B. Type 3.

C. Type 2B.

D. Type 2A.

E. Type 2M.

B. Factor II.

C. Factor X.

D. Factor XI.

E. Prekallikrein.

B. Cryoprecipitate challenge.

C. Platelet function analyzer (PFA)-100.

D. Partial thromboplastin time (PTT) mixing study.

E. von Willebrand factor (vWF) multimer analysis.

B. Type 2N.

C. Type 2A.

D. Type 2M.

E. Type 1.

4b. The initial diagnosis of the patient was delayed because vWF antigen and activity results were within the reference range when she was initially evaluated 2 years earlier. In the interim, however, she continued to have menorrhagia, prompting reevaluation and the subsequent diagnosis of type of vWD in Question 4a. Which one of the following is the most likely reason that vWD was not diagnosed on her initial visit?

B. At the time of initial evaluation the patient was taking oral contraceptives, which are known to increase vWF levels.

C. The blood sample for testing was transported to the laboratory at room temperature instead of the preferred temperature of 4°C.

D. At the time of the initial evaluation, the patient had not recently exercised, and a lack of physical activity is known to increase vWF levels.

E. vWF levels are known to decrease with increasing age.

5. Which one of the following is an acquired cause of factor VII deficiency?

B. Inflammatory bowel disease.

C. Phytonadione therapy.

D. Heparin therapy.

E. Ganciclovir therapy.

6. Which one of the following clotting factors has the longest plasma half-life?

B. Factor X.

C. Factor II.

D. Factor XIII.

E. Factor VIII.

7. Which one of the following is a rare complication of congenital factor VII deficiency?

B. Thrombosis.

C. Menorrhagia.

D. Central nervous system bleeding.

E. Postoperative bleeding.

8. A 3-day-old infant, born by normal spontaneous delivery without complications, is brought to the emergency department with vomiting, lethargy, and pallor. On physical examination, the anterior fontanelle is bulging, and a cephalohematoma is diagnosed. A complete blood cell count shows a hemoglobin of 7.2 g/dL (11.0 to 13.0 g/dL), platelet count of 254 (165 to 415 × 10⁹/L), prothrombin time (PT) of 62.4 seconds (12.3 to 14.6 seconds), and activated partial thromboplastin time (aPTT) of 32.1 seconds (24.0 to 34.6 seconds). The PT corrects into the reference range when mixed 1:1 with pooled normal plasma. Which one of the following factors is most likely to be decreased in this patient?

B. Factor VII.

C. Factor VIII.

D. Factor IX.

E. Factor XI.

9. Which one of the following reasons makes it difficult to treat actively bleeding patients with congenital factor VII deficiency using recombinant factor VIIa?

B. Short half-life of recombinant factor VIIa.

C. Requirement for high doses (120 μg/kg).

D. Contamination with activated factors II, IX, and X.

E. Need to overlap with heparin therapy.

B. Consistent with inadequate dosing of factor VIII.

C. Inadequate reconstitution of lyophilized product.

D. Presence of a lupus anticoagulant (LAC).

E. Factor VIII inhibitor.

B. 1000 IU.

C. 2500 IU.

D. 3500 IU.

E. 5000 IU.

B. Factor VII.

C. Factor VIII.

D. Factor X.

E. Factor V.

11b. The patient has never been transfused and has not received factor replacement therapy. A factor IX activity is reported as less than 1% and factor VIII as 82%. The patient weighs 30 kg, and a target of 50% factor IX activity is desired. Which one of the following is the appropriate dose of recombinant factor IX replacement?

B. 210 IU.

C. 750 IU.

D. 1500 IU.

E. 5000 IU.

12. What is the biologic half-life of factor IX?

B. 12 hours.

C. 24 hours.

D. 48 hours.

E. 96 hours.

13. Which one of the following clinical manifestations is characteristic of patients with a coagulation factor deficiency?

B. Superficial hematomas.

C. Epistaxis.

D. Delayed bleeding.

E. Gum bleeding.

14. Which one of the following coagulation factor proteins is vitamin K dependent?

B. Factor II.

C. Factor VIII.

D. Factor V.

E. Factor XIII.

15. Polymorphisms in which one of the following genes is associated with variable response to warfarin therapy?

B. CYP2D9.

C. ABCB1.

D. CYP3A4.

E. CYP2J2.

16. A 32-year-old Ashkenazi Jewish woman, 38 weeks pregnant (G2P1001), is in active labor and is failing to progress. A cesarean delivery is planned. During the delivery of her first child, she bled profusely and required transfusion of 2 U of packed red cells. At her obstetrician’s office, records show that she has a complete blood cell count with a hemoglobin of 10.2 g/dL (11.0 to 13.0 g/dL), platelets 254 (165 to 415 × 109/L), PT 13.4 seconds (12.3 to 14.6 seconds), and aPTT 54.1 seconds (24.0 to 34.6 seconds). The patient’s aPTT corrected into the reference range when mixed 1:1 with pooled normal plasma. Factor levels are factor VIII 180% (50% to 150%), factor IX 107% (50% to 150%), and factor XI 12% (50% to 150%). Which one of the following is the best choice to treat active bleeding in this patient?

B. Fresh frozen plasma (FFP), 15 mL/kg.

C. Recombinant factor VIII.

D. Fibrin glue.

E. Recombinant factor XI.

17. Which one of the following coagulation protein deficiencies is not associated with a bleeding disorder?

B. Factor VII.

C. Factor XI.

D. Factor XII.

E. Factor XIII.

B. Protein C deficiency.

C. Factor XIII deficiency.

D. Prekallikrein deficiency.

E. High-molecular-weight kininogen deficiency.

19. Your laboratory performs a factor X activity assay using a minimum of three dilutions of patient plasma. The three plasma dilutions exceed the allowable within-run coefficient of variation. Which one of the following disclaimers should be reported with the result?

B. Inhibitor pattern.

C. Noncalibrated curve.

D. Linear range.

E. Between-subject variability.

20. Which one of the following acquired coagulation factor deficiencies is associated with primary systemic amyloidosis?

B. Factor VII.

C. Factor VIII.

D. Factor X.

E. Factor XII.

B. Dysfibrinogenemia.

C. Factor XIII deficiency.

D. Fibrinolysis.

E. High-molecular-weight kininogen deficiency.

B. 40 to 60 seconds.

C. 60 to 80 seconds.

D. 60 to 120 seconds.

E. 80 to 120 seconds.

23. For which one of the following clinical situations would thrombophilia testing be most indicated?

B. 49-year-old man with hypertension, hyperlipidemia, and diabetes with first stroke.

C. 55-year-old man with first mesenteric vein thrombosis.

D. 35-year-old woman with first spontaneous abortion in first trimester.

E. 45-year-old woman taking oral contraceptives.

24. Which of the following can lead to falsely low protein S activity on a PTT-based protein S activity assay?

B. Decreased factor V activity.

C. Direct thrombin inhibitor.

D. Factor V Leiden mutation.

E. Pregnancy.

B. Severe factor VII deficiency.

C. LAC.

D. Disseminated intravascular coagulation (DIC).

E. Factor IX inhibitor.

25b. Which one of the following must be demonstrated during laboratory testing to identify an LAC?

B. Phospholipid dependence of the inhibitor.

C. Mixing study in which the aPTT corrects into the reference range.

D. Prolongation of a phospholipid-independent clotting test.

E. Mixing study where the aPTT prolongation is time dependent.

25c. Which one of the following is an antiphospholipid antibody that is evaluated for the diagnosis of antiphospholipid antibody syndrome?

B. Antinuclear antibody.

C. Anti–Jo-1 antibody.

D. Anti–ADAMTS-13 antibodies.

E. Anti–β₂-glycoprotein 1 antibodies.

B. A DVT and positive mixing study showing the presence of an inhibitor are diagnostic for APS.

C. An arterial thrombosis is necessary to diagnose APS.

D. An antiphospholipid antibody must be demonstrated on more than one occasion to meet the diagnostic criteria for APS.

E. Detection of any antiphospholipid antibody is itself diagnostic of APS.

B. Aspirin.

C. Packed red blood cell transfusion.

D. Plasma transfusion.

E. Platelet transfusion.

B. Heparin.

C. Packed red blood cell transfusion.

D. Plasma transfusion.

E. Platelet transfusion.

B. Heparin.

C. Packed red blood cell transfusion.

D. Plasma transfusion.

E. Platelet transfusion.

29. Which one of the following assays is the best method for measuring antithrombin (AT) activity?

B. Enzyme-linked immunoassay (ELISA).

C. Latex particle immunoturbidometric.

D. DNA analysis.

E. Heparin-binding site inhibition.

30. Which one of the following tests is the most sensitive in detecting dysfibrinogenemia?

B. Fibrometer-based fibrinogen.

C. Nephelometry.

D. Reptilase time.

E. Thrombin time.

31. Which one of the following is the closest approximation of the prevalence of the factor V Leiden mutation in Caucasian Americans?

B. 1%.

C. 5%.

D. 15%.

E. 25%.

32. Which one of the following is an acquired cause of low AT activity?

B. l-Asparaginase therapy.

C. Hematuria.

D. Menopause.

E. Warfarin therapy.

33. Your laboratory performs a second-generation modified functional activated protein C (APC) resistance screen by determining the activated protein C resistance (APCR) ratio of an aPTT. Which one of the following situations will most likely cause a false-positive result?

B. Warfarin therapy.

C. Heparin 0.7 U/mL.

D. LAC.

E. Liver dysfunction.

34. Which one of the following can increase protein C activity in an aPTT-based assay?

B. Factor V Leiden.

C. LAC.

D. Warfarin therapy.

E. Thrombosis.

35. Which one of the following disorders can present with purpura fulminans and DIC in newborns?

B. Dysfibrinogenemia.

C. Homozygous factor V Leiden.

D. Homozygous protein C deficiency.

E. Homozygous prothrombin 20210G>A mutation.

36. Which one of the following is an effect associated with heparin therapy?

B. Development of anti–heparin polyvinyl sulfate antibodies.

C. Development of Hollenhorst plaques.

D. Hypokalemia.

E. Hirsutism.

37. The international sensitivity index (ISI) of the thromboplastin used in your laboratory is 1.26 according to the package insert. You perform the PT with this reagent in 120 normal donors and find the arithmetic mean to be 13.5 seconds, the geometric mean 13.7 seconds, and the standard error 3.5%. A patient’s PT is 19.8 seconds using this reagent. Which one of the following is the closest value for the international normalized ratio (INR) for this patient? (Note: A calculator to determine the value is allowed.)

B. 1.47

C. 1.59

D. 1.62

E. 5.67

38. A 64-year-old man develops a DVT after a right total hip replacement. Before initiating heparin therapy, laboratory studies show a PT of 14.8 seconds (12.7 to 15.4) and an aPTT of 29.3 seconds (26.3 to 39.4). The heparin therapeutic range is an aPTT of 60 to 80 seconds. Heparin therapy is initiated with a bolus of 5000 IU and then 1000 IU/hr. Six hours later, the aPTT is 35 seconds. Another bolus of 5000 IU of heparin is administered, and the infusion is increased to 1500 IU/hr. The aPTT 6 hours later is 44 seconds. An anti-Xa level is performed and found to be 1.1 IU/mL. Which one of the following is the most likely explanation for these findings?

B. AT deficiency.

C. Acquired factor VIII deficiency.

D. APCR.

E. Increased heparin cofactor II activity.

39. A 55-year-old man requires heparin reversal after cardiopulmonary bypass for aortic valve repair. Which one of the following is the best method for reversal of the anticoagulant effect of unfractionated heparin (UFH) in this patient?

B. Vitamin K.

C. Cryoprecipitate.

D. Prothrombin complex concentrates.

E. Protamine sulfate.

B. 0.90

C. 0.95

D. 1.0

E. 1.1

41. Which one of the following is an adverse effect of warfarin therapy?

B. Transmissible spongiform encephalopathy.

C. Purple toe syndrome.

D. Immune thrombocytopenia.

E. Hollenhorst plaque.

42. Several hemostatic changes occur during the different stages of the liver transplant procedure. Which one of the following stages of liver transplantation is associated with hypercoagulability?

B. Anhepatic to reperfusion.

C. Postreperfusion.

D. Postoperative.

E. During surgical dissection and mobilization of the diseased liver.

43. Which one of the following is the mechanism of thrombosis associated with AT III deficiency?

B. Failure to generate APC.

C. Failure of APC to inactivate factors Va and VIIIa.

D. Failure to generate plasmin.

E. Failure to inhibit factors IIa (thrombin), Xa, and other activated factors.

44. Which one of the following is a recognized mechanism for thrombosis in patients with dysfibrinogenemia?

B. Abnormal fibrinogen inhibits protein C activity.

C. Abnormal fibrinogen resists fibrinolysis.

D. Abnormal fibrinogen induces cytotoxicity and disrupts vascular hemostatic mechanisms.

E. Abnormal fibrinogen disrupts the activation of plasminogen.

45. Which one of the following is the mechanism of thrombosis in the setting of factor V Leiden?

B. Neutralization of tissue plasminogen activator (tPA).

C. Endothelial cell toxicity and disruption of vascular hemostatic mechanisms.

D. Activated protein C is unable to inactivate factor Va.

E. Failure to inhibit thrombin.

46. Which one of the following is the mechanism for thrombosis in the setting of heparin cofactor II deficiency?

B. Failure to activate plasminogen.

C. Failure to inhibit factor IIa (i.e., thrombin), factor Xa, and other activated factors.

D. Failure to generate APC.

E. Neutralization of tPA.

47. Which one of the following anticoagulant factors is solely produced by the liver?

B. AT.

C. Protein S.

D. Plasminogen.

E. Tissue factor pathway inhibitor.

48. Which one of the following procoagulant hemostatic factors is produced solely by the liver?

B. Fibrinogen.

C. Factor V.

D. Plasminogen activator inhibitor.

E. Plasminogen.

49. Which one of the following is the mechanism of thrombosis associated with hyperhomocysteinemia?

B. Failure to inhibit factor IIa (i.e. thrombin), factor Xa, and other activated factors.

C. Failure to activate plasminogen.

D. Endothelial cell toxicity and disruption of vascular hemostatic mechanisms.

E. Failure of APC to inactivate factors Va and VIIIa.

50. Which one of the following statements most accurately reflects the utility of routine coagulation tests to predict bleeding or thrombotic risk in liver disease?

B. There is good evidence that the PT, not the PTT, is useful in predicting bleeding or thrombotic risk in liver disease.

C. There is no good evidence to support the use of routine coagulation tests for predicting bleeding or thrombotic risk in liver disease.

D. There is good evidence to support the use of the PTT for predicting bleeding or thrombotic risk in liver disease.

E. There is good evidence to support the use of the thrombin time for predicting bleeding or thrombotic risk in liver disease.

51. Which one of the following disorders is associated with neutralization of tPA?

B. AT deficiency.

C. Factor V Leiden.

D. Heparin cofactor II deficiency.

E. Excess plasminogen activator inhibitor 1 activity.

52. Which one of the following mechanisms for thrombosis is associated with plasminogen deficiency?

B. Failure to inhibit thrombin.

C. Failure to generate plasmin.

D. Failure to generate APC.

E. Failure of activated protein C to inactivate factor Va.

53. Which one of the following is the mechanism of thrombosis in protein C deficiency?

B. Failure to generate APC.

C. Failure to generate plasmin.

D. Endothelial cell toxicity and disturbance of vascular hemostatic mechanisms.

E. Failure of APC to inactivate factors Va and VIIIa.

54. Which one of the following is the mechanism of thrombosis in protein S deficiency?

B. Failure of APC to inactivate factors Va and VIIIa.

C. APC fails to inactivate factor Va because of a highly conserved point mutation.

D. Failure to activate plasminogen.

E. An acquired abnormality in fibrin results in decreased fibrinolysis.

55. Which one of the following is the mechanism of thrombosis associated with tPA deficiency?

B. Failure of APC to inactivate factors Va and VIIIa.

C. Failure to generate APC.

D. Failure to activate plasminogen.

E. Failure to inhibit factor IIa, factor Xa, and other activated factors.

Major points of discussion

■ Types 1 and 3 vWD are characterized by quantitative deficiencies of normal vWF.

■ Type 2A vWD is caused by a mutation in the vWF protease cleavage site, leading to increased enzymatic cleavage and a lack of high- and intermediate-molecular-weight multimers.

■ Type 2B vWD is caused by a gain-of-function-mutation in the platelet GPIb/V/IX binding site of vWF, leading to a “stickier” vWF.

■ Type 2M vWD is caused by a loss-of-function-mutation in the platelet GPIb/V/IX binding site of vWF, leading to decreased binding of vWF to platelets.

■ Type 2N vWD is caused by a mutation in the factor VIII binding site of vWF.

■ Thrombocytopenia following DDAVP administration in type 2B vWD is usually transient and often is not associated with bleeding or thrombosis.

■ Interpretation of screening tests, as well as clotting factor activities, should be based on age-specific reference intervals.

■ Clotting factors with lower levels in young children than in adults include the vitamin K–dependent factors (II, VII, IX, and X), as well as factors V, XI, and XII, prekallikrein, and high-molecular-weight kininogen.

■ Clotting factors with similar or higher levels in young children than in adults include fibrinogen, factor VIII, and vWF.

■ PT-vWD is caused by a gain-of-function mutation in GPIbα that leads to increased binding to vWF.

■ Type 2B vWD and PT-vWD have similar clinical and laboratory findings. Although variable, patients classically present with thrombocytopenia, decreased vWF:Ag and vWF:ristocetin cofactor activity (RCo), increased vWF:Ag/vWF:RCo ratio, lack of high-molecular-weight vWF multimers, decreased FVIII:C, and abnormal ristocetin-induced platelet aggregation.

■ It is important to distinguish these two disorders because the treatment will differ. Administration of vWF/FVIII concentrates in vWF type 2B would be effective, whereas in PT-vWD, it would not. Desmopressin (DDAVP) is contraindicated in PT-vWD.

■ vWF is an acute-phase reactant and therefore is increased by stress, inflammation, pregnancy, and oral contraceptives. Repeated testing is necessary in some cases to identify low vWF levels.

■ Medical conditions that can decrease vWF levels or activity include autoimmune/antibody-induced inhibition or clearance and destruction of vWF by increased shear stress (e.g., left ventricular assist devices).

■ ABO blood type has an effect on plasma vWF levels. Individuals with group O blood type have approximately 25% lower vWF levels than individuals with other blood types.

■ vWF levels tend to be higher in neonates than adults.

■ Acquired factor VII deficiency can be due to acquired vitamin K deficiency (e.g., diet, antibiotic therapy, total parenteral nutrition, and gastrointestinal malabsorption), liver disease, and warfarin therapy.

■ Vitamin K is required for the γ-carboxylation of the vitamin K–dependent coagulation factors (factors II, VII, IX, and X and proteins C and S). In the absence of vitamin K and γ-carboxylation, these factors are not active.

■ Recycling vitamin K requires enzymes that are inhibited by warfarin, which leads to inactive vitamin K–dependent coagulation and anticoagulation factors. Exogenous administration of vitamin K (phytonadione) is used as an antidote for the warfarin effect.

■ Antibiotics can alter gut flora, which leads to vitamin K deficiency. Hospitalized patients are often taking antibiotics and may be NPO or receiving total parenteral nutrition, which can make them vitamin K deficient.

■ Hemorrhagic disease of the newborn can be seen in neonates who do not get adequate vitamin K across the placenta before delivery.

■ The vitamin K–dependent factors II, VII, IX, and X and proteins C and S are produced in the liver. Decreased synthetic ability of the liver leads to decreased coagulation factor levels.

6. A. Factor VII.
Rationale: Factor VII has a half-life of approximately 5 hours.
B. Factor X.
Rationale: Factor X has a half-life of approximately 40 hours.
C. Factor II.
Rationale: Factor II has a half-life of approximately 65 hours.
D. Factor XIII.
Rationale: Factor XIII has a half-life of approximately 200 hours.
E. Factor VIII.
Rationale: Factor VIII has a half-life of approximately 10 hours.

Major points of discussion

■ For example, recombinant factor VIIa is dosed every 2 hours, factor VIII every 12 hours, and factor IX every 24 hours.

■ Factor VII has the shortest half-life.

■ Factor XIII has the longest half-life.

■ The approximate plasma half-lives of the clotting factors are as follows:

• Factor II: 65 hours

• Factor V: 15 to 36 hours

• Factor VII: 5 hours

• Factor VIII: 10 hours

• Factor IX: 25 hours

• Factor X: 40 hours

• Factor XI: 45 to 80 hours

• Factor XII: 50 to 70 hours

• Factor XIII: 200 hours

7. A. Epistaxis.
Rationale: This is a common complication.
B. Thrombosis.
Rationale: This is a rare complication of congenital factor VII deficiency and is associated with surgery and sometimes recombinant factor VIIa therapy.
C. Menorrhagia.
Rationale: Up to 60% of women with congenital factor VII deficiency have menorrhagia.
D. Central nervous system bleeding.
Rationale: Occurs most commonly in the first 6 months of life.
E. Postoperative bleeding.
Rationale: Up to 30% of patient will bleed after surgery.

Major points of discussion

■ It is an autosomal recessive disorder and has an incidence of 1 in 500,000.

■ The genotype and factor activity level in congenital factor VII deficiency do not correlate well with clinical bleeding.

■ Clinical manifestations include epistaxis, mucosal bleeding, soft tissue hematomas, hemarthrosis, menorrhagia, and postoperative bleeding.

■ A mixing study with pooled normal plasma should correct the prolonged PT when a factor deficiency is present.

■ Clinical manifestations of congenital factor VII deficiency include epistaxis, mucosal bleeding, soft tissue hematomas, hemarthrosis, menorrhagia, postoperative bleeding, and uncommonly, thrombosis.

■ Recombinant factor VIIa at a low dose of 20 to 30 μg/kg is used to treat congenital factor VII deficiency.

■ Although plasma infusions can be used for treatment in this setting, there is a risk for volume overload, other transfusion reactions, and transfusion transmitted infection.

9. A. Formation of anti–factor VII antibodies.
Rationale: Formation is possible but extremely rare in patients with congenital factor VII deficiency.
B. Short half-life of recombinant factor VIIa.
Rationale: Recombinant factor VIIa must be administered every 2 hours because of the short half-life.
C. Requirement for high doses (120 μg/kg).
Rationale: Low doses (20 to 30 μg/kg) are usually given; this is a much lower dose than for patients with factor VIII inhibitors.
D. Contamination with activated factors II, IX, and X.
Rationale: Activated prothrombin concentrates can have activated factors II, VII, IX, and X. However, recombinant factor VIIa does not contain the activated factors II, IX, and X.
E. Need to overlap with heparin therapy.
Rationale: Initiation of warfarin requires heparin overlap, but recombinant factor VIIa is not used for anticoagulation and does not require heparin overlap.

Major points of discussion

■ Although plasma infusions have been used, there is risk for volume overload and other transfusion reactions, as well as transfusion-transmitted infection.

■ The biological half-life of rFVIIa is 2 hours, so it may have to be administered frequently until bleeding ceases.

■ rFVIIa in vivo action is inhibited by tissue factor pathway inhibitor (TFPI).

■ rFVIIa is also used to treat bleeding in patients with hemophilia A or B with inhibitors or patients with acquired factor VIII or IX inhibitors.

10a. A. Consistent with factor VIII half-life.
Rationale: The half-life of factor VIII is 12 hours, so approximately 25% of factor VIII activity should remain after 12 hours (the administered dose should result in 50% peak factor VIII levels). The mixing study in this case is consistent with an inhibitor.
B. Consistent with inadequate dosing of factor VIII.
Rationale: The dose is calculated correctly for factor VIII replacement: body weight (kg) × desired % increase × 0.5 IU/kg.
C. Inadequate reconstitution of lyophilized product.
Rationale: The mixing study is not consistent with a factor deficiency.
D. Presence of a lupus anticoagulant (LAC).
Rationale: An LAC could produce an isolated elevated PTT without correction on a mixing study. However, the time dependence of the antibody and bleeding, rather than thrombosis, suggests that this is less likely an LAC.
E. Factor VIII inhibitor.
Rationale: The percentage recovery of factor activity is too low after 12 hours, and the mixing study shows a time-dependent inhibitor. These findings are most consistent with a factor inhibitor.

Major points of discussion

■ Anti–factor VIII antibodies can be detected on coagulation studies by performing a mixing study. A mixing study is performed by mixing one volume of patient plasma with one volume of pooled normal plasma. If a factor deficiency is present, the pooled normal plasma will correct the prolonged aPTT. However, the presence of an alloantibody will inhibit the factor VIII in both the patient plasma and the pooled normal plasma.

■ LACs can also inhibit coagulation by interfering with the phospholipid surfaces that coagulation factors require for their activity. However, the inhibitor activity is usually seen upon immediate mixing and with incubation. In addition, LACs are more likely to cause thrombosis, not bleeding.

■ Anti–factor VIII alloantibodies are generally weaker antibodies than LACs, and with incubation, their inhibition will strengthen. Therefore, the immediate mix may correct, and then with incubation for 1 to 2 hours, the aPTT will prolong as the antibody’s binding is enhanced.

■ The strength of an antibody should be determined using a Bethesda assay, which measures the titer of the antibody. The higher the Bethesda units (BUs), the greater the strength of the antibody. A BU reflects the amount of inhibitor that will inactivate half of the factor activity.

■ If there are fewer than 5 BUs, then overwhelming the inhibitor with increasing factor replacement can be attempted (i.e., give a very large dose of factor VIII). However, if there are more than 5 BUs, then alternate concentrates (e.g., porcine factor VIII) or bypassing agents (e.g., factor VIII inhibitor bypassing activity, prothrombin complex concentrates, recombinant factor VIIa) should be used to treat active bleeding.

10b. A. 750 IU.
B. 1000 IU.
Rationale: These doses are too low for weight and target goal.
C. 2500 IU.
Rationale: This dose may be appropriate for factor VIII replacement.
D. 3500 IU.
E. 5000 IU.
Rationale: These doses are too high. The target should be 50%.

Major points of discussion

■ Calculated dose in units = [% Change in factor activity desired] × [Body weight (kg)] × 0.5 IU/kg.

■ For this patient, assuming factor VIII activity of 0%, a target of 50%, and body weight of 100 kg: Calculated dose = 50% × 100 kg × 0.5 IU/kg = 2500 IU.

■ The target goal for hemarthrosis, gastrointestinal bleeding, and dental extraction is 50%; major surgery and intracranial hemorrhage should have a target goal of 100%.

■ The half-life of factor VIII is 12 hours, so the maintenance dose (50% of loading dose) should be administered approximately every 12 hours.

11a. A. Factor II.
Rationale: Factor II deficiency should affect both the PT and the aPTT.
B. Factor VII.
Rationale: Factor VII deficiency is usually associated with an elevated PT and a normal aPTT.
C. Factor VIII.
Rationale: An isolated aPTT in the setting of clinical bleeding can be caused by factor VIII, factor IX, and factor XI deficiencies.
D. Factor X.
Rationale: The PT and aPTT are usually affected in factor X deficiency.
E. Factor V.
Rationale: The PT and aPTT are usually affected in factor V deficiency.

Major points of discussion

■ Hemophilia A is inherited in an X-linked recessive manner so that males are affected more than females. Females can develop hemophilia A by inheriting two affected alleles, having Turner syndrome (XO), or having skewed lyonization.

■ Factor deficiencies associated with an elevated aPTT but a normal PT include factors VIII, IX, XI, and XII; prekallikrein; and high-molecular-weight kininogen deficiencies. Deficiencies in prekallikrein, factor XII, and high-molecular-weight kininogen are not associated with an increased risk for bleeding.

■ An elevated PTT caused by a factor deficiency can be corrected in vitro by mixing patient plasma with pooled normal plasma (mixing study) if an inhibitor is not present.

■ Hemophilia A is classified into severe (< 1% activity), moderate (1% to 5%), and mild (> 5% to 30%).

• Moderate hemophilia is associated with bleeding with minor trauma or surgery.

• Mild hemophilia usually requires a greater degree of trauma or invasive procedures to cause bleeding.

■ Clinical manifestations include hemarthrosis, deep soft tissue hematomas, hematuria, intracranial hemorrhage, and gastrointestinal bleeding.

11b. A. 75 IU.
B. 210 IU.
C. 750 IU.
Rationale: These doses are too low for weight and target goal.
D. 1500 IU.
Rationale: This dose may be appropriate.
E. 5000 IU.
Rationale: This dose is too high.

Major points of discussion

■ Calculated dose in units = % Change in factor activity desired × Body weight (kg) × 1.0 IU/kg.

■ For this patient, assuming factor IX activity of 0%, weight of 30 kg, and desired factor level of 50%: Calculated dose = 50% × 30 kg × 1.0 IU/kg = 1500 IU.

■ Target goal for hemarthrosis is 50%; major surgery has a target of 100%.

■ The half-life of factor IX is 18 to 24 hours, so the maintenance dose (50% of loading dose) should be administered approximately every 24 hours.

■ In addition, factor IX distributes into the extravascular space, so recovery of factor activity after infusion may not reliably be 100%. Therefore, a correction factor to increase the dose to account for the loss of intravascular recovery may be required based on empirical studies for a specific patient.

12. A. 5 hours.
Rationale: The half-life of factor VII is approximately 5 hours.
B. 12 hours.
Rationale: The half-life of factor VIII is 12 hours.
C. 24 hours.
Rationale: The half-life of factor IX is 18 to 24 hours.
D. 48 hours.
Rationale: The half-life of factor XI is approximately 50 hours.
E. 96 hours.
Rationale: The half-life of fibrinogen is approximately 4 days.

Major points of discussion

■ Targets of 50% are desirable for joint bleeding, dental procedures, hematuria, and gastrointestinal bleeding.

■ Higher targets of 100% are desirable for soft tissue hematomas, intracranial hemorrhage, and surgery.

■ The half-life of factor IX is 18 to 24 hours, which means a loading dose should be provided and then a maintenance dose (usually one half the loading dose) administered every 24 hours.

■ Administration of recombinant factor IX can have 60% to 80% recovery because of the rapid extravascular distribution of the factor after administration.

13. A. Petechiae.
B. Superficial hematomas.
Rationale: Deep hematomas are characteristic of coagulation deficiencies.
C. Epistaxis.
D. Delayed bleeding.
Rationale: Delayed postoperative bleeding is characteristic of coagulation deficiencies.
E. Gum bleeding.
Rationale for A, C, and E: Seen in platelet-type bleeding.

Major points of discussion

■ Coagulation factor deficiencies are inherited in an X-linked recessive (factor VIII or factor IX deficiency), autosomal recessive (factor VII or XI deficiency), or autosomal dominant (vWD) manner.

■ Petechiae, persistent bleeding from superficial cuts, and mucosal bleeding such as gum bleeding characterize platelet-type bleeding.

■ Coagulation factor deficiencies are more commonly seen in men because of sex-linked inheritance.

■ It is important to keep in mind that abnormalities of the blood vessels may also lead to bleeding disorders.

14. A. Factor XII.
B. Factor II.
Rationale: Factors II, VII, IX, and X and proteins C and S are vitamin K–dependent factors.
C. Factor VIII.
D. Factor V.
E. Factor XIII.
Rationale for A, C, D, and E: These factors are not vitamin K dependent.

Major points of discussion

■ Vitamin K is a cofactor for γ-carboxylation of glutamic acid residues of the vitamin K–dependent coagulation proteins.

■ The carboxylation of glutamic acid residues results in the chelation of calcium ions and allows the proteins to bind to the phospholipids on the platelet cell membranes to promote the assembly of coagulation factor complexes on the platelet surface.

■ Without sufficient vitamin K, γ-carboxylation does not take place, the proteins are unable to bind to phospholipids, and coagulation proteins cannot interact, making them functionally inactive.

■ Warfarin exerts its effect by inhibiting vitamin K 2,3-epoxide reductase, thereby blocking regeneration of the active form of vitamin K, leading to a lack of γ-carboxylation.

15. A. VKORC1.
Rationale: VKORC1 single-nucleotide polymorphisms are associated with variable metabolism of warfarin.
B. CYP2D9.
Rationale: CYP2C9, not CYP2D9, is a hepatic P450 enzyme associated with variable metabolism of warfarin.
C. ABCB1.
Rationale: Encodes P-glycoprotein, a drug efflux pump, that may be involved in dabigatran availability.
D. CYP3A4.
E. CYP2J2.
Rationale: Involved in the metabolism of rivaroxaban.

Major points of discussion

■ VKORC1 and CYP2C9 are two genes with single-nucleotide polymorphisms that predict variable response to warfarin metabolism.

■ CYP2C9 is a P450 enzyme involved in warfarin metabolism. The variant alleles CYP2C9*2 and CYP2C9*3 metabolize warfarin more slowly and may require a lower initial dose.

■ VKORC1 encodes the vitamin K epoxide reductase gene. Variable alleles are associated with variable metabolism of warfarin. Individuals with the group A haplotype produce less VKORC1, and lower warfarin doses are needed in these patients.

■ It is more commonly seen in patients with an Ashkenazi Jewish background.

■ Patients may have a range of clinical manifestations from no bleeding tendency to severe bleeding. Factor XI levels do not correlate well with the bleeding risk.

■ The aPTT is prolonged, and the PT is normal. The prolonged aPTT can be corrected by a 1:1 mix with pooled normal plasma. Homozygotes and compound heterozygotes have factor XI levels of less than 15%.

■ Mild bleeding episodes may not have to be treated. However, factor levels above 30% are generally desired before surgery.

■ If treatment is required, a loading dose of 15 to 20 mL/kg of FFP is administered, followed by a 3 to 6 mL/kg dose every 12 hours. Factor XI has a half-life of approximately 50 hours.

■ For severe bleeding, recombinant factor VIIa has been used, including in patients with factor XI inhibitors. These indications are not approved by the FDA for use with recombinant factor VIIa.

17. A. Factor II (prothrombin).
Rationale: Factor II deficiency is associated with hematomas and hemarthroses.
B. Factor VII.
Rationale: Congenital and acquired factor VII deficiencies are associated with bleeding.
C. Factor XI.
Rationale: Factor XI deficiency is associated with variable degrees of bleeding from asymptomatic to severe. Factor levels do not predict bleeding manifestations.
D. Factor XII.
Rationale: Factor XII deficiency is not associated with bleeding.
E. Factor XIII.
Rationale: Factor XIII deficiency is associated with bleeding.

Major points of discussion

■ Factor XII deficiency results in an elevated aPTT with a normal PT. However, factor XII deficiency is not associated with bleeding. Rather, factor XII deficiency is thought to have delayed arterial thrombosis.

■ Factor VII deficiency is associated with bleeding that is treated with recombinant factor VIIa or plasma.

■ Deficiencies in the contact system proteins can result in an elevated aPTT with a normal PT. A mixing study with pooled normal plasma would correct the aPTT (i.e., deficiency, not an inhibitor).

■ Factor XII, prekallikrein, and high-molecular-weight kininogen deficiencies are not associated with bleeding.

■ Complete normalization of prolonged aPTT following prolonged incubation is characteristic of prekallikrein deficiency and is caused by autoactivation of factor XII (activation of factor XII by factor XIIa).

■ The one-stage coagulation factor assay is an example of a parallel line bioassay. In this type of assay, the results are reliable when the curves of reference and patient plasma are parallel. If there is nonparallelism, the factor activity results should be considered incorrect.

■ A common reason for nonparallelism is the presence of LACs or other nonspecific inhibitors.

■ If three or more patient dilutions are used and they do not agree within a set tolerance limit, the assay is demonstrating an inhibitor pattern. A nonspecific inhibitor should be suspected.

■ The presence of anticoagulants will interfere with clotting times and may underestimate factor activities.

■ The presence of LACs will interfere with coagulation factor activity and can underestimate coagulation factor activity.

■ Elevated factor VIII levels (acute-phase reactant) can shorten the PTT and thus overestimate factor activity.

■ Amyloid fibers are thought to bind directly to factor X.

■ Treatment consists of FFP or prothrombin complex concentrate infusions.

■ Acquired factor X deficiency can result from vitamin K deficiency or liver disease, although the deficiency is usually not isolated; it is usually associated with decreased factors II, VII, and IX.

■ Respiratory infections, thymoma, and malignancies (e.g., renal carcinoma, adrenal adenocarcinoma, acute myeloid leukemia) have also been reported to be associated with acquired factor X deficiency.

■ Congenital factor X deficiency is rare (1 per 500,000). Acquired inhibitors to factor X are uncommon.

21. A. α₂-Antiplasmin deficiency.
Rationale: α₂-Antiplasmin deficiency would show normal clot dissolution in the 5M urea test.
B. Dysfibrinogenemia.
Rationale: Dysfibrinogenemia would show a prolonged thrombin time.
C. Factor XIII deficiency.
Rationale: The rapid clot dissolution in 5M urea is characteristic of factor XIII deficiency.
D. Fibrinolysis.
Rationale: An elevated d-dimer would be expected if bleeding were caused by fibrinolysis.
E. High-molecular-weight kininogen deficiency.
Rationale: High-molecular-weight kininogen deficiency would cause a prolongation of the aPTT and is not a cause of bleeding.

Major points of discussion

■ Factor XIII deficiency is a rare, autosomal recessive disorder. Factor XIII cross-links fibrin through peptide bonds. The clots from these patients dissolve rapidly in 5M urea or in 1% monochloracetic acid.

■ Factor XIII deficiency is associated with delayed umbilical stump bleeding, bleeding after circumcision, hematomas, soft tissue bleeding, and recurrent spontaneous abortions.

■ Factor XIII deficiency is treated with cryoprecipitate or factor XIII concentrates (Europe).

■ Patients with α₂-antiplasmin deficiency have a bleeding tendency because of decreased inactivation of plasmin, which leads to excess fibrinolysis.

■ The PT and aPTT are normal in α₂-antiplasmin deficiency. Solubility in 5M urea is normal. However, the euglobulin lysis time is shortened. Euglobulin is formed when plasma is acidified, and it contains important fibrinolytic factors such as plasminogen, tPA, and α₂-antiplasmin. Deficiency will result in a markedly accelerated lysis time.

■ Heparin binds to AT and accelerates its activity 1000-fold, leading to the inactivation of coagulation factors IIa, Xa, IXa, and XIa.

■ UFH is the most common injectable anticoagulant and is used for the treatment of acute coronary syndromes, venous thromboembolism (VTE), and atrial fibrillation. It is also used as the anticoagulant in cardiac bypass and extracorporeal membrane oxygenation circuits.

■ UFH is monitored by following the aPTT. The sensitivity of the aPTT to heparin concentration varies from lot to lot of aPTT reagent. Therefore, the therapeutic range should be established with each new lot of aPTT reagent.

■ The Brill-Edwards method of establishing the UFH therapeutic range requires samples from patients currently on UFH therapy, but not on warfarin (the PT should be in the reference range). UFH anti-Xa activity and the aPTT are measured on each sample. A linear regression is performed; the aPTT corresponding to 0.3 to 0.7 U/mL anti-Xa activity is read from the regression line. The aPTT range is set as the therapeutic range. If heparin-protamine titration is used, then 0.2 to 0.4 U/mL heparin would be used to determine the therapeutic range.

■ UFH is usually administered as a bolus dose and then a constant infusion. The half-life of heparin is 1 to 2 hours. Therefore, heparin therapy is usually monitored by measuring the aPTT every 6 hours while it is adjusted.

■ Adverse reactions to heparin include heparin-induced thrombocytopenia, osteopenia, and hyperkalemia.

■ Acquired risk factors for thrombophilia include malignancy, older age, estrogen replacement, immobilization (e.g., orthopedic surgery), chronic inflammation, and antiphospholipid antibodies.

■ Laboratory testing is recommended only for a select population of patients.

■ Indications for testing for patients with VTE:

• Recurrent VTE

• First-degree relatives with VTE

• Thrombosis at unusual sites, such as mesenteric vein, portal vein, splenic vein, renal vein, and cerebral sinus, without other risk factors

■ Acquired causes of protein S deficiency include vitamin K deficiency, warfarin therapy, liver disease, thrombosis, oral contraceptives, and estrogen replacement therapy.

■ PTT-based protein S assays are performed by diluting patient plasma with protein S–deficient plasma. Fixed amounts of APC and factor Va are added. Prolongation of the PTT is proportional to the amount of protein S in the sample (low protein S → less factor Va inactivated → shorter PTT).

■ PTT-based assays can be affected by LACs, APCR/factor V Leiden, elevated factor VIII activity, direct thrombin inhibitors, and heparin therapy.

■ Protein S circulates as free protein S and as C4 binding protein (C4bp)-bound protein S. C4bp-bound protein S is not active.

■ Thrombosis can be venous, arterial, or microvascular. Microvascular thrombosis may manifest as “catastrophic antiphospholipid syndrome” that typically involves multiorgan failure (lungs, brain, kidneys).

■ According to criteria defining APS, APS is present if at least one of the following clinical and one of the following laboratory criteria are met:

■ Clinical criteria:

• Pregnancy morbidity: (a) one or more unexplained deaths of a morphologically normal fetus at or beyond the tenth week of gestation; (b) one or more preterm births of a morphologically normal neonate before the thirty-fourth week of gestation because of eclampsia or severe pre-eclampsia, or recognized features of placental insufficiency; (c) three or more unexplained consecutive spontaneous miscarriages before the tenth week of gestation, with maternal anatomic or hormonal abnormalities and paternal and maternal chromosomal causes excluded.

■ Laboratory criteria:

• ACL antibody of immunoglobulin G (IgG) and/or IgM isotype in serum or plasma, present in medium or high titer, on two or more occasions, at least 12 weeks apart

• Anti–β₂-glycoprotein 1 antibody of IgG and/or IgM isotype in serum or plasma, present on two or more occasions at least 12 weeks apart

■ LAC activity is an in vitro phenomenon defined by prolongation of a phospholipid-dependent coagulation test that is not caused by an inhibitor to a specific coagulation factor. Criteria for the laboratory diagnosis of an LAC include the following:

• Demonstration of presence of an inhibitor by mixing tests

• Demonstration of the phospholipid dependence of the inhibitor

Present on two or more occasions 12 weeks apart

■ Two or more phospholipid-based screening tests should be used to detect LACs. Options for these tests include a low phospholipid concentration PTT, dilute Russell viper venom time, kaolin clotting time, and dilute PT. A confirmatory step using high phospholipid concentration, platelet neutralizing agent, or LAC-insensitive reagent should be performed to demonstrate phospholipid dependence.

■ A small wire is rotated in a well filled with whole blood (or the well is rotated), and an activator is added. In addition, heparin neutralizers can be added so that testing can be performed during cardiac bypass. As the clot forms, the resistance on the wire or cup increases. This resistance (measured in arbitrary millimeter units) is graphed over time to produce the thromboelastogram.

■ Thromboelastography can show specific defects of platelets, coagulation, and/or fibrinolysis.

■ The time from initiation of the test to the beginning of clot resistance is called the R time and indicates the function of coagulation factors. The rate of clot formation is indicated by the time it takes to reach 20 mm of clot strength.

■ The α angle gives information about fibrin production and clot strength.

■ The maximal amplitude achieved reflects platelet function.

■ Excess fibrinolysis is reflected in the formation of a clot that dissolves faster than normal.

■ The LY30 measures the percentage decrease in amplitude 30 minutes after maximal amplitude. The LY60 measures the percentage decrease in amplitude 60 minutes after maximal amplitude.

■ The reference range for the LY30 is about 7.5% but is activator dependent (e.g., celite activated or whole blood).

■ The LY30 is a measure of clot lysis (i.e., fibrinolysis).

■ Excess fibrinolysis is reflected in the formation of a clot that dissolves faster than normal.

■ Antifibrinolytic medications include aminocaproic acid, aprotinin, and tranexamic acid.

28. A. Tranexamic acid.
Rationale: Tranexamic acid is an antifibrinolytic agent. This thromboelastogram shows a hypercoagulable state.
B. Heparin.
Rationale: Heparin would be the most appropriate therapy for a hypercoagulable state.
C. Packed red blood cell transfusion.
Rationale: Packed red cells are used to treat anemia, not hemostatic defects. This thromboelastogram shows a hypercoagulable state.
D. Plasma transfusion.
Rationale: Plasma is used to replace a balance of normal levels of coagulation factors and natural anticoagulants. This thromboelastogram shows a hypercoagulable state, and the levels of natural anticoagulants in plasma would not be therapeutically sufficient.
E. Platelet transfusion.
Rationale: Platelet transfusions are used to treat quantitative or qualitative platelet disorders. This thromboelastogram shows a hypercoagulable state.

Major points of discussion

■ A hypercoagulable state can be measured using TEG by detecting a decreased R time and increased maximal amplitude (MA).

■ Causes of hypercoagulability include the postoperative state, oral contraceptives, pregnancy, deficiencies of natural anticoagulants, and cancer.

■ Thrombocytosis can increase coagulability on TEG by increasing the MA.

■ There are several assays that measure AT antigen levels. ELISA-based assays, immunodiffusion, and latex particle immunoturbidometric methods are the most common.

■ Type I AT deficiency is a decreased level of functionally normal AT. Antigen assays would be low, and functional assay would be low (i.e., quantitative defect).

■ Type II AT deficiency is a functionally abnormal AT present in normal quantities. Antigen assays would be normal, and functional assay would be low (i.e., qualitative defect).

■ Dysfibrinogenemias can be congenital or acquired. The most common causes of acquired dysfibrinogenemia are associated with liver disease.

■ Among dysfibrinogenemias, 55% are asymptomatic, 25% are associated with bleeding only, and 20% are associated with thrombosis with or without bleeding.

■ The thrombin time is performed by adding a low concentration of thrombin to platelet-poor plasma and measuring the time to clot formation. It is sensitive but is not a very specific test for dysfibrinogenemias. It can be prolonged in the presence of heparin, fibrin degradation products, hypofibrinogenemia, antibovine thrombin antibodies (secondary to fibrin glue), and others.

■ The reptilase time is performed by adding Bothrops atrox snake venom to platelet-poor plasma and measuring the time to clot formation. Reptilase time is prolonged in similar situations such as thrombin time, but it is not sensitive to heparin inhibition. Therefore, a prolonged thrombin time with normal reptilase time suggests the presence of heparin.

■ The FVL mutation is a mutation of factor V at the APC cleavage site (1691G>A). APC is 10 times slower in inactivating FVL than wild-type factor V.

■ FVL is the most common mutation (> 90%) causing APCR. APCR is caused by other factor V mutations (i.e., FV Cambridge, FV Liverpool, R485K mutation, R2 haplotype, A/G allele).

■ FVL is the most common genetic risk factor for VTE. It is found in 20% to 25% of patients with VTE and 50% of cases of familial VTE.

■ Administration of UFH can accelerate AT activity 1000-fold.

■ Congenital AT deficiency is seen in 0.02% to 0.17% of the general population and 0.5% to 4.9% of patients with VTE (i.e., very rare).

■ AT levels are decreased in liver disease, active thrombosis, surgery, DIC, sepsis, inflammatory bowel disease, l-asparaginase therapy, nephrotic syndrome, oral contraceptive therapy, and pregnancy.

■ First-generation assays were subject to false-negative and false-positive results because of factor deficiencies, the presence of heparin, warfarin therapy, and LACs.

■ Second-generation, modified functional APCR assays are performed after diluting the sample with factor V–deficient plasma to replace any factor deficiencies, thus removing the effect of warfarin therapy or protein C deficiency. The assay also adds polybrene to neutralize unfractionated and low-molecular-weight heparin.

■ Factor V–deficient plasma could have low protein S activity because of the manufacturing process. Therefore, protein S deficiency can still produce false-positive results.

■ LACs are still problematic in second-generation assays, although dilution with factor V–deficient plasma dilutes the antibody and its effect.

■ Protein C deficiency can be inherited, acquired, or an artifact of the testing system.

■ Inherited causes of protein C deficiency are seen in 0.14% to 0.5% of the general population and in 3% of patients with a first VTE; it increases the risk for VTE by sevenfold.

■ Acquired causes of protein C deficiency include vitamin K deficiency, warfarin therapy, liver disease, thrombosis, surgery, DIC, and l-asparaginase therapy.

■ In aPTT-based assays, protein C levels may be overestimated when there is an LAC or DTI. aPTT-based assays may also underestimate protein C activity in the presence of increased factor VIII activity and FVL.

■ Purpura fulminans is characterized by ecchymotic skin lesions that spontaneously become necrotic. Purpura fulminans is often associated with DIC.

■ Protein C is a vitamin K–dependent natural anticoagulant. Activated protein C, with its cofactor protein S, inactivates factors VIIIa and Va.

■ Heterozygous hereditary protein C deficiency is seen in 3% of patients with their first VTE and in only 0.14% to 0.5% of the general population.

■ Osteoporosis caused by osteoclast activation and osteoblast suppression can develop, especially with long-term therapy with low-molecular-weight heparin.

■ Less common complications include increased liver enzymes, alopecia, and hyperkalemia.

■ The PT measures extrinsic coagulation factors and is used to monitor warfarin therapy. Warfarin anticoagulation prevents the carboxylation of vitamin K–dependent coagulation factors (factor II, VII, IX, and X and proteins C and S).

■ The sensitivity of each thromboplastin reagent to vitamin K–dependent coagulation factors can differ significantly from manufacturer to manufacturer and lot to lot. The INR was developed so that patients taking warfarin could have their warfarin therapy monitored by different laboratories.

■ Each thromboplastin should have an ISI determined by testing it against an international or working reference preparation, which are thromboplastins available in several different preparations (e.g., rabbit, human, recombinant) from the World Health Organization.

■ To determine the INR of a patient’s PT, the geometric mean of the normal donor population for an individual laboratory must be determined by testing 120 normal donors. The arithmetic mean is not acceptable.

■ INR = (PT/PT_GeoMean)^ISI.

■ Log (INR) = ISI × PT/ PT_GeoMean.

■ PT/ PT_GeoMean is often referred to as the PT ratio.

■ The target INR depends on the patient’s condition; in most cases, the treatment goal is an INR between 2 and 3.

38. A. LAC.
Rationale: Although an LAC increases the risk for DVT, it does not explain the heparin resistance. LACs usually prolong the PT and/or aPTT.
B. AT deficiency.
Rationale: Inherited or acquired AT deficiency affects heparin therapy because heparin is not able to accelerate AT effectively if AT is present in low amounts.
C. Acquired factor VIII deficiency.
Rationale: The aPTT should be prolonged before initiation of therapy. Heparin resistance is seen with elevated factor VIII activity.
D. APCR.
Rationale: APCR, most commonly caused by the FVL mutation, is a risk factor for DVT. However, it should not lead to heparin resistance.
E. Increased heparin cofactor II activity.
Rationale: Heparin cofactor II binds heparin, which leads to inactivation of factor IIa. This would not lead to heparin resistance.

Major points of discussion

■ The aPTT is usually used to monitor UFH therapy. After a bolus dose and infusion, the aPTT is checked after 6 hours to determine whether the aPTT is in the therapeutic range. Depending on the aPTT, heparin can be adjusted to achieve the target based on being over, under, or at target.

■ However, if heparin is administered and the aPTT does not increase, heparin resistance should be considered. Usually, if more than 25,000 to 35,000 IU of heparin is administered in a 24-hour period without achieving the therapeutic target (as measured by aPTT or activated clotting time) constitutes heparin resistance. Anti-Xa activity of the heparin can be determined. If the anti-Xa is elevated out of proportion to the measured aPTT, then heparin resistance should be considered.

■ Common causes for heparin resistance are elevated factor VIII activity (acute-phase reactant), congenital or acquired AT deficiency, increased heparin clearance, and increased heparin-binding proteins.

■ Treatment can consist of switching to an alternate anticoagulant such as argatroban. In addition, administering AT through infusion of plasma or AT concentrates may be useful. Monitoring with anti-Xa activity instead of aPTT may be useful.

39. A. FFP.
Rationale: FFP may accentuate the anticoagulant effect by providing additional AT.
B. Vitamin K.
Rationale: Vitamin K is used for the reversal of warfarin.
C. Cryoprecipitate.
Rationale: Cryoprecipitate has no role in the reversal of heparin therapy.
D. Prothrombin complex concentrates.
Rationale: Prothrombin complex concentrates can be used for warfarin therapy but are not useful for heparin reversal.
E. Protamine sulfate.
Rationale: Protamine sulfate is the treatment of choice for reversal of heparin.

Major points of discussion

■ UFH has a half-life of 1 hour, so it takes 5 to 6 hours to be cleared.

■ UFH can be reversed by administering protamine, which is an arginine-rich (positively charged) protein that binds to the negatively charged heparin.

■ Dosing of protamine for heparin reversal: Administer 100 mg of protamine for each 100 IU/mL of heparin activity.

■ The main side effect of protamine is allergic reactions that can be severe (anaphylaxis). It can also cause hypotension, bradycardia, and hypertension/pulmonary hypertension.

■ Protamine is also used to reverse the anticoagulant effect of low-molecular-weight heparin, but it only partially reverses the effect of low-molecular-weight heparin.

40. A. 0.1
Rationale: The y-intercept is not used in the calculation of the ISI of a test reagent.
B. 0.90
Rationale: This is equal to the ISI + y-intercept, which is not the correct equation.
C. 0.95
Rationale: The ISI of the test reagent = ISI reference preparation × Slope of regression line.
D. 1.0
Rationale: The ISI of the test reagent would be 1.0 if the slope of the regression line were 1.0.
E. 1.1
Rationale: This is equal to the ISI of the reference preparation – the y-intercept, which is not the correct equation.

Major points of discussion

■ The sensitivity of each thromboplastin reagent to vitamin K–dependent coagulation factors can differ significantly from manufacturer to manufacturer and lot to lot. The INR was developed so that patients taking warfarin could have their warfarin therapy monitored by different laboratories.

■ Each thromboplastin should have an ISI determined by testing it against an international or working reference preparation, which are thromboplastins available in several different preparations (e.g., rabbit, human, recombinant) from the World Health Organization.

■ To determine the ISI of a reagent, the PT is performed using both the international reference preparation or working reference preparation on a group of normal donors and patients on stable warfarin therapy. A double logarithmic plot of a donor/patient’s PT using the reference and test reagent is plotted, and a linear regression is calculated. The ISI is determined in terms of this slope.

■ ISI of test reagent = ISI of the reference preparation × Slope.

■ In this example, ISI = 1.0 × 0.95 = 0.95.

■ The ISI is used to determine the international normalized ratio:

• Log (INR) = ISI × PT/ PT_GeoMean.

■ Warfarin skin necrosis typically appears 3 to 10 days after warfarin initiation if heparin is not overlapped. Lack of the overlap results in clot formation and necrosis of skin, which is more common on the penis, thighs, buttocks, and breasts. It is more common in obese, middle-aged women.

■ Purple toe syndrome is a rare complication appearing 3 to 8 weeks after initiation of therapy. Cholesterol emboli deposit in the skin of the feet, causing pain and the purple color.

■ Most adverse events associated with warfarin are related to hemorrhage, which is seen in 1% to 3% of patients treated with warfarin. There are many drug-drug interactions that may increase the risk for bleeding. Bleeding can be mild with bruising and epistaxis, or it can be severe with intracranial hemorrhage and stroke, gastrointestinal bleeding, intraabdominal bleeding, and compartment syndrome.

■ Osteoporosis and allergic reactions can also occur.

42. A. Pre-anhepatic.
Rationale: The pre-anhepatic stage is characterized by a mild deterioration in baseline coagulopathy.
B. Anhepatic to reperfusion.
Rationale: During this stage, there is a loss of coagulation factor synthesis and clearance as well as hyperfibrinolysis, which can lead to severe bleeding.
C. Postreperfusion.
Rationale: During the postreperfusion stage, there is resolution of fibrinolysis, restoration of coagulation factor synthesis and clearance, and a heparin-like effect from the donor liver.
D. Postoperative.
Rationale: Hypercoagulability during the postoperative stage is related to the early recovery of procoagulants, elevated factor VIII, and delayed recovery of AT, protein C, and protein S.
E. During surgical dissection and mobilization of the diseased liver.
Rationale: This refers to the pre-anhepatic stage. The pre-anhepatic stage is characterized by a mild deterioration in baseline coagulopathy.

Major points of discussion

■ The stages of liver transplantation are:

• Anhepatic to reperfusion (from occlusion of hepatic vasculature to reperfusion)

• Postreperfusion

• Postoperative

■ The hemostatic changes associated with each stage of transplantation have been described.

■ A mild deterioration in baseline coagulopathy is noted in the pre-anhepatic stage and is mostly related to the preexisting coagulopathy.

■ The anhepatic to reperfusion stage is associated with a loss of coagulation factor synthesis and clearance and hyperfibrinolysis that can lead to severe bleeding. There is also an increase in tPA that may be caused by reduced clearance and increased release from the ischemic endothelium of the donor liver.

■ During the postreperfusion stage, there is restoration of coagulation factor synthesis and clearance by the transplanted liver, resolution of hyperfibrinolysis, thrombocytopenia, and a heparin-like effect from the donor liver. The resolution of hyperfibrinolysis may be attributed to increased clearance of tPA and increased production of plasminogen activator inhibitor 1. Thrombocytopenia may be caused by sequestration of platelets in the sinusoids of the donor liver. And the heparin-like effect from the donor liver is associated with heparinization of the donor before harvest and the release of heparinoids from the ischemic donor liver endothelium.

■ Heparin is a cofactor that accelerates the binding of AT to thrombin and factor Xa.

■ AT deficiency is associated with an increased risk for VTE.

■ AT deficiency can be inherited or acquired (e.g., liver disease, sepsis, nephrotic syndrome, cardiothoracic surgery/bypass).

■ The pattern of inheritance is autosomal dominant.

■ Dysfibrinogenemias classically show an elevated thrombin time and an elevated reptilase time. An elevated thrombin time and normal reptilase time would suggest the presence of heparin.

■ Clinically, approximately half of the patients with dysfibrinogenemias are asymptomatic, and the other half present with bleeding or thrombosis, or both.

■ APCR is an inherited thrombophilia caused by an abnormal factor V that is resistant to inactivation by APC.

■ The factor V Leiden (FVL) gene mutation is the most common cause of APCR. Approximately 5% of North American Caucasians carry the FVL gene.

■ In patients with an FVL mutation, APC fails to inactivate factor Va because of a highly conserved point mutation (G1628A), leading to an amino acid substitution (R485K) in factor V, leading to APCR and an increased risk for thrombosis.

■ The factor V Liverpool variant is caused by a point mutation (T1250C), resulting in an amino acid substitution (I359T) that also leads to APCR and an increased risk for thrombosis.

■ HCII requires heparin, heparan sulfate, or dermatan sulfate as a cofactor.

■ HCII appears to act as an adjunct to AT, and evidence suggests that its deficiency alone does not lead to an increased risk for thrombosis; however, it may increase the risk for thrombosis when there is a concomitant AT deficiency.

■ HCII activity can be measured in vitro by measuring its anti–factor IIa activity in the presence of dermatan sulfate. Alternatively, HCII can be measured antigenically using an ELISA-based method.

■ The procoagulant factors produced by the liver include factors II (prothrombin), VII, IX, X, XI, XII, V, and VIII, plasminogen activator inhibitor 1 (PAI-I), α₂-antiplasmin, and thrombin activatable fibrinolysis inhibitor (TAFI).

■ The anticoagulant factors produced by the liver include AT, protein C, protein S, TFPI, and plasminogen.

■ Factor V, factor VIII, PAI-1, AT, protein C, protein S, and TFPI are also synthesized by extrahepatic sites.

■ Hemostatic changes found in liver disease lead to rebalancing of the coagulation system; reduction of procoagulant and fibrinolytic factors is offset by a concomitant decrease of anticoagulant and antifibrinolytic proteins.

■ The procoagulant factors produced by the liver include factors II, VII, IX, X, XI, XII, V, and VIII, PAI-I, α₂-antiplasmin, and TAFI.

■ The anticoagulant factors produced by the liver include AT, protein C, protein S, TFPI, and plasminogen.

■ Factor V, factor VIII, PAI-1, AT, protein C, protein S, and TFPI are also synthesized by extrahepatic sites.

■ Hemostatic changes found in liver disease lead to rebalancing of the coagulation system; reduction of procoagulant and fibrinolytic factors is offset by a concomitant decrease of anticoagulant and antifibrinolytic proteins.

■ Patients with liver disease therefore may be at risk for both bleeding and thrombotic complications.

49. A. Failure to generate APC.
Rationale: This is the mechanism of activation associated with protein C deficiency.
B. Failure to inhibit factor IIa (i.e., thrombin), factor Xa, and other activated factors.
Rationale: This is the mechanism of thrombosis associated with AT deficiency.
C. Failure to activate plasminogen.
Rationale: This is the mechanism of thrombosis associated with tPA deficiency.
D. Endothelial cell toxicity and disruption of vascular hemostatic mechanisms.
Rationale: This is the mechanism of thrombosis associated with hyperhomocysteinemia.
E. Failure of APC to inactivate factors Va and VIIIa.
Rationale: This is the mechanism of thrombosis associated with protein S deficiency.

Major points of discussion

■ Homocysteine is an intermediate amino acid formed during the conversion of methionine to cysteine; the enzyme cystathionine-β-synthase is required.

■ Moderate elevations of homocysteine may be a risk factor for atherothrombotic disease and VTE.

■ Elevation of homocysteine can be caused by inherited defects of metabolism, nutritional deficiencies of vitamin cofactors, acquired diseases, and medications.

■ The procoagulant factors produced by the liver include factors II, VII, IX, X, XI, XII, V, and VIII, PAI-I, α₂-antiplasmin, and TAFI.

■ The anticoagulant factors produced by the liver include AT, protein C, protein S, TFPI, and plasminogen.

■ Factor V, factor VIII, PAI-1, AT, protein C, protein S,and TFPI are also synthesized by extrahepatic sites.

■ Hemostatic changes found in liver disease lead to rebalancing of the coagulation system; reduction of procoagulant and fibrinolytic factors is offset by a concomitant decrease of anticoagulant and antifibrinolytic proteins.

■ Patients with liver disease therefore may be at risk for both bleeding and thrombotic complications.

■ tPA activates plasminogen to plasmin. Plasmin degrades fibrinogen and fibrin to its degradation products.

■ Excess PAI-1 activity decreases tPA levels and decreases plasminogen activation, leading to decreased plasmin and decreased fibrinolysis.

■ Excess PAI-1 activity can lead to arterial and venous thrombotic disorders, such as VTE and myocardial infarction.

■ Plasmin degrades fibrinogen and fibrin with the primary role of dissolving thrombi.

■ Both homozygous and heterozygous forms of plasminogen deficiency have been described; they can be qualitative defects (active site mutations; type II) or quantitative defects (type I). It is a very rare disorder.

■ Patients can be asymptomatic or present with a history of thrombosis. They can also have ligneous conjunctivitis with pseudomembranes covering the eyes (other organs can also be affected). The pseudomembranes are caused by the accumulation of fibrin and are more commonly found in patients with type I deficiencies.

■ Thrombomodulin on endothelial surfaces accelerates the activation of protein C to APC.

■ APC, along with its cofactor protein S, inactivates factors Va and VIIIa.

■ Protein C deficiency is a rare inherited disorder that is associated with an increased risk for VTE. Type I defects are quantitative defects of normal protein C and type II defects are qualitative defects.

■ Protein S circulates bound to C4 binding protein (C4bp) or as free protein S (Total protein S = Free protein S + Protein S:C4bp).

■ Approximately 65% of protein S is bound to C4bp, and the remainder is free protein S. Only the free form has cofactor activity.

■ Protein S deficiency is an inherited, autosomal dominant disorder that is a risk factor for VTE. However, inherited protein S deficiency is not common.

■ Assays for protein S include total protein S antigen, free protein S antigen, and total protein S activity. Antigenic assays are immunologically based, whereas activity is measured by clot-based assays using platelet poor plasma.

■ Type I protein S deficiency is a quantitative disorder of normal protein S. Type II is a qualitative disorder of quantitatively normal protein S. Type III is a disorder of normal total protein S, but decreased free protein S and increased bound protein S.

■ tPA converts plasminogen to plasmin. Plasmin functions to degrade fibrinogen and fibrin.

■ Recombinant tPA is a pharmaceutical agent that is used therapeutically for acute ischemic cerebrovascular events, pulmonary embolism, and other thrombotic disorders.

■ Increased endogenous tPA activity can lead to excessive fibrinolysis and bleeding.

■ tPA deficiency can lead to hypofibrinolysis and a risk for thrombosis.