Molecular Mechanisms of Heparin-Induced Platelet Dysfunction

Introduction

Hemostasis involves many components, all of which must function properly in order for the process to be efficient. Maintaining adequate hemostasis is extremely important for patients undergoing cardiothoracic surgery, due to the continuing problem of postoperative bleeding. The event of excessive postoperative bleeding for heart surgery patients varies from 4% to 32%, depending on the institutional definition of excessive.1 Cardiothoracic surgical programs account for 25% of the demand put on United States blood banks each year.1 Platelets play an essential role in the coagulation process and maintenance of hemostasis. Second only to a break in vascular integrity, a decrease in platelet count and function are the major causes of postoperative bleeding.1

Cardiopulmonary bypass (CPB) is a key contributor to platelet dysfunction in cardiac surgery.1,2 Utilization of heparin as an anticoagulant is an important and necessary component during the conduct of CPB. Although most anticoagulation regimens employ heparin primarily, heparin has been linked to several detrimental outcomes on platelet count and function.2-7 Several causative agents and mechanisms occur to facilitate platelet dysfunction at the level of the receptor. A complete discussion of all the molecular and biological aspects of platelet receptors is beyond the scope of this review. However, the goal of this discussion is to present the basic physiological aspects of certain platelet adhesion receptors, glycoprotein IIb/IIIa (GP IIb/IIIa), von Willebrand factor (GP Ib-IX-V), and P-Selectin, and describe the role of these receptors in heparin-induced platelet dysfunction.

GP IIb/IIIa Receptor

The GP IIb/IIIa receptor is the more abundant receptor complex on the surface of the platelet.8 The GP IIb/IIIa receptor complex is an integral membrane protein with a molecular weight of 236,000 Kda.8 The GP IIb/IIIa receptor, also known as the fibrinogen receptor, can bind a variety of soluble adhesive proteins such as; fibrinogen, vWf, fibronectin, thrombospondin, and vitronectin.9,10 Platelet activation is necessary for the GP IIb/IIIa receptor to bind these aforementioned ligands. GP IIb/IIIa activation is calcium dependent11 and is initiated via G-protein- dependent inositol triphosphate (IP3) hydrolysis, which increases the intracellular concentration of calcium. As intracellular calcium increases, protein kinase C is activated, which is associated with platelet aggregation, secretion and release of arachidonic acid metabolites, and facilitation of fibrinogen-binding.12 As fibrinogen is the common ligand for the GP IIb/IIIa receptor, its binding results in platelet activation and instigation of the platelet-platelet interaction.

The pathologic effects of CPB on the GP IIb/IIIa receptor are well documented. However, the GP IIb/IIIa receptor has been implicated in heparin-induced platelet dysfunction.13 In this study, Kozek-Langenecker and coworkers suggest heparin and protamine diminish the response from fibrinogen binding resulting in concomitant decreases in platelet granule secretion and aggregation. Other research has suggested minimal platelet dysfunction occurs as a result of the heparin-induced GP IIb/IIIa receptor disruption. So, the role of the GP IIb/IIIa receptor in heparin-induced platelet dysfunction remains a topic for further study.

The GP IIb/IIIa receptor is the target of several pharmacologic therapies. Two such pharmaceuticals are available; GP IIb/IIIa receptor antagonists and monoclonal antibodies against the GP IIb/IIIa receptor. GP IIb/IIIa receptor antagonists are a new class of potent drugs that profoundly inhibit platelet function by blocking the key receptor involved in platelet aggregation. GP IIb/IIIa receptor antagonists, such as AggrastatÒ occupy the active site of the receptor obstructing ligand entry, therefore inhibiting platelet activation. Monoclonal antibodies to the GP IIb/IIIa receptor, AbciximabÒ have been developed and demonstrate tremendous clinical efficacy. The antibodies adhere directly to the receptor preventing ligand binding and subsequent platelet activation.

GP Ib-IX-V Receptor

von Willebrand factor receptor (GP Ib-IX-V) is the second most abundant receptor on the surface of the platelet.14 The GP Ib-IX-V receptor complex is an integral membrane protein with a molecular weight of approximately 200,000 Kda.15 The binding of vWf to the GP Ib-IX-V receptor results in arachidonic acid metabolism, activation of protein kinase C, increased intracellular calcium concentrations, cytoskeletal reorganization, and functional changes to other adhesion receptors.15 In addition to vWf , thrombin is also capable of binding the GP Ib-IX-V receptor,15 however activation following thrombin binding is quite different with respect to its downstream effects.

Heparin-induced platelet dysfunction via the GP Ib-IX-V receptor occurs directly at the level of the receptor. Heparin disrupts the GP Ib-IX-V receptor by blocking vWf binding in the active site, the GP Ib subunit.3 The inability of vWf to bind to platelet receptor GP Ib results in the prevention of platelet adhesion to the subendothelium of the vessel, and therefore, bleeding is prolonged.3

Currently, there are no pharmaceutical agents that directly target the GP Ib-IX-V receptor to antagonize platelet function.

P-Selectins

The P-Selectin receptor is an integral membrane protein with a molecular weight of 140,000 Kda.16 P-Selectin is a member of the selectin family of vascular cell surface receptors. The selectins can be found on the surface of the endothelium, leukocytes, and platelets. P-Selectins have only been localized on the surface of the platelet and endothelial cell.15 P-Selectins are stored in the Weibel-Palade bodies of the endothelial cell and the a-granules of the platelet. Upon activation of the platelet or vascular injury, P-Selectins are released and appear within minutes on the platelet plasma membrane.15 The P-Selectins mediate the platelet- endothelium interaction, as well as the platelet-leukocyte interaction in the presence of P-Selectin glycoprotein ligand –1 (PSGL-1) along with L-Selectin.17 P-Selectin has been demonstrated to have an inflammatory component. As activated neutrophils or monocytes migrate to a site of inflammation, P-Selectins on the endothelial surface bind to exposed Selectin receptors on the Platelet-Leukocyte complex initiating the rolling phase of leukocyte migration.15

Heparin has been shown to have negative effects on the platelet as evidenced by the circulating level of P-Selectin.18 This finding suggests using circulating levels of P-Selectin as an indication of platelet destruction prior to surgical intervention. Xie and colleagues demonstrated heparin derivatives can directly bind to the Selectin receptors abrogating the adhesion interaction.19 Finally, Kestin and coworkers suggest that heparin indirectly blocks the activation and expression of P-Selectins on the surface of platelets via inhibition of endogenous thrombin binding.20

Conclusions

Significant evidence elucidates that heparin induces platelet dysfunction. The mechanism, in which heparin inhibits platelets, occurs at the receptor level. The adhesion receptors, GP IIb/IIIa, GP Ib-IX-V, and P-Selectin, are one class of platelet membrane receptors, which are susceptible to detrimental effects of heparin. Of the three adhesion receptors, GP IIb/IIIa shows the least ill response in interaction with heparin. However, the heparin/protamine complex has demonstrated a decrease in fibrinogen bonds with platelet receptor GP IIb/IIIa, which results in a decrease secretion of platelet granules and decrease platelet aggregation. Heparin causes dysfunction of the GP Ib-IX-V receptor by preventing the binding of vWf. By preventing vWf from binding to the GP Ib-IX-V receptor, there is a decrease in platelet aggregability at the site of injury on the subendothelium. Finally, heparin has an indirect effect in decreasing the activation and expression of P-Selectin on the platelet and endothelial membrane. Heparin produces its inhibitory effects on P-Selectins through the inhibition of endogenous thrombin. Although it is necessary to incorporate adequate anticoagulation in the CPB process, the detrimental effects of heparin on platelets and platelet adhesion receptors should not be ignored, because dysfunctional platelets lead to inefficient hemostasis and consequently, increased postoperative bleeding.

References

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