia, namely, ceftazidime (1 g every 12 hours) and vancomycin (1 g every 24 hours). In addition, as the patient’s heart function was slightly reduced (ejection fraction, 40%-50%) and dilatation disorder was observed, which suggests acute heart failure, nitrate treatment was given alongside the antibiotic therapy. Nevertheless, the patient’s respiratory status did not improve. On day 91, after high eosinophil counts were observed in the blood smear (30%; reference range, 0%-6%), bronchoalveolar lavage was performed. The eosinophil count in the bronchoalveolar lavage was very high (45%; reference range, 0%-1%), and the patient was diagnosed with acute eosinophilic pneumonia. Hydrocortisone at a dose of 200 mg/d was initiated on the same day. A marked improvement was seen in the respiratory status, and radiographic images showed improvement in signs of infiltrative. On day 94, the patient was extubated. His condition was stable, and he was discharged from the hospital.

In 1969, Gowland et al [6] investigated, for the first time, the occurrence of EDTA-PTCP. However, as yet, the mechanism underly- ing the development of EDTA-PTCP is not fully understood. EDTA- dependent pseudothrombocytopenia is thought to occur when the cryptic platelet antigen on the platelet GP IIb/IIIa is activated by an autoimmune antibody [12,13]. Cryptic platelet antigen is not often expressed, but it can be expressed as a result of the chelating effect of EDTA on Ca²⁺. In addition, EDTA can initiate an antigen-antibody

Fig. 1. Thoracal radiography. Radiographic image show bilateral lung infiltrative, particularly in the middle portion.

reaction, which can result in platelet agglutination [14]. Autoimmune antibodies are known to have no response in Glanzmann disease, a congenital disorder in which the platelet surface GP, the GP IIb/IIIa complex, is absent [12]. In several cases, these antibodies were shown to respond to GP IIb [2], suggesting that the potential antigen is within the GP IIb/IIIa complex. Because of the chelating effect of EDTA, the GP IIb/IIIa complex becomes dissociated, and this can lead to the appearance of a potential antigen. Moreover, factors other than EDTA that result in the expression of cryptic platelet antigens and the maintenance of the autoimmune antibody include sepsis [15]), viral infection [16,17], administration of antibiotics [18], and the platelet aggregation inhibitor abciximab [19] as well as aging [20].

Autoimmune antibodies, which induce EDTA-PTCP, are present in Healthy adults and expressed in high levels in malignant tumors, autoimmune disease, liver disease, and lymph node proliferative disease [4,21]. Thus far, no clear correlation between age or sex and EDTA-PTCP has been reported [4]. In the present case, the patient had sepsis due to a liver abscess. He also had rectal cancer and had previous chronic hepatitis B infection. We believe that this is what caused the expression of cryptic platelet antigens, which resulted in the retention of autoimmune antibodies of EDTA-PTCP. EDTA-dependent pseudo- thrombocytopenia is often misdiagnosed as one of the true in vivo thrombocytopenias such as ITP and disseminated intravascular coagulation, leading to various unnecessary treatments, including platelet transfusion [21], bone marrow puncture [22], steroid treat- ment [23], and splenectomy [13]. Hence, in patients diagnosed with EDTA-PTCP, it is important to identify potential underlying diseases.

To our knowledge, this is the first case of EDTA-PTCP complicated by eosinophilic pneumonia, although ITP complicated with eosino- philic disorders has been reported previously [7,8]. Although the mechanism underlying the development of EDTA-PTCP is not fully understood, it was previously shown that when the release of P- selectin from platelets is activated, the recruitment of eosinophil in the lungs is promoted [9]. In addition, it is thought that platelet stimulation in ITP causes eosinophilic disorder.

EDTA-dependent pseudothrombocytopenia is similar to ITP in that platelet agglutination occurs because of an antigen-antibody reaction that follows the expression of antigen on GP IIb/IIIa [9,10]. EDTA- dependent pseudothrombocytopenia is an in vitro phenomenon, and it is usually not associated with in vivo disorders. Nevertheless, various conditions such as sepsis, malignancy, transfusion, and administration of antibiotics can result in the expression of cryptic platelet antigens. Because platelet agglutination can also occur in citric acid blood samples [4], a high risk of antigen-antibody reaction can be expected in patients who receive blood transfusions that contain citric acid and antibiotics with chelating effects. Although this is not enough to cause

Fig. 2. Computed tomographic image. Thoracal CT image shows infiltrative in the right upper and middle lung lobes and the dorsal portion of the left upper lung lobe. Bilateral accumulation of pleural fluid can be seen.

N. Ohashi et al. / American Journal of Emergency Medicine 31 (2013) 1157.e5-1157.e7 1157.e7

agglutination, platelet activation induced by antigen-antibody reac- tion can trigger eosinophilic disorder. Therefore, the risk of eosino- philic disorder might be high in cases of EDTA-PTCP.

This case presents 2 findings. First, in patients diagnosed with EDTA-PTCP, it is important to identify potential underlying diseases. Second, EDTA-PTCP is an in vitro phenomenon, but it may develop into an eosinophilic disorder, similar to ITP.

Acknowledgments

We would like to acknowledge Makiko Hirahata and Shigemi Kobayashi for their assistance.

Naoko Ohashi MD Kensuke Nakamura MD Ryota Inokuchi MD

Department of Emergency and Critical Care Medicine The University of Tokyo Hospital, Bunkyo-ku, Tokyo 113-8655, Japan

E-mail address: [email protected]

Hajime Sato, MD, PhD

Department of Health Policy and Technology Assessment National Institute of Public Health, Wako, Saitama 351-0197, Japan

Kurato Tokunaga MD Tatsuma Fukuda MD Susumu Nakajima MD, PhD Naoki Yahagi MD, PhD

Department of Emergency and Critical Care Medicine The University of Tokyo Hospital, Bunkyo-ku, Tokyo 113-8655, Japan

http://dx.doi.org/10.1016/j.ajem.2013.03.033

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