Coagulation Cascade and Coagulation Tests

Related synonyms: Bleeding diathesis, clotting disorder, coagulation disorder, coagulopathy, haemostatic disorder

When a blood vessel is injured, a series of biochemical reactions is brought into play, called the coagulation cascade. The result of this series of reactions is to achieve haemostasis by developing a clot, stopping its formation at the right time, and eventually facilitating clot dissolution when the vessel has healed.

Most of the proteins required for the cascade are produced by the liver as inactive precursors (zymogens) which are then modified into clotting factors. The implication of the term 'cascade' is that a small stimulus can produce a reaction which is sufficiently widespread to produce a significant clot.
There are two routes for activation of the coagulation system. The intrinsic pathway is activated by contact with collagen from damaged blood vessels (or indeed any negatively-charged surface). Platelet activation is normally involved. The extrinsic pathway is activated by contact with tissue factor from the surface of extravascular cells.

Both routes end in a final common pathway - the proteolytic activation of thrombin and the cleaving of fibrinogen to form a fibrin clot. The intrinsic pathway is the main 'player' in this scenario, with the extrinsic pathway acting as an enhancer.

When a patient presents with symptoms which might relate to an abnormality of the haemostatic system it is essential to take a good history. Ask about:

• Bruising
  • Spontaneous or recurrent
  • Large bruises on sun-exposed areas of limbs in the elderly are usually due to cumulative UV vessel damage (rarely significant)³
  • Large bruises on trunk are more indicative of a bleeding disorder
• Prolonged bleeding e.g.
  • After minor cuts/abrasions
  • Nosebleeds lasting >10 minutes despite compression
  • Severe menorrhagia (causing anaemia)
  • Following dental extraction
  • Post-partum
  • After injections or surgical procedures
• Anaemia of unknown cause or recurrent anaemia of known cause
• Any episodes of unusual spontaneous bleeding e.g. haemarthrosis
• Current medication
  • Including aspirin, NSAIDs, warfarin and complementary/alternative preparations
  • Remember drug interactions between warfarin and other medications that prolong the INR
• Family history of bleeding tendency
• Alcohol intake
• Other constitutional symptoms e.g. malaise, weight loss
• Past history or thrombosis (can be suggestive of thrombophilia)

• Temperature
• Haemodynamic status - pulse, blood pressure
• Examine skin closely
  • For signs of petechiae and purpura
  • Skin creases or where clothing exerts pressure on skin
• Examine mouth for signs of bleeding around gums (nearly always pathological in absence of severe gum disease) or opportunistic oral infection e.g. Candida
• Check sclera for evidence of petechiae conjunctivae for anaemia and consider fundoscopy to look for retinal haemorrhage
• Check axillary, cervical and groin nodes for evidence of lymphadenopathy
• Abdominal examination looking for splenomegaly and/or hepatomegaly
• Check joints if suspect haemarthrosis
• If suspect acute bleeding diathesis consider per rectum or per vaginam examination to look for occult bleeding

Examples of when bleeding tendency should be investigated are

• Any internal haemorrhage not related to trauma e.g. gastrointestinal bleed
• Prolonged bleeding of any cause
• Recurrent haemorrhage e.g. bleeding gums, epistaxis
• Recurrent spontaneous bruising
• Haemorrhage into unusual places e.g. joints (with no explanation)
• Acutely unwell patients in whom disseminated intravascular coagulation (DIC) is suspected e.g. septicaemia

• FBC, blood film and platelet count - may detect leukaemia, lymphoma or thrombocytopenia or abnormal platelets⁶
• Consider checking U&E to exclude uraemia causing a platelet disorder
• Consider LFTs to detect hepatic cause of acquired coagulation factor deficiency (and ethanol damage)
• Bone marrow biopsy

Investigation of bleeding tendency⁷

A coagulation screen usually involves taking blood in a mixture of citrate, EDTA and clotted sample bottles.

• Activated partial thromboplastin time (APTT) - measures the intrinsic pathway (which includes factors I, II, V, VIII, IX, X, XI, and XII) and the common pathway. It shows possible coagulation factor deficiency as in haemophilia A & B. A plasma sample is used and the intrinsic pathway is activated by adding phosopholipid, an activator such as kaolin (which acts as a negatively-charged surface), and calcium ions. The formation of prothrombinase complexes on the surface of the phospholipid enables the formation of thrombin and a subsequent clot. The result is reported as the time in seconds for this reaction. The test is a used to assess the overall competence of the coagulation system, as a routine test to monitor heparin therapy, and as a preoperative screen for bleeding tendencies.
• Prothrombin time (PT) - this assesses the extrinsic and final common pathway of the coagulation cascade thus can detect factor I, II, V, VII or X deficiency or the effects of warfarin. It is performed by adding thromboplastin and calcium ions to a plasma sample. The time for clot formation is measured. Prolonged time suggests the presence of an inhibitor to, or a deficiency of one or more coagulation factors, the presence of warfarin, the existence of vitamin K deficiency or liver dysfunction. The International Normalised Reaction (INR), used to monitor warfarin, is derived by comparing the patient's clotting time to that of a standardised sample.
• Thrombin clotting time test - this measures the rate of a patient's clot formation compared to a normal plasma control. The plasma is first depleted of platelets, and a standard amount of thrombin added. The test is used in the diagnosis of disseminated intravascular coagulation (DIC), and other conditions that can affect fibrinogen level such as liver disease.

If the above tests are all normal then the vast majority of common haemostatic disorders will have been excluded. However, if symptoms persist and/or there is a suggestion of family history, refer to haematologist for further tests, e.g.:

• Bleeding time - this tests the interaction between the platelets and the vessel walls. A standardised spring-loaded lancet is used to make a small cut in the patient's forearm and the time for the bleeding to stop is then measured. The test is not useful as a screening test as it has a high false positive result, but is sometimes useful in certain clinical situations, e.g. the investigation of von Willebrand's Disease and the effect of drugs on the coagulation system.⁸
• A new investigation that may replace the in-vivo bleeding time test is being developed, the platelet function analyser.⁹
• Fibrinogen - the level can be determined by immunological or functional assay. it is usually performed when APTT or PT screening tests are prolonged. The main disorders detected are afibrinogenaemia or hypofibrinoginaemia (due to absence or a low level of fibrinogen production) and dysfibrinoginaemia (due to a molecular alteration of fibrinogen, causing poor function). Differences in the level of fibrinogen measured by the two methods is suggestive of dysfibrinogenaemia.¹⁰
• D-dimers - this test is used in the diagnosis of deep vein thrombosis. High concentrations also occur in malignancy and pregnancy.

Remember that some diseases can be associated with both bleeding and thrombosis e.g. polycythaemia rubra vera and essential thrombocythaemia.¹¹

Investigation of thrombophilia^12,13

Because of the difficulty in interpretation of results and the need to provide specific advice and management for each individual patient, the current guidance suggests that only physicians with a specialist knowledge should undertake screening tests for hereditary thrombophilia, and only after appropriate counselling of the patient.¹⁴

• When an individual is thought to have a hereditary form of thrombophilia, it is now possible to perform a range of tests in order to determine the precise mutation.
• It has been suggested that the range of tests performed should include, Factor V Leiden, prothrombin G20210A, protein C, protein S and anti-thrombin III deficiency mutations.
• Although the polymerase chain reaction for the presence of the factor V Leiden mutation is 99% accurate, the other tests are less so, and are influenced by various factors such as age, gender, pregnancy, the pill or HRT and use of anti-coagulants.
• Other appropriate screening tests include:
  • Clotting screen: prothrombin time, APTT, raised fibrinogen, raised prothrombin, raised Factor VIII, plasminogen, Factor XII
  • Full blood count and film: myeloproliferative disorders, paroxysmal nocturnal haemoglobinuria, thrombocytosis, polycythaemia
  • ESR and/or CRP, antinuclear antibodies: to screen for possibility of connective tissue disorders
  • Homocysteine levels
  • Investigations for cardiac disease, liver disease, nephrotic syndrome and other causes acquired thrombophilia as appropriate
  • Consider occult malignancy and investigate appropriately
• Some believe that thrombophilia testing should not routinely be performed on young people who have already suffered a thrombosis as the results are unlikely to change the management. They may however be appropriately used in those patients with a strong family history of venous thrombosis, or a history of recurrent miscarriage.