Heparin Induced Platelet Aggregation Presentation and Surgical Consequences

Heparin induced thrombocytopenia and its vascular complications were identified in 25 patients. Aggregation studies performed using ADP and/or heparin in these patients were compared with responses in 18 normal. individuals. The specific aggregating factor was demonstrated in plasma. Other findings of note were normal fibrinogen, minimal elevation of fibrin split products, a normal or slightly prolonged prothrombin time and severe thrombocytopenia. Arterial and venous thrombosis required surgery in seven patients, four of whom required major amputation. Although an antibody has not been identified, the time of development of this phenomenon suggests an immune response. Only those patients who had previous exposure to heparin developed this phenomenon in less than six days. Thrombotic complications occurred with either bovine lung or porcine gut preparations from different manufacturers and appeared following both subcutaneous and intravenous administration. Electron microscopy of clots removed from four patients revealed characteristic platelet fibrin thrombi. Early recognition, prompt discontinuance of heparin and the use of platelet antiaggregants will prevent disastrous consequences. All patients undergoing heparin therapy should be carefully observed for thrombocytopenia with specific platelet aggregation studies when appropriate.

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The Clinical Usefulness of the Platelet Aggregation Test for the Diagnosis of Heparin-Induced Thrombocytopenia

Thrombosis and Haemostasis ◽

10.1055/s-0038-1651610 ◽

1993 ◽

Vol 69 (04) ◽

pp. 344-350 ◽

Cited By ~ 175

Author(s):

B H Chong ◽

J Burgess ◽

F Ismail

Keyword(s):

Platelet Aggregation ◽

Heparin Therapy ◽

Serotonin Release ◽

Point System ◽

Control Groups ◽

Heparin Induced Thrombocytopenia ◽

Heparin Concentration ◽

Release Test ◽

The Mean ◽

Normal Controls

SummaryThe platelet aggregation test is widely used for the diagnosis of heparin-induced thrombocytopenia (HIT), a potentially serious complication of heparin therapy. We have evaluated its sensitivity and specificity in comparison with those of the 14C-serotonin release test. The sensitivity of the platelet aggregation test was found to vary with the heparin concentration and the donor of the platelets used in the test. The optimal heparin concentrations were between 0.1 and 1.0 U/ml. Using these heparin concentrations, the mean sensitivity varied from 39% (with the least reactive platelets) to 81% (with the most reactive platelets). In comparison, the sensitivity of the release test ranged from 65% to 94%. The specificities of the platelet aggregation test were 82%, 90% and 100% for the following control groups: (1) non-thrombocytopenic patients given heparin, (2) patients with thrombocytopenia due to other causes, and (3) normal controls not given heparin, respectively. The corresponding specificities for the release test was 94%, 90% and 100%. The specificities can be further increased to 100% for all controls with the adoption of a two-point system which defines a positive result as one in which platelet aggregation occurs with a low heparin concentration (0.5 U/ml) but not with 100 U heparin/ml. For optimal results, a two-point platelet aggregation test should be performed with heparin concentrations of 0.5 and 100 U/ml and using platelets of more reactive donors.

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Heparin-induced thrombocytopenia: when a low platelet count is a mandate for anticoagulation

Hematology ◽

10.1182/asheducation-2009.1.225 ◽

2009 ◽

Vol 2009 (1) ◽

pp. 225-232 ◽

Cited By ~ 14

Author(s):

Thomas L. Ortel

Keyword(s):

Platelet Count ◽

Heparin Therapy ◽

Severe Thrombocytopenia ◽

Drug Induced ◽

Platelet Factor ◽

Heparin Induced Thrombocytopenia ◽

Immune Mediated ◽

Number Of Patients ◽

Increased Risk ◽

Antibody Levels

Abstract Heparin-induced thrombocytopenia (HIT) is an immune-mediated disorder caused by the development of antibodies to platelet factor 4 (PF4) and heparin. The thrombocytopenia is typically moderate, with a median platelet count nadir of ~50 to 60 × 109 platelets/L. Severe thrombocytopenia has been described in patients with HIT, and in these patients antibody levels are high and severe clinical outcomes have been reported (eg, disseminated intravascular coagulation with microvascular thrombosis). The timing of the thrombocytopenia in relation to the initiation of heparin therapy is critically important, with the platelet count beginning to drop within 5 to 10 days of starting heparin. A more rapid drop in the platelet count can occur in patients who have been recently exposed to heparin (within the preceding 3 months), due to preformed anti-heparin/PF4 antibodies. A delayed form of HIT has also been described that develops within days or weeks after the heparin has been discontinued. In contrast to other drug-induced thrombocytopenias, HIT is characterized by an increased risk for thromboembolic complications, primarily venous thromboembolism. Heparin and all heparin-containing products should be discontinued and an alternative, non-heparin anticoagulant initiated. Alternative agents that have been used effectively in patients with HIT include lepirudin, argatroban, bivalirudin, and danaparoid, although the last agent is not available in North America. Fondaparinux has been used in a small number of patients with HIT and generally appears to be safe. Warfarin therapy should not be initiated until the platelet count has recovered and the patient is systemically anticoagulated, and vitamin K should be administered to patients receiving warfarin at the time of diagnosis of HIT.

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Sera from patients with heparin-induced thrombocytopenia generate platelet-derived microparticles with procoagulant activity: an explanation for the thrombotic complications of heparin-induced thrombocytopenia

Blood ◽

10.1182/blood.v84.11.3691.bloodjournal84113691 ◽

1994 ◽

Vol 84 (11) ◽

pp. 3691-3699 ◽

Cited By ~ 336

Author(s):

TE Warkentin ◽

CP Hayward ◽

LK Boshkov ◽

AV Santos ◽

JA Sheppard ◽

...

Keyword(s):

Flow Cytometry ◽

Procoagulant Activity ◽

Severe Thrombocytopenia ◽

Drug Reactions ◽

Heparin Induced Thrombocytopenia ◽

Circulating Microparticles ◽

Thrombotic Complications ◽

Drug Dependent

Heparin-induced thrombocytopenia is characterized by moderate thrombocytopenia and thrombotic complications, whereas quinine/quinidine-induced thrombocytopenia usually presents with severe thrombocytopenia and bleeding. Using flow cytometry and assays of procoagulant activity, we investigated whether sera from patients with these immune drug reactions could stimulate normal platelets to generate platelet-derived microparticles with procoagulant activity. Sera or purified IgG from patients with heparin-induced thrombocytopenia stimulated the formation of platelet-derived microparticles in a heparin-dependent fashion. Further studies showed that heparin-induced thrombocytopenia sera also produced a marked increase in procoagulant activity. In contrast, sera from patients with quinine- or quinidine-induced thrombocytopenia did not generate platelet-derived microparticles nor generate increased procoagulant activity. However, quinine/quinidine-induced thrombocytopenia sera produced a significant increase in the binding of IgG to platelets in a drug-dependent fashion, whereas sera from patients with heparin-induced thrombocytopenia demonstrated no drug-dependent binding of IgG to platelets. We also observed increased levels of circulating microparticles in patients with acute heparin-induced thrombocytopenia compared with control patients. Our observations indicate that the generation of procoagulant platelet-derived microparticles in vivo is a plausible explanation for the thrombotic complications observed in some patients with heparin-induced thrombocytopenia.

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Early Management of Heparin-Induced Thrombocytopenia Using a Simple Scoring System

Blood ◽

10.1182/blood.v118.21.4679.4679 ◽

2011 ◽

Vol 118 (21) ◽

pp. 4679-4679

Author(s):

Harry L. Messmore ◽

Nancy Fabbrini ◽

Mary L. Bird ◽

Abdul M. Choudhury ◽

Miguel Cerejo ◽

...

Keyword(s):

Decision Making ◽

Platelet Aggregation ◽

Clinical Management ◽

Scoring System ◽

Laboratory Tests ◽

Heparin Therapy ◽

Test Results ◽

Heparin Induced Thrombocytopenia ◽

Clinical Scoring ◽

Lab Test

Abstract Abstract 4679 Since stopping heparin therapy lessens the probability of severe complications of heparin-induced thrombocytopenia (HIT) such as arterial and venous thromboembolism, it is important to stop heparin as soon as the diagnosis is suspected. As HIT is a complex disorder and its diagnosis is difficult, the use of a clinical scoring system would facilitate clinical decision making. One of the foremost issues in these patients is whether to discontinue heparin (to avoid HIT complications) or not (to avoid thrombotic complications). This study was performed to develop a simple scoring system to aid in the early clinical management of suspected HIT patients with regard to decisions for continued heparin therapy. The system was designed to arrive at low (0) or possible (1) probability scores based on clinical criteria without knowledge of lab test results, except platelet counts, to avoid delays. As the safest clinical approach is to discontinue heparin, intermediate and high scores were combined. Historically, laboratory tests such as the heparin-induced platelet aggregation test (PA) and 14C-serotonin release assay (SRA) and nonfunctional tests such as the enzyme-linked-immunosorbent assay (ELISA) have been used for the diagnosis of HIT. These test results are frequently available only after 12 to 36 hours which, for some patients, is a risk due to continued heparin therapy while waiting for test results. One solution to this problem is to have a clinical scoring system to guide the clinician before test results are available. This study enrolled 100 critically ill VA hospitalized patients with a >=30% fall in platelet count. Assessment of platelet aggregation (PA), 14C-SRA and GTI ELISA was also performed. In this population 53% were scored 1 and of these 43% were positive by lab testing for HIT. Emphasizing the decision to discontinue heparin, the clinical signs of HIT were paramount for the immediate determination of a diagnosis of HIT. Specifically, this study demonstrated the value of a simple clinical score to aid in the clinical management decision making to continue or discontinue heparin in patients suspected of having HIT, without dependence on a positive HIT lab test. An algorithm is provided. This scoring system does not preclude reassessment of patients in terms of continuing heparin if subsequent laboratory tests and/or thrombosis become positive. Disclosures: No relevant conflicts of interest to declare.

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Adenosine Diphosphate (ADP) and ADP Receptor Play a Major Role in Platelet Activation/Aggregation Induced by Sera From Heparin-Induced Thrombocytopenia Patients

Blood ◽

10.1182/blood.v91.2.549.549_549_554 ◽

1998 ◽

Vol 91 (2) ◽

pp. 549-554 ◽

Cited By ~ 1

Author(s):

János Polgár ◽

Petra Eichler ◽

Andreas Greinacher ◽

Kenneth J. Clemetson

Keyword(s):

Platelet Aggregation ◽

Platelet Activation ◽

Atp Release ◽

Adenosine Diphosphate ◽

Heparin Therapy ◽

Dense Granule ◽

Heparin Induced Thrombocytopenia ◽

Adp Receptor ◽

Induced Aggregation ◽

Granule Release

The molecular basis for heparin-induced thrombocytopenia (HIT), a relatively common complication of heparin therapy, is not yet fully understood. We found that pretreatment of platelets with AR-C66096 (formerly FPL 66096), a specific platelet adenosine diphosphate (ADP) receptor antagonist, at a concentration of 100 to 200 nmol/L that blocked ADP-dependent platelet aggregation, resulted in complete loss of platelet aggregation responses to HIT sera. AR-C66096 also totally inhibited HIT serum-induced dense granule release, as judged by measurement of adenosine triphosphate (ATP) release. Apyrase, added to platelets at a concentration that had only minor effects on thrombin- or arachidonic acid-induced aggregation, also blocked completely HIT serum-induced platelet aggregation. Furthermore, AR-C66096 inhibited platelet aggregation and ATP release induced by cross-linking FcγRIIA with specific antibodies. These data show that released ADP and the platelet ADP receptor play a pivotal role in HIT serum-induced platelet activation/aggregation. The thromboxane receptor inhibitor, Daltroban, had no effect on HIT serum-induced platelet activation whereas GPIIb-IIIa antagonists blocked platelet aggregation but had only a moderate effect on HIT serum-induced dense granule release. Pretreatment of platelets with chondroitinases but not with heparinases resulted in concentration dependent inhibition of HIT serum-induced platelet aggregation. These novel data relating to the mechanism of platelet activation induced by HIT sera suggest that the possibility should be examined that ADP receptor antagonists or compounds that inhibit ADP release may be effective as therapeutic agents for the prevention or treatment of complications associated with heparin therapy.

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Heparin-Induced Thrombocytopenia and Thrombosis: Therapeutic Strategy Using a Single-Chain Variable Fragment (scFv) Antibody

Blood ◽

10.1182/blood.v120.21.3346.3346 ◽

2012 ◽

Vol 120 (21) ◽

pp. 3346-3346

Author(s):

Jaa Yien New ◽

Jose Perdomo ◽

Xing-Mai Jiang ◽

Beng Chong

Keyword(s):

Monoclonal Antibody ◽

Platelet Aggregation ◽

Heparin Therapy ◽

Human Platelets ◽

Binding Activity ◽

Single Chain Variable Fragment ◽

Single Chain ◽

Heparin Induced Thrombocytopenia ◽

E Coli

Abstract Abstract 3346 Introduction and Aim Heparin-Induced Thrombocytopenia and Thrombosis (HIT) is a life threatening disorder that affects 1–5% of patients receiving heparin therapy. A low platelet count is usually recorded (<150,000 per cubic millimetre) with a decrease of 50% or more from the baseline. The occurrence of HIT is due to the presence of an IgG antibody that recognizes the immune complex formed between Platelet Factor 4 (PF4) and heparin. The antibody/PF4/Heparin complex binds to the FcγRIIa receptor on platelets, leading to platelet activation and thrombotic complications in patients receiving heparin. IV.3 is a murine monoclonal antibody that was raised against the FcγRIIa receptor and has been used as an inhibitor in specificity assays to confirm HIT in patients. We have developed a humanized single-chain variable fragment (scFv) antibody based on the IV.3 monoclonal antibody that binds to the FcγRIIa receptor on platelets and prevents platelet aggregation induced by HIT antibodies. Methods The variable heavy chain (VH) and light chain (VL) of the IV.3 antigen binding fragment (Fab) moiety were amplified using polymerase chain reaction (PCR). These two fragments were then coupled with a linker (Glycine4 and Serine)6. This was followed by introduction of several components including fusion tags (FLAG and c-Myc) at both termini for cloning, detection and purification purposes. The construct was transformed into E. coli (strain-BL21) for protein expression of the scFv. The presence of the protein was detected via immunostaining using anti-FLAG and anti-c-Myc antibodies. The scFv was purified by affinity chromatography and the binding activity was detected using flow cytometry and confocal microscopy. The functional activity was determined using Platelet Aggregation Assay. The scFv was then humanized to minimize potential immunogenicity. Humanization was achieved by introducing specific mutations that rendered the molecule human-like but did not affect binding specificity. The humanized scFv was also expressed in E. coli, purified and tested as before. Results The scFv protein (32kDa) was expressed, purified and confirmed via immunostaining. The created humanized scFv exhibits binding activity against the FcγRIIa on human platelets as determined by flow cytometry and confocal microscopy. In addition, the protein successfully inhibits platelet aggregation at micro molar concentrations in aggregation assays conducted in vitro in the presence of HIT antibodies. Conclusions The humanized scFv was successful in recapitulating the properties of the IV.3 murine monoclonal antibody. It demonstrated binding activity against the FcgRIIa on human platelets and exhibited functional activity by inhibiting platelet activation and aggregation in vitro. This implies that our scFv is able to stop binding of the antibody/PF4/Heparin immune complex to platelets, thus hindering one of the critical initial steps in HIT. The scFv described here may be able to ameliorate the unwanted side effects of heparin therapy and could serve as a potential therapeutic drug for HIT patients. Disclosures: No relevant conflicts of interest to declare.

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Recidivne tromboze arterij spodnjega uda pri bolniku s trombocitopenijo, povzročeno s heparinom – Prikaz primera

Slovenian Medical Journal ◽

10.6016/zdravvestn.1864 ◽

2017 ◽

Vol 86 ◽

Author(s):

Blanka Mahne ◽

Mladen Gasparini ◽

Matija Kozak

Keyword(s):

Platelet Count ◽

Venous Thrombosis ◽

Lower Limb ◽

Early Recognition ◽

Coronary Bypass ◽

Posterior Tibial Artery ◽

Heparin Therapy ◽

Limb Amputation ◽

Heparin Induced Thrombocytopenia ◽

Acute Thrombosis

Background: Heparin-induced thrombocytopenia (HIT) is an immune-mediated disorder following heparin therapy presenting with thrombocytopenia and associated arterial or/and venous thrombosis (heparin induced thrombocytopenia with thrombosis–HITT). Unrecognised HIT can lead to severe complications like limb amputation and death.Case report: We report a case of a patient who presented with HIT-associated recurrent lower limb arterial thrombotic occlusions and popliteal venous thrombosis 29 days after coronary bypass graf surgery. The patient underwent urgent thrombectomy of superfcial femoral, popliteal and posterior tibial artery. Because of recurrent thrombotic occlusions of lower limb arteries three surgical revisions were performed. Te platelet count decreased from 124 × 109/l to 53 × 109/l on the fifth day after the first intervention. After clinical suspicion of HIT, heparin was discontinued and fondaparinux was started. Arterial thrombosis did not recur and the patient recovered without consequences.Conclusions: HIT occurs in 1–3 % of patients after cardiac surgery. Strict following of international guidelines regarding the frequency of platelet count monitoring, assessing probability for HIT and laboratory testing is mandatory in order not to miss the diagnosis of HIT. HIT can manifest clinically several days after the first exposure to heparin. If a patient presents with acute thrombosis and thrombocytopenia, HITT should be suspected. Postoperative HIT is associated with higher morbidity and mortality. Early recognition is crucial to prevent severe complications and death.

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Adenosine Diphosphate (ADP) and ADP Receptor Play a Major Role in Platelet Activation/Aggregation Induced by Sera From Heparin-Induced Thrombocytopenia Patients

Blood ◽

10.1182/blood.v91.2.549 ◽

1998 ◽

Vol 91 (2) ◽

pp. 549-554 ◽

Cited By ~ 68

Author(s):

János Polgár ◽

Petra Eichler ◽

Andreas Greinacher ◽

Kenneth J. Clemetson

Keyword(s):

Platelet Aggregation ◽

Platelet Activation ◽

Atp Release ◽

Adenosine Diphosphate ◽

Heparin Therapy ◽

Dense Granule ◽

Heparin Induced Thrombocytopenia ◽

Adp Receptor ◽

Induced Aggregation ◽

Granule Release

AbstractThe molecular basis for heparin-induced thrombocytopenia (HIT), a relatively common complication of heparin therapy, is not yet fully understood. We found that pretreatment of platelets with AR-C66096 (formerly FPL 66096), a specific platelet adenosine diphosphate (ADP) receptor antagonist, at a concentration of 100 to 200 nmol/L that blocked ADP-dependent platelet aggregation, resulted in complete loss of platelet aggregation responses to HIT sera. AR-C66096 also totally inhibited HIT serum-induced dense granule release, as judged by measurement of adenosine triphosphate (ATP) release. Apyrase, added to platelets at a concentration that had only minor effects on thrombin- or arachidonic acid-induced aggregation, also blocked completely HIT serum-induced platelet aggregation. Furthermore, AR-C66096 inhibited platelet aggregation and ATP release induced by cross-linking FcγRIIA with specific antibodies. These data show that released ADP and the platelet ADP receptor play a pivotal role in HIT serum-induced platelet activation/aggregation. The thromboxane receptor inhibitor, Daltroban, had no effect on HIT serum-induced platelet activation whereas GPIIb-IIIa antagonists blocked platelet aggregation but had only a moderate effect on HIT serum-induced dense granule release. Pretreatment of platelets with chondroitinases but not with heparinases resulted in concentration dependent inhibition of HIT serum-induced platelet aggregation. These novel data relating to the mechanism of platelet activation induced by HIT sera suggest that the possibility should be examined that ADP receptor antagonists or compounds that inhibit ADP release may be effective as therapeutic agents for the prevention or treatment of complications associated with heparin therapy.

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Lepirudin treatment in a boy with suspected HIT II after surgery because of tetralogy of Fallot

Hämostaseologie ◽

10.1055/s-0037-1617021 ◽

2009 ◽

Vol 29 (02) ◽

pp. 168-170

Author(s):

K. Barth ◽

L. Nakamura ◽

U. Budde ◽

R. Arnold ◽

B. Zieger ◽

...

Keyword(s):

Tetralogy Of Fallot ◽

Heparin Therapy ◽

Severe Thrombocytopenia ◽

Repeated Testing ◽

Heparin Induced Thrombocytopenia ◽

Life Threatening ◽

Therapeutic Measures ◽

First Time ◽

Factor Antigen ◽

Heparin Exposure

SummaryHeparin-induced thrombocytopenia (HIT II) in childhood is rare. Suspected HIT II requires immediate diagnostic and therapeutic measures in order to avoid potentially life threatening complications. Heparin must be stopped immediately.We report on a 6-year old boy who required cardiac surgery due to tetralogy of Fallot. To our knowledge he had been exposed to heparin for the first time during cardiac catheterization on the day before surgery. Preoperatively, platelet count was normal. Postoperatively (3 days after heparin exposure), he developed pulmonary and renal failure and required inotropic cardiac support and dia -lysis. He also developed progressive (severe) thrombocytopenia under heparin therapy on day 2–3 postoperatively. The dialysis filter required daily exchanges due to clotting despite increasing heparin doses. The first ELISA for HIT on postop day 4 was negative. 3 days later a repeated test was positive. Von Wille-brand factor antigen and D-dimers were markedly increased. The patient was immediately switched to lepirudin and subsequently stabilized slowly. No major systemic thrombosis occurred. After lepirudin treatment for 6 weeks the patient was fully recovered and HIT II-testing was negative again. Conclusion: In children with progressive thrombocytopenia in the setting of heparin exposure and signs of major or micro thrombosis HIT II must be ruled out. Even if a first early test turns out negative repeated testing should be performed. Lepirudin anticoagulation is effective and should be monitored correctly. Platelet transfusion should be avoided in HIT II.

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EFFECTS OF THROMBOXANE PATHWAY ANTAGONISTS ON HEPARIN-INDUCED PLATELET AGGREGATION (H-IPA) AND TESTING FOR HEPARIN-INDUCED THROMBOCYTOPENIA (HIT)

10.1055/s-0038-1643383 ◽

1987 ◽

Author(s):

L D Brace ◽

J Fareed ◽

D Hoppensteadt

Keyword(s):

Platelet Aggregation ◽

Normal Donor ◽

Severe Thrombocytopenia ◽

Induce Platelet Aggregation ◽

Heparin Induced Thrombocytopenia ◽

Thromboxane Receptor ◽

Heparin Fractions ◽

Acid Metabolites ◽

Thromboxane Production ◽

Drug Free

We have reported that heparin and heparin fractions can induce platelet aggregation (PA) in a substantial number of normal healthy drug-free donors. H-IPA was shown to depend upon the molecular weight and concentration of the heparin preparation used, but its mechanism remains unknown. Therefore, we performed experiments with antagonists of the thromboxane pathway to determine whether arachidonic acid metabolites contribute to H-IPA. When indomethacin or 13-azaprostanoic (a thromboxane receptor anatagonist) was added to the PRP of donors whose platelets had been shown to aggregate in response to heparin, H-IPA was completely inhibited: heparin (bovine or porcine) caused 75% PA, while pretreatment with indomethacin or 13-APA reduced the response to 6%. Similarly, if the same donors ingested 650 mg aspirin 3 hours prior to phlebotomy, the PA response to heparin was reduced to approx. 10%. These results demonstrate that at least part of the mechanism of H-IPA is mediated through thromboxane generation. However, the mechanism by which'heparin stimulates thromboxane production in platelets remains unknown.In some patients, heparin is known to induce an immune response that causes severe thrombocytopenia (HIT) and is associated with arterial and venous thrombosis. Fratantoni, et al. (Blood 45:395-401, 1975) have introduced a PA method for the diagnosis of HIT. We have used a modification of this method to show that the PA observed when heparin is added to a mixture of normal donor PRP and HIT patient’s serum or plasma can be inhibited by antagonists of the thromboxane pathway. When normal donor PRP was pretreated with indomethacin or 13-APA and then mixed with serum from a HIT patient (290 uL PRP:160 uL serum), the PA response to heparin was reduced from 75% to 10% or less. Similarly, if the PRP donors ingested 650 mg aspirin prior to phlebotomy, PA in the HIT test was reduced from 75% to 10% or less. Thus, the interaction of heparin with the antibody and platelets causes thromboxane generation and leads to PA. Cyclo-oxygenase specific antiplatelet drugs and inhibitors of thromboxane generation may be useful in the clinical management of HIT and H-IPA.

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