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The following is the complete blood count (CBC) at the time of admission:

WBC (109/L)Hb (g/dL)MCV (fL)PLT (109/L)
9.77.492259

What’s what: WBC, white blood cell count; Hb, hemoglobin; MCV, mean cell volume; MCHC, mean cellular hemoglobin concentration; RDW-SD, red cell distribution width-standard deviation; platelets, PLT; Normal values: WBC 5-10 x 109/L, RBC 4-6 x 1012/L, Hb 12-16 g/dL, Hct 35-47%, MCV 80-100 fL, MCHC 32-36 g/dL, RDW-SD < 45 fL, platelets (PLT) 150-450 x 109/L

PT and aPTT results:

PT (seconds)aPTT (seconds)
13.881.6
What’s what: PT, prothrombin time; aPTT, activated partial thromboplastin time; Normal values: PT 9.4-12.5 seconds, aPTT 25-36.5 seconds

PT is minimally elevated. For all intents and purposes, we can approach this case as an isolated elevation in the aPTT.

Let’s look at how a PT and aPTT are carried out:

Notes:

  • Step 1 – Draw whole blood from a blood vessel (typically an arm vein) into a blue top tube containing the anticoagulant, sodium citrate.
  • Step 2 – Spin the liquid blood sample in a centrifuge so that red cells layer on the bottom, plasma on the top.
  • Step 3 – Pipette plasma (top layer) into a clean, empty test tube.
  • Step 4 – Add phospholipid, calcium and activator (tissue factor for PT, silica for aPTT).
  • Step 5 – Incubate at 37 degrees C and measure time to clot formation, either with machine or more rarely by vision.

Note that the major difference between the PT and aPTT is the nature of the activator added to the sample.

What are possible causes of an isolated elevation in the aPTT (i.e., the PT is normal)?

a
Factor X deficiency
This would be expected to increase both the PT and aPTT.
b
Lupus anticoagulant
Yes, a lupus anticoagulant typically elevates the aPTT alone. The PT test includes excess phospholipid, which may quench the LA.
c
Factor V deficiency
This would be expected to increase both the PT and aPTT.
d
Factor VIII deficiency
Correct. Factor VIII is in the intrinsic pathway, and its deficiency causes an isolated increase in the aPTT.
e
Factor VIII inhibitor
Correct. Factor VIII is in the intrinsic pathway, and an inhibitor against this clotting factor causes an isolated increase in the aPTT.

Back to the patient. He presents with bilateral arm swelling and a high aPTT. What are the two most likely causes of the arm swelling?

a
Bilateral arm infiltration with cancer
b
Lymphedema
c
Superior vena cava syndrome (SVC syndrome)
d
Bilateral upper extremity DVT
Yes, this might be seen in a patient with lupus anticoagulant, though bilateral clots in the upper extremity would be highly unusual.
e
Bilateral upper extremity hematoma
This seems like the most likely possibility given that the findings are bilateral and there is extensive bruising.

A Doppler ultrasound of the upper extremities was ordered in the emergency room.

These are the results:

Therefore, the swelling in the arms is most certainly secondary to hematoma.

Further work-up of this case requires some knowledge of the clotting cascade. Let’s review the basics:

Notes:

  • The bottom line is formation of an insoluble fibrin plug, which, along with platelets, stems blood loss.
  • Fibrin is derived from thrombin-mediated cleavage of soluble fibrinogen (which is a structural protein).
  • Thrombin (which is a type of enzyme called a serine protease) is formed when another serine protease, activated factor X (FXa), cleaves prothrombin.
  • Two pathways may activate factor X:
    • The intrinsic pathway, which consists of a series of linked reactions involving serine proteases FXII, FXI and FIX, and cofactor FVIII.
    • The extrinsic pathway, which consists of tissue factor-mediated activation of FVII (FVIIa).
  • In vivo, the clotting cascade is always initiated by tissue factor activation of FVII (extrinsic pathway) and amplified by the intrinsic pathway via cross-talk (FVIIa activates FIX) and feedback (thrombin activates FXI and FVIII) mechanisms.
  • aPTT measures the integrity of the intrinsic pathway.
  • PT/INR measures the integrity of the extrinsic pathway.

According to the scheme above, an isolated elevation in the aPTT indicates a deficiency of or inhibitor against a clotting factor in the intrinsic pathway, namely FXII, FXI, FIX or FVIII.

Heparin is often considered to be a cause of isolated aPTT elevation. Heparin inhibits components of the intrinsic, extrinsic and common pathways, so in theory it should raise both the PT and aPTT. In fact, heparin does increase the PT, but the degree of change is small (5-10%) and at therapeutic doses barely noticeable:

Pharmacotherapy. 1991;11:312-6.

This patient provides no history of heparin exposure. But there are cases, especially in hospitalized patients, where accidental or unidentified heparin exposure may be an issue.

Let’s consider each assay in turn:

Thrombin time

  • Measures the time required for formation of a stable clot, following addition of thrombin to citrated plasma.
  • Provides the measure of conversion of fibrinogen to fibrin and therefore an estimation of the quantity of fibrinogen.
  • Causes of prolonged thrombin time include:
    • Afibrinogenemia
    • Hypofibrinogenemia
    • Dysfibrinogenemia
    • Factors that inhibit thrombin-mediated conversion of fibrinogen to fibrin:
      • Heparin
      • Hirudin
      • Other direct thrombin inhibitors
      • Autoantibodies, for example in multiple myeloma
      • Fibrin degradation products, for example in disseminated intravascular coagulation (these inhibit thrombin-mediated cleavage of fibrinogen)

Reptilase time

  • A venom-based functional plasma clotting assay based on the enzymatic activity of batroxobin, the thrombin-like proteolytic enzyme isolated from the venom of the South American pit viper.
  • Unlike thrombin, which cleaves fibrinopeptides A and B from fibrinogen, batroxobin splits specifically fibrinopeptide A, converting fibrinogen into fibrin monomers. The resulting fibrin monomers polymerize spontaneously and lead to the formation of a stable fibrin clot.
  • The time, starting from the addition of batroxobin to the plasma sample, until clot formation is the reptilase time and is given in seconds.
  • The reptilase time is independent of thrombin generation disturbances or disturbances in the action of thrombin on fibrinogen. Therefore, it can be used to confirm heparin contamination or to obtain similar information as with the thrombin clotting time in heparinized and hemophiliac patients.
  • The clotting time obtained by RT is independent of thrombin generation disturbances and disturbances of thrombin–fibrinogen interaction and is not affected by anticoagulants such as heparin and hirudin.
  • Causes of prolonged reptilase time include:
    • Afibrinogenemia
    • Hypofibrinogenemia
    • Dysfibrinogenemia
    • Factors that inhibit conversion of fibrinogen to fibrin:
      • Especially fibrin degradation products
      • Not sensitive to antithrombin inhibitors

Source.

Which of the following is true (more than one answer may apply)?

a
An elevated reptilase time indicates presence of heparin
b
An elevated thrombin time is diagnostic of heparin contamination
c
An elevated thrombin is consistent with the presence of heparin
d
An elevated thrombin time together with an elevated reptilase time indicates presence of heparin
e
An elevated thrombin time together with a normal reptilase time is highly suggestive of the presence of heparin

Back to the patient. What would you like to test for next?

a
von Willebrand factor (vWF)
While von Willebrand disease (VWD) may increase the aPTT because of reduced FVIII levels (VWF carries and stabilizes FVIII), VWD would be unlikely to prolong the aPTT to this degree.
b
Clot lysis time
This test is used to assess the fibrinolytic system.
c
Mixing study
Correct. This is a quick screening test for the presence of clotting factor deficiency vs. inhibitor, including LA.
d
Lupus anticoagulant (LA)
While it might be reasonable to test for LA, it is not a priority in a patient who is hemorrhaging into his arms with an elevated aPTT. LA is associated with thrombosis, not bleeding.

A mixing study is a reasonable next step in a patient with unexplained elevation in PT and/or aPTT. What is a mixing study?

a
aPTT of 1 part patient plasma mixed with 3 parts normal pooled plasma
b
aPTT of 1 part patient plasma mixed with platelet rich plasma
c
Mix of PT and aPTT
d
aPTT of 1 part patient plasma mixed with 1 part normal pooled plasma
Correct! See next slide for details.

Let’s review the basics of the mixing study:

Notes:

  1. Mixing study involves mixing patient plasma and pooled normal plasma in 1:1 ratio.
  2. The resulting mixture is then processed for clotting time (PT or aPTT) immediately (time 0) and then after 2 hours of incubation.
  3. A clotting factor must drop below about 30% before the PT or PTT begins to prolong.
  4. Therefore, even if a clotting factor is completely missing (i.e. 0% level), the 1:1 mix will restore it to 50% (equal volume of plasma containing 0% of the factor and normal pooled plasma containing 100% of the factor), more than enough to restore a normal PT or aPTT (the example above shows 10% activity of imaginary factor Y in patient plasma, with 1:1 mix resulting in 60% activity).
  5. However, if there is an inhibitor or antibody to a clotting factor, the inhibitor will carry over from patient to normal plasma in the mix and interfere with factor activity, leading to continued prolongation of the PT or aPTT.

Mixing study (50:50 mix) results

(baseline aPTT 76.7, PT 13.1)


What is the result of the above mixing study?

a
Negative for inhibitor
b
Positive for inhibitor
Correct. The failure of the aPTT to correct with 1:1 mix indicates the presence of a factor inhibitor.

Mixing study (50:50 mix) results


If the PT, and not the aPTT was prolonged and the prolongation failed to correct with the 1:1 mix, what factor would you predict is being inhibited?

a
Factor II (prothrombin)
b
Factor V
c
Factor VII
The extrinsic pathway, which is measured by the PT, consists of factor VII alone.
d
Factor XII
e
Fibrinogen

To summarize at this point, we have an elevated aPTT (and normal PT) in a patient with recent rapid onset of bilateral arm swelling with ecchymoses, and a positive inhibitor screen (mixing study), indicating the presence of an inhibitor against one or more factors (almost certainly a single factor) in the intrinsic pathway.

Which factor level would you prioritize investigating?

a
Factor XIII (FXIII) level
FXIII functions to crosslink fibrin. Because the end point of clotting times does not require that fibrin be crosslinked, even severe deficiency in FXIII is not associated with a change in the PT or aPTT.
b
Factor XII (FXII) level
FXII deficiency does not confer an increased risk of bleeding.
c
Factor VIII (FVIII) level
We are considering an inhibitor against a factor in the intrinsic pathway. Inhibitors are most common against FVIII.
d
Factor IX (FIX) level
Inhibitors against FIX are much rarer than those against FVIII.

the patient’s factor VIII level was <1% (normal 50-150%)

What is the diagnosis?

a
Hemophilia A
b
Hemophilia B
c
Acquired von Willebrand syndrome
d
Acquired hemophilia A
Correct. We use the term “acquired hemophilia A” to describe the presence of an acquired inhibitor against factor VIII.

Are you surprised there is no family history of bleeding?

a
Yes
b
No
Correct. Acquired hemophilia A is by definition not inherited.

About 50% of cases of acquired hemophilia A are secondary to an underlying medical condition. Which of the following conditions may be associated with acquired hemophilia A?

a
Autoimmune disorders (e.g. systemic lupus erythematosus)
b
Malignancy
Both solid tumors and hematological malignancies.
c
Pregnancy
d
Hypothyroidism
e
Drugs

Acquired and congenital hemophilia A have a similar bleeding phenotype, with one notable exception.

Let’s consider the anemia for a moment. The patient’s Hb is 7.3

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