A67-year-old man who had undergoneuncomplicated bioprosthetic replacementof the aortic valve and coronary artery

bypass grafting developed transient postoperativethrombocytopenia, followed by a second decline inplatelet count. He was clinically stable and did nothave new symptoms or postoperative complica-tions. There was no evidence of sepsis, thrombosisor hemorrhage. The patient had received unfrac-tionated heparin (70 000 IU in total) with prota-mine reversal (600 mg), one unit of pooledplatelets and one unit of packed red blood cellsduring the operation. Enoxaparin (30 mg twicedaily) had been given postoperatively forantithrombotic prophylaxis.

The patient’s platelet count fell to 43 × 109/Lby the second postoperative day and then gradu-ally rose to 85 × 109/L by the fifth day. His plateletcount began to decline for a second time on day 6after surgery and reached 47 × 109/L on day 7,which prompted a consultation with the hematol-ogy service (Box 1). A complete blood countshowed that the patient had mild, stable, postoper-ative anemia (hemoglobin level 82 g/L and meancorpuscular volume 95 fL) and a normal leuko-cyte count (6.2 × 109/L) with a normal differential.A peripheral blood film confirmed thrombocy-topenia, with no morphologic abnormalities in theerythrocytes or leukocytes. Prothrombin and acti-vated partial thromboplastin times were withinnormal limits. The patient’s lactate dehydrogenaselevel was 207 (normal 125–220) U/L.

What is the most likely diagnosisat the seventh postoperative day?

a. Dilutional thrombocytopeniab. Heparin-induced thrombocytopeniac. Post-transfusion purpura d. Thrombotic thrombocytopenic purpura

A tentative diagnosis of heparin-induced thrombo-cytopenia, an immune-mediated thrombocytopenia

resulting in platelet activation and hypercoagula-bility, was made (b). Dilutional thrombocytopeniacould explain the initial decline in platelet count,which reached its nadir on the second postopera-tive day, and the subsequent recovery of theplatelet count to 85 × 109/L. The nadir of postoper-ative thrombocytopenia can occur anytimebetween days 1 and 4 after surgery (most com-monly occuring on day 2). Once the platelet countbegins to rise, the ultimate peak count occurs onabout day 14, with a gradual restoration to preop-erative levels over the following one to two weeks.1

In contrast, new-onset thrombocytopenia on orafter postoperative day 5, which was observed inour patient, is not consistent with a diagnosis ofdilutional thrombocytopenia.

Post-transfusion purpura is a rare disorder thatoccurs almost exclusively in women with a remotehistory of pregnancy or transfusion; less than 5% ofcases are seen in men who have had previous bloodtransfusion.2 Post- transfusion purpura presents withsevere thrombocytopenia (platelet count < 10 ×109/L) and mucocutaneous bleeding that occurs 5–10 days after transfusion with a platelet-containingproduct (usually packed red blood cells).2

What is your call?

Progressive thrombocytopenia after cardiac surgeryin a 67-year-old man

Ilana Kopolovic MD, Theodore E. Warkentin MD

Competing interests:Theodore Warkentin hasreceived a grant from theHeart and StrokeFoundation of Ontario. Hehas received lecturehonoraria from CanyonPharmaceuticals,Instrumentation Laboratoryand Pfizer Canada; and hehas received consulting feesfrom ParinGenix and W.L.Gore, as well as fees formedical–legal testimony. Hehas received royalties fromTaylor & Francis. Nocompeting interests weredeclared by IlanaKopolovic.

This article has been peerreviewed.

Correspondence to:Ilana Kopolovic,kopolovi@ualberta.ca

CMAJ 2014. DOI:10.1503/cmaj.131449

PracticeCMAJ

© 2014 Canadian Medical Association or its licensors CMAJ, September 2, 2014, 186(12) 929

Box 1: Sequence of platelet counts in a67-year-old man who underwent cardiacsurgery*

• Preoperative: 158

• Intraoperative: 71

• Postoperative

- Immediate: 103†

- Day 1: 86

- Day 2: 43

- Day 3: 61

- Day 4: 78

- Day 5: 85

- Day 6: 73

- Day 7: 47

*All platelet counts are × 109/L (normal range 150–400).†Patient received an intraoperative platelet transfusion.

CM

E

Thrombotic thrombocytopenic purpura canoccur postoperatively, in particular followingcardiac surgery,3 but it is rare. The absence ofschistocytes on the blood film and a normal lac-tate dehydrogenase level ruled out this diagnosisin our patient.

A presumptive diagnosis of heparin-inducedthrombocytopenia was made, and a therapeuticdose of fondaparinux (7.5 mg/d) was startedbased on expert opinion.4 A polyspecificenzyme-linked immunosorbent assay (ELISA)for antiplatelet factor 4/heparin antibodies wasstrongly positive (optical density 2.5 [normal< 0.40]). Bilateral Doppler and compressionultrasound scans of the legs were obtained torule out subclinical thrombosis,5 because its pres-ence would influence the required duration ofanticoagulation. The ultrasound images showedno evidence of deep vein thrombosis, and therewere no clinical signs or symptoms of othervenous or arterial thromboembolism.

Despite the change to treatment with fonda-parinux, the patient’s platelet count continued todecline over the next four days, from 47 × 109/Lon postoperative day 7 to 13 × 109/L on day 11.All other parameters of the complete blood countand peripheral blood film were unchanged. Pro-thrombin and activated thromboplastin timesremained within normal limits, and the fibrino-gen level was elevated (4.5 [normal 1.5–3.5] g/L). The hematology service was asked toreevaluate therapy.

What is the most appropriateintervention for this patient?

a. Stop treatment with fondaparinux and starthigh-dose intravenous immunoglobulintreatment

b. Stop treatment with fondaparinux and starttreatment with a direct thrombin inhibitor

c. Continue treatment with fondaparinuxd. Stop treatment with fondaparinux without start-

ing treatment with a different anticoagulant

Fondaparinux treatment was continued (c) for apresumptive diagnosis of heparin-inducedthrombocytopenia, based on the presence of anti-bodies characteristic of the “delayed-onset” type.High-dose intravenous immunoglobulin treat-ment would be an appropriate treatment for post-transfusion purpura. A direct thrombin inhibitor,such as argatroban or bivalirudin, would be anappropriate treatment for heparin-induced throm-bocytopenia, in keeping with recent evidence-based guidelines;6 however, titration of the doseof these agents using the laboratory coagulation

parameters can be confounded by concomitantdisseminated intravascular coagulation,7 a com-plication occurring in up to 25% of patients withdelayed-onset heparin-induced thrombocytope-nia.8,9 A case series of patients with well- documented heparin-induced thrombocytopeniareported that fondaparinux was a safe and effec-tive treatment for this condition.10 The occur-rence or perpetuation of heparin-induced throm-bocytopenia in patients taking fondaparinux israre.11 Most importantly, the usual progression ofthe delayed-onset variant of heparin-inducedthrombocytopenia is characterized by progres-sive thrombocytopenia that reaches its nadirbetween days 10 and 17 after surgery.8,9

Given that our patient was otherwise doingwell, without clinical evidence of thrombosis orovert disseminated intravascular coagulation, thedecision was made to continue treatment withfondaparinux. Discontinuation of fondaparinuxwithout starting treatment with a different anti-coagulant would put the patient at risk of venousor arterial thromboembolism, which occurs inover 50% of patients with heparin-inducedthrombocytopenia if adequate anticoagulation isnot provided.12 Current guidelines and an expertreview recommend a minimum of four weeks oftherapeutic treatment with a non-heparin anti -coagulant in patients who have heparin-inducedthrombocytopenia without thrombosis.5,6

Which result of a serotonin releaseassay would confirm thediagnosis?

a. Serotonin release at a dose of 0.1 IU/mLheparin, but not at a dose of 0 or 100 IU/mLheparin

b. Serotonin release at doses of 0 and 0.1 IU/mLheparin, but not at a dose of 100 IU/mL heparin

c. No serotonin release at any concentration ofheparin

d. Serotonin release only with doses of fonda-parinux

The serotonin release assay is a diagnostic testfor heparin-induced thrombocytopenia that mea-sures the ability of the antiplatelet factor 4/heparin immunoglobulin G (IgG) antibodies toactivate platelets and induce serotonin release ina sample of serum. In our patient, the results ofthe assay confirmed strong reactivity (b) both inthe presence of heparin at a therapeutic dose(day 11 serum sample: peak 92% serotoninrelease at 0.1 IU/mL unfractionated heparin) andin the absence of heparin (peak 81% serotoninrelease at 0 IU/mL unfractionated heparin)

Practice

930 CMAJ, September 2, 2014, 186(12)

(Appendix 1, available at www.cmaj.ca/lookup/suppl /doi:10.1503 /cmaj.131449/-/DC1). Plateletactivation was inhibited at a high concentrationof heparin (0% serotonin release at a dose of100 IU/mL unfractionated heparin). The strongserum-induced activation of platelets observed inthe absence of heparin (buffer control) is animportant serologic feature of delayed-onsetheparin-induced thrombocytopenia.9

Serotonin release at a dose of 0.1 IU/mLheparin, but not a dose of 0 or 100 IU/mLheparin, would be characteristic of most patientswith heparin-induced thrombocytopenia, inwhom strong reactivity in the serotonin releaseassay is observed only when the patient is receiv-ing therapeutic concentrations of heparin. Noserotonin release at any concentration of heparinargues against a diagnosis of heparin-inducedthrombocytopenia.

Serotonin release only with doses of fonda-parinux would implicate fondaparinux as a causalagent of the thrombocytopenia and would suggestthat this agent was provoking platelet activationand an associated prothrombotic state. There wasno increase in reactivity over the buffer control atany dose of fondaparinux tested; this includedtests of patient serum diluted 1:16 and 1:64, inwhich less than 5% reactivity was observed at adose of 0 IU/mL heparin and at all concentrationsof fondaparinux, but strong heparin-dependentactivation of platelets (> 80% serotonin release)remained (Appendix 1). Because the addition offondaparinux did not increase the degree of sero-tonin release compared with that observed withthe buffer control, fondaparinux cross-reactivitywas ruled out.

The patient continued to receive therapeuticanticoagulation (which was changed to treatmentwith rivaroxaban upon discharge because ofpatient preference). The gradual elevation of hisplatelet count corresponded to an observed pro-gressive decline in serum-induced activation ofplatelets in the absence of heparin.

Discussion

Heparin-induced thrombocytopenia is a well-characterized, prothrombotic disorder caused byplatelet-activating IgG antibodies that recognizemultimolecular complexes of platelet factor 4(a cationic protein) bound to anionic heparinthrough charge–charge interactions.8 Plateletactivation occurs when the platelet factor 4/heparin/IgG complexes form in situ on the sur-face of platelets, cross-linking the platelet FcγIIareceptors, thereby producing strong platelet acti-vation in association with a marked procoagu-lant response.

Clinically, heparin-induced thrombocytopeniatypically presents 5–10 days after an immunizingexposure to heparin, which is usually heparingiven intraoperatively (e.g., during cardiac or vas-cular surgery) or soon after surgery (e.g., as usualpostoperative thromboprophylaxis).12 The com-mon occurrence of heparin-induced thrombocy-topenia postoperatively is believed to result fromthe combination of surgery (with the release ofplatelet factor 4 from activated platelets) and theadministration of heparin during or after surgery,triggering the formation of the immunogenicplatelet factor 4/heparin complexes.13

In our patient, the initial picture of a seconddecline in the platelet count that began on thefifth postoperative day and while the patient wasreceiving low-molecular-weight heparin wasconsistent with heparin-induced thrombocytope-nia. However, as the case evolved, several pointsseemed to argue against the diagnosis, in particu-lar, the progressive decline of the patient’splatelet count for nearly a week following thecessation of heparin treatment. One possibleexplanation for this was concomitant dissemi-nated intravascular coagulation, a well-describedcomplication of heparin-induced thrombocytope-nia.8,9 However, disseminated intravascular coag-ulation is unlikely to progress if the patient isreceiving effective anticoagulation; our patientwas receiving fondaparinux at a therapeuticdose. In addition, stable fibrinogen levels andcoagulation parameters (activated partial throm-boplastin time and prothrombin time/interna-tional normalized ratio) within the normal rangesuggested that there was no progression to dis-seminated intravascular coagulation.

It is commonly thought that severe thrombo-cytopenia (platelet count < 20 × 109/L) arguesagainst a diagnosis of heparin-induced thrombo-cytopenia; however, it occurs in 5%–10% ofpatients with heparin-induced thrombocytopeniaand in about 50% of patients with delayed-onsetheparin-induced thrombocytopenia.9 Suchpatients are at increased risk of disseminatedintravascular coagulation and microthrombosisassociated with heparin-induced thrombocytope-nia. The lowest recorded nadir for a plateletcount in a patient with serologically confirmedheparin-induced thrombocytopenia is 2 × 109/L.8

Delayed-onset heparin-inducedthrombocytopeniaDelayed-onset heparin-induced thrombocytopeniawas initially reported in a case series of 12patients in whom heparin-induced thrombocy-topenia began five or more days after their lastexposure to heparin.9 Recently, the term “delayed-onset heparin-induced thrombocytopenia” also

Practice

CMAJ, September 2, 2014, 186(12) 931

has been used to describe heparin-induced throm-bocytopenia that worsens after heparin treatmentends,8 which occurred in our case. Our patient’splatelet count recovered relatively slowly; this isconsistent with the finding that the normalizationof the platelet count may follow a protractedcourse in patients with delayed-onset heparin-induced thrombocytopenia and associated strongreactivity in the absence of heparin in the sero-tonin release assay.9 An evaluation on day 44 aftersurgery showed that the serum of our patientremained strongly positive in the serotonin releaseassay at therapeutic heparin doses (Appendix 1).In addition, whereas the reactivity of the serotoninrelease assay in the absence of heparin declinedover time (with a corresponding gradual increasein the patient’s platelet count to 61 × 109/L), therewas still some residual reactivity (10% serotoninrelease), which explains the persisting degree ofthrombocytopenia in the patient. At day 88 aftersurgery, the platelet count was 111 × 109/L, andthere was essentially no reactivity in the serotoninrelease assay in the absence of heparin (3% sero-tonin release).

Diagnostic testing for heparin-inducedthrombocytopeniaThe need to make a timely diagnosis and startappropriate therapy requires an awareness ofhow heparin-induced thrombocytopenia can pre-sent and progress in a patient, because it oftentakes several days to confirm the diagnosis withlaboratory tests. However, these tests are criticaland can inform the clinician’s understanding andcharacterization of the disease process.

Enzyme-linked immunosorbent assay isundertaken initially and identifies the presence ofhigh levels of antiplatelet factor 4 /heparin anti-bodies in patient serum; its high sensitivity (about99%)14,15 allows heparin-induced thrombocy-topenia to be ruled out if the result is negative,whereas the strength of a positive result can pre-dict the likelihood of true heparin-induced throm-bocytopenia. As in our patient, higher opticaldensity levels (> 2.00) have been strongly associ-ated with the presence of heparin-dependent,platelet-activating IgG antibodies, which areimplicated in heparin-induced thrombocytopenia:up to 90% of patients with optical density levelsgreater than 2.0 in the ELISA will have a positiveresult in the serotonin release assay.16,17 When theoptical density is not considered, the specificityof a positive ELISA result is limited and has beenestimated to be as low as 26%.18

As described earlier, the presence ofantiplatelet factor 4/heparin antibodies in apatient’s serum is not sufficient for a diagnosis ofheparin-induced thrombocytopenia. In fact, the

serum of most patients with such antibodies willnot be able to activate platelets in a serotoninrelease assay, which indicates that these antibod-ies are not pathogenic. Conversely, in most truecases of heparin-induced thrombocytopenia, theserotonin release assay will demonstrate the abil-ity of the patient’s serum to induce platelet acti-vation only at therapeutic concentrations ofheparin, with minor or no platelet activation inthe absence of heparin, and an inhibition ofplatelet activation at very high concentrations ofheparin.9 Currently, the serotonin release assay isperformed in only one laboratory in Canada(McMaster Platelet Immunology Laboratory,Hamilton, Ont.).

The serotonin release assay provides threeimportant contributions to our case. First, the test ismore specific for heparin-induced thrombocytope-nia than the ELISA and therefore strongly suggestsa diagnosis of heparin-induced thrombocytopenia,despite our patient’s atypical clinical features. Sec-ond, the strong, serum-induced platelet activationat a concentration of 0 IU/mL heparin is character-istic of delayed-onset heparin-induced thrombocy-topenia, which helps to explain both the dramaticprogression of thrombocytopenia in the absence offurther treatment with heparin and the subsequentslow recovery of the platelet count in our patient.The sera of patients with delayed-onset heparin-induced thrombocytopenia, when compared withthe sera of patients with usual heparin-inducedthrombocytopenia, have strong serotonin release(≥ 50%, and often > 80%) both in the presence andin the absence of therapeutic concentrations ofheparin. Serotonin release will be inhibited at adose of 100 IU/mL heparin in all sera of patientswith heparin-induced thrombocytopenia.9 Finally,the serotonin release assay ruled out fondaparinuxcross-reactivity as the cause of the progressivedecline in the platelet count in our patient while hereceived this drug.

References1. Greinacher A Warkentin TE. Acquired non-immune thrombocy-

topenia. In: Marder VJ, Aird, WC, Bennett JS, et al., editors.Hemostasis and thrombosis: basic principles and clinical prac-tice. 6th ed. Philadelphia (PA): Lippincott Williams & Wilkins;2013: 796-804.

2. Taaning E, Svejgaard A. Post-transfusion purpura: a survey of12 Danish cases with special reference to immunoglobulin Gsubclasses of the platelet antibodies. Transfus Med 1994;4:1-8.

3. Naqvi TA, Baumann MA, Chang JC. Post-operative thromboticthrombocytopenic purpura: a review. Int J Clin Pract 2004; 58:169-72.

4. Warkentin TE. How I diagnose and manage HIT. HematologyAm Soc Hematol Educ Program. 2011;2011:143-9.

5. Lee GM, Arepally GM. Heparin-induced thrombocytopenia.Hematology Am Soc Hematol Educ Program. 2013;2013:668-74.

6. Linkins LA, Dans AL, Moores LK, et al. Treatment and preven-tion of heparin-induced thrombocytopenia: antithrombotic ther-apy and prevention of thrombosis, 9th ed: American College ofChest Physicians Evidence-Based Clinical Practice Guidelines.Chest. 2012; 141(2 Suppl): e495S-530S.

7. Warkentin TE. Anticoagulant failure in coagulopathic patients:

Practice

932 CMAJ, September 2, 2014, 186(12)

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PTT confounding and other pitfalls. Expert Opin Drug Saf2014;13:25-43.

8. Warkentin TE. Agents for the treatment of heparin-inducedthrombocytopenia. Hematol Oncol Clin North Am 2010; 24:755-75.

9. Warkentin TE, Kelton JG. Delayed-onset heparin-inducedthrombocytopenia and thrombosis. Ann Intern Med2001;135:502-6.

10. Warkentin TE, Pai M, Sheppard JI, et al. Fondaparinux treat-ment of acute heparin-induced thrombocytopenia confirmed bythe serotonin-release assay: a 30-month, 16-patient case series.J Thromb Haemost 2011;9:2389-96.

11. Warkentin TE. Fondaparinux: Does it cause HIT? Can it treatHIT? Expert Rev Hematol 2010;3:567-81.

12. Warkentin TE, Kelton JG. A 14-year study of heparin-inducedthrombocytopenia. Am J Med 1996;101:502-7.

13. Warkentin TE. HIT paradigms and paradoxes. J ThrombHaemost 2011;9(Suppl 1):105-17.

14. Pouplard C, Amiral J, Borg JY, et al. Decision analysis for use ofplatelet aggregation test, carbon 14-serotonin release assay, andheparin-platelet factor 4 enzyme-linked immunosorbent assayfor diagnosis of heparin-induced thrombocytopenia. Am J ClinPathol 1999;111:700-6.

15. Bakchoul T, Giptner A, Najaoui A, et al. Prospective evaluationof PF4/heparin immunoassays for the diagnosis of heparin-induced thrombocytopenia. J Thromb Haemost 2009;7:1260-5.

16. Warkentin TE, Sheppard JI, Moore JC, et al. Quantitative inter-pretation of optical density measurements using PF4-dependentenzyme-immunoassays. J Thromb Haemost 2008;6:1304-12.

17. Pearson MA, Nadeau C, Blais N. Correlation of ELISA opticaldensity with clinical diagnosis of heparin-induced thrombocy-

topenia: a retrospective study of 104 patients with positive anti-PF4/heparin ELISA. Clin Appl Thromb Hemost 2013 Nov. 27

18. Cuker A, Cines DB. How I treat heparin-induced thrombocy-topenia. Blood 2012;119:2209-18.

Affiliations: Department of Medicine (Kopolovic), Univer-sity of Toronto, Toronto, Ont.; Pathology and MolecularMedicine (Warkentin), Hamilton Regional Laboratory Medi-cine Program, McMaster University, Hamilton, Ont.

Contributors: Ilana Kopolovic drafted the manuscript, andTheodore Warkentin reviewed its intellectual content. Both ofthe authors approved the final version of the manuscript sub-mitted for publication.

CMAJ, September 2, 2014, 186(12) 933

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