Understanding Lupus Anticoagulant and APTT Testing

Blood Homeostasis and Coagulation (MCLS7211)

• This course covers blood homeostasis and coagulation, part of a master's program in Clinical Laboratory Sciences.
• The material is from the first semester of 2024/2025.
• The lecturer is Dr. Ola Karmi, with email address: [email protected].

Heritable Thrombophilia (Lecture 9)

• This lecture discusses heritable thrombophilia, including testing for factor V Leiden deficiency.
• Testing includes diluting the sample with factor-deficient plasma for APC sensitivity ratio testing.

APC Resistance Testing

• APC/SR test involves diluting the test sample using FV-deficient plasma;
• APC/SR is calculated as (aPTT of sample + APC (sec) / aPTT of sample without added APC (sec)).
• Confirmation is done through DNA analysis.

APC Resistance (Functional Assay)

• This is a modified APTT assay to assess APC resistance.
• Patient PPP is mixed with FV-deficient substrate to enhance sensitivity.
• The modified APTT is measured after adding a specific amount of APC, it should be prolonged.
• Genetic testing (PCR analysis of FV mutations) is part of the diagnosis.

Heritable APC Resistance (Not Due to FVL)

• Caused by rare mutations (Arg306Thr and Arg306Gly).
• A gene haplotype (HR2) mutation (His1299Arg) exists
• Co-inheritance of HR2 and FVL increases VTE risk by 3-4 folds.

Von Willebrand Disease vs. Factor V Leiden

• Von Willebrand Disease:
  • Recurrent bleeding episodes
  • Family history
  • Increased bleeding time
  • Normal or prolonged aPTT
  • Normal PT
  • Normal platelet count
  • Ristocetin cofactor assay shows no platelet aggregation
  • Cofactor level <30 IU/dL
• Factor V Leiden:
  • Unprovoked thromboembolic events
  • Family history of thromboembolism
  • Normal bleeding time, aPTT, PT, and platelet counts
  • Positive Activated protein C resistance assay
  • Genetic testing for Factor V Leiden

Prothrombin G20210A Mutation

• A single nucleotide change (G20210A) in the 3'UTR of the prothrombin gene.
• Found in 2% of the general population, 6% with first VTE episode, and up to 18% with thrombosis history.
• Double heterozygosity with FVL is uncommon (1 in 1000 general population and 1-5% with VTE).

Prothrombin G20210A Mutation (Continued)

• Heterozygosity is linked with 25% higher prothrombin levels in plasma.
• Increased prothrombin plasma levels and a higher risk of VTE
• Elevated FII levels increase VTE risk
• Three fold risk increase for VTE (G20210A mutation)
• DNA analysis (PCR) is used for diagnosis.

A-RFLP-PCR for FII G20210A

• A diagnostic technique identifying the G20210A mutation using polymerase chain reaction and restriction fragment length polymorphisms.

Dysfibrinogenemia

• A rare condition (0.8% of VTE patients, but high postpartum thrombosis incidence)
• Defective thrombin binding to fibrin results in elevated thrombin levels in plasma.
• Diagnosis is based on discordance between functional and immunological fibrinogen assay results – functional activity is notably lower than immunological values in patients with a prolonged thrombin time.

Mixed Etiology Thrombophilia

• Elevated FVIII levels are a risk factor for venous and arterial thrombosis, especially in non-O blood groups.
• Elevated FVIII (>1500 IU/L) is an independent risk factor for VTE.
• Increased levels of FIX and FXI above the 90th percentile for the general population also increase VTE risk.

Thrombophilia Screening

• Clinical Assessment:
  • Personal history
  • Family history
  • Clinical examination
• Risk factor search:
  • Age, immobility, trauma, surgery, hormone use, pregnancy, and obesity.
• Lab tests:
  • Complete blood count (CBC) and platelet counts
  • Liver function tests
  • Imaging and further investigations.

Thrombophilia Screening (Continued)

• First-Line Tests: Coagulation screen (APTT, PT, TT), AT activity, PC, and PS.
• Modified Tests: APC/SR, FV Leiden, FII G20210A, lupus inhibitor screen, anticardiolipin assays.
• Second-Line Tests: FVIII levels, homocysteine, fibrinogen levels.

Diagnostic Algorithm for Thrombophilia Screening

• A multi-step approach for diagnosing thrombophilia in patients with VTE.

Lecture 10: Acquired Thrombophilia

• This lecture focuses on acquired (rather than inherited) blood clotting disorders.

Epidemiology of Venous Thromboembolism (VTE)

• Deep vein thrombosis (DVT) is common, predominantly affecting the lower limbs.
• Annual incidence in Western populations is around 1 per 1000 people.
• VTE incidence is age-dependent, increasing with age, with significant rates in the elderly.

Pregnancy and Venous Thromboembolism (continued)

• Pregnancy increases risk of VTE, with significant instances in the postpartum period, but also occurring in the antepartum phase, including the first trimester.
• Several factors contribute to this risk, including obesity, increased intra-abdominal pressure, decreased fibrinolysis from the placenta, and increased clotting factors.
• Delivery, especially emergency caesarean sections, further increase this risk

Immobility and VTE

• Prolonged immobility, whether from surgery, illness, or prolonged sitting, greatly increases VTE risk.
• Studies have associated prolonged sitting (even in office settings) with venous thrombosis.

Hospital-Acquired Thrombosis and Iatrogenic VTE

• Hospital-acquired thrombosis (HAT) can occur within 90 days of admission, often linked to factors like immobility and/or procedures.
• Indwelling devices, infections, and certain chemotherapy drugs contribute to VTE risk
• Combined oral contraceptives and Hormone Replacement Therapy (HRT) are linked to VTE risk.

Antiphospholipid Syndrome

• This is an acquired thrombophilia affecting both arterial and venous systems.
• Characterized by recurrent thrombosis or pregnancy complications.
• Presence of antiphospholipid antibodies (tested via the lupus anticoagulant test among other tests and markers)

Venous Thromboembolism and Cancer

• A well-established link exists between cancer and venous thromboembolism (VTE).
• Cancer-related prothrombotic tendencies include tissue factor expression, elevated prothrombin levels, and tumour cell-associated factors.

Thrombotic Risk in Myeloproliferative Diseases

• Malignant diseases may increase the risk of thrombosis.
• Polycythemia vera (PV) and essential thrombocythemia (ET) result in vascular occlusions (PV untreated and likely in ET).
• Acute promyelocytic leukaemia (APL) sometimes exhibits arterial thrombosis, and the use of all-trans retinoic acid might increase the risk.

Inflammation and Thrombosis

• Inflammation modifies coagulation factors. It activates procoagulants while down-regulating anticoagulants and fibrinolysis (by activating PAI-1).
• Inflammation factors also increase platelet counts and stimulate fibrinogen levels.

Haematological, Non-Malignant Conditions and Thrombosis

• A range of conditions associated with blood disorders including PNH, TTP, sickle cell disease influence thrombosis risk.
• In PNH, thrombosis occurs often in certain vascular sites such as the hepatic, portal veins, and cerebral veins.
• Sickle cell disease is characterized by sickled red blood cell clustering, thrombosis, and often increased prothrombin levels.

This quiz explores the implications of lupus anticoagulant on clot formation, its characteristics, and the relationship with APTT in blood testing. Additionally, it addresses issues related to indwelling venous devices and postoperative patient care. Test your knowledge on this complex topic in hemostasis and thrombosis.