Hemostasis Flashcards
Identify the elements that compose the hemostatic system. Understand the basic paradigm for coagulation factor activation
Components:
Coagulation factors (13 known)
Platelets
Endothelial wall (site of injury)
General idea: Endothelial wall becomes damaged, exposing collagen, tissue factor, and von Willebrand factor. (vWF, if you’re interested, is produced constitutively in endothelia and binds platelets once the endothelium is damaged. Also binds Factor VIII and keeps it viable.)
Platelets bind to the site of injury and change configuration, exposing certain proteins that attract fibrinogen and vWF (thus binding more platelets). They also release granules containing thromboxane (leading to vasoconstriction of damaged vessel), vWF and ADP (to bind and activate other platelets), various clotting factors, etc. They also serve as a ground on which clotting factors can easily activate.
Note that the endothelium is normally actively preventing coagulation; prevents platelet aggregation, promotes clot breakdown. As with so many other things, it’s a balancing act. When the endothelium is breached, the balance shifts (towards pro-coagulation) until it’s resolved.
Describe the blood clotting pathway. List which components of the system are vitamin K dependent factors. Distinguish the extrinsic and intrinsic pathways and describe the screening tests to measure both (PT and APTT, respectively)
A few notes about the components of the clotting pathway:
Several clotting factors are called by name, most are called by numeral. An “a” after the numeral indicates that the factor has been activated and is enzymatic.
By name: Prothrombin (factor II), Fibrinogen (factor I). These, when activated, are called thrombin and fibrin respectively.
By numeral: Factors III-XIII.
Several clotting factors are large enzymes that serve to orient the others.
These are: Factors V and VIII. Tissue factor also serves this role.
Most of the rest of the clotting factors are proenzyme serine proteases which, when cleaved and activated, cleave and activate each other.
These are: Factors II, VII, IX, X, XI, and XII.
The others seem to have particular, difficult-to-generalize-about roles. For example, Factor XIII cross-links fibrin fibers to make a ‘hard clot.’
Some clotting factors require carboxylation in the liver by vitamin K (recall from the MCAT that it’s a fat-soluble vitamin stored in the liver). If there’s no vitamin K available or there’s a lot of liver damage, these factors may not function correctly. Note that she emphasized this a lot in lecture.
Vitamin K-dependent factors: Factors II, VII, IX, and X.
Note that several important anti-clotting factors are also vitamin K-dependent: proteins C and S.
So the end goal of clotting is to produce, cross-link, and harden a mat of fibrin fibers to plug the endothelial damage. There are two pathways, the extrinsic and the intrinsic, to get things set up to do this; both of them end by activating Factor X to feed into the same clot formation mechanism, the so-called “common pathway.”
Extrinsic pathway (Factor VII-mediated, triggered by release of tissue factor from damaged endothelium):
Tissue factor serves as a co-factor with Factor VIIa (it’s not clear what activates VII) to activate Factor X to Xa.
Note VIIa can activate elements in the intrinsic pathway as well (XI, IX).
Intrinsic pathway (Factor VIII-mediated, Factor VII and tissue factor independent):
Factor XII is activated (unclear what does this).
XIIa activates XI.
XIa activates IX.
IXa, with Factor VIII as its co-factor, activates Factor X to Xa.
Common pathway (Factor V-mediated):
Factor Xa, oriented by Factor V, cleaves prothrombin (Factor II) to thrombin (Factor IIa).
Thrombin cleaves fibrinogen (I) to fibrin (Ia), a soluble fiber that begins to polymerize.
Factor XIII cross-links fibrin polymers to form a hard, insoluble clot.
And that’s clotting. To reiterate:
Extrinsic: tissue factor + VIIa -> Xa.
Intrinsic: XIIa -> XIa -> IXa + VIII -> Xa.
Common: Xa -> IIa -> Ia + XIII -> clot.
Several notes:
Notice that many steps in these pathways require Ca2+. No calcium, no (or little) clotting.
Fibrin clots binds the platelets at the site together and attaches them more firmly to the vessel wall.
Explain how thrombin acts as the central regulation point for coagulation. Describe normal homeostasis in the coagulation system
Thrombin (IIa) is the central regulation point for coagulation. Activates platelets, cleaves fibrinogen to form fibrin. It also activates Factors VIII and V, which further accelerates the clotting cascade (recall that these are the two large orientation co-factors).
Normal hemostasis involves a certain amount of regulation of thrombin. There are anti-thrombin proteins that normally circulate to bind free thrombin to prevent its being activated. Thrombin also activates another anti-coagulation protein, protein C, which cleaves Factors Va and VIIIa to slow down clotting. There’s also a protein called tissue factor pathway inhibitor that inhibits Xa and the tissue factor-factor VIIa complex. Point is that there’s a balance going on.
Explain the function of platelets in hemostasis. Describe the process of platelet aggregation
As mentioned, adhere to site of endothelial injury, partly mediated by vWF. Activated by thrombin (IIa) to release vasoconstrictors (TXA2), calcium, and ADP, as well as changing conformation, exposing proteins (factors IIb and IIIa, if I’m reading my notes right) to bind more vWF and fibrinogen, thus binding and activating more platelets.
Platelets also have receptors for Factor V, which (recall) is the orienting co-factor for the conversion of prothrombin to thrombin. This speeds up that conversion.
Platelets also expose the phospholipid complex, which provides a greatly pro-clotting enzymatic effect on the coagulation cascade.
As mentioned just now, platelets adhere due mainly to vWF but are activated largely by thrombin (from clotting factor activity). Once activated, platelets expose receptors which bind vWF and thus bind other platelets to the site, as well as releasing vWF, ADP, etc in granules to recruit and activate them.
Note epinephrine, ADP, and collagen also activate platelets.
Drugs: see “Pharmacology of Anticoagulation Therapy.” Essentially aspirin, ADP receptor blockers, and adhesion glycoprotein inhibitors.
Explain the contribution and influence of endothelial cells on coagulation
As mentioned, when damaged, they expose collagen (mainly type II), vWF, and tissue factor.
Collagen and vWF bind platelets.
Tissue factor activates the extrinsic clotting pathway.
Describe the regulatory mechanisms of coagulation, i.e., protein C pathway, antithrombin, fibrinolytic pathway
Fibrinolytics:
Fibrin (clot material) is degraded by plasmin, which is cleaved and activated by tissue plasminogen activator (TPA). After degradation of fibrin, fibrin split products (also called fibrin degradation products) are formed, which are also anti-coagulation factors.
Protein C pathway:
Activated thrombin (IIa) activates Protein C, with thrombomodulin as a cofactor.
Protein C, with Protein S as co-factor, inactivates Factors Va and VIIIa.
Note that a mutation in Va can cause resistance to its inactivation.
Antithrombin:
Antithrombin III forms a complex with thrombin and other serine proteases to block their activity.
It’s activated by heparin (endogenously produced anti-clotting substance).
Note that, in the absence of endothelial damage, the balance is generally in favor of anti-clotting activity.
Review events occuring during hemostasis. Compare primary and secondary hemostasis.
Adhesion, activation, and aggregation of platelets to form a platelet plug constitute the first events in formation of a clot (primary hemostasis). The platelet plug is stabilized by formation of a fibrin network generated through the coagulation cascade (secondary hemostasis). Optimal numbers and function of platelets are key to cessation of bleeding from small vascular injuries.
Diagram the structure of a mature platelet and show the location of: dense granules, alpha granules, glycoprotein 1a, glycoprotein Ib, glycoprotein IIb/IIIa, and phospholipids.
Dense Granules: located inside the platelet cell, release ADP which stimulates local platelets to aggregrate and become activated. Dense granules are less common inside the platelet cell than alpha granules.
Alpha Granules: located inside the platelet cell, release growth factors, fibrinogen, VWF
Glycoprotein 1a: Receptor on the surface of the platelet cell that binds collagen of the injured subendothelial tissue.
Glycoprotein 1b: Receptor on the surface of the platelet cell that binds to VWF
Glycoprotein IIb/IIIa: Receptor on the surface of the platelet cell that binds fibrinogen and links platelets together to form clot.
List three functions of platelets.
- Platelets play several important roles in hemostasis, including adhesion to the vascular subendothelium at sites of injury to begin the hemostatic process.
- Activation of intracellular signaling pathways leading to cytoskeletal changes and release of intracellular granules to enhance platelet plug formation, aggregation to form the platelet plug, and
- Support of thrombin generation by providing a phospholipid surface for the coagulation cascade to take place.
Construct a simple diagram that depicts the process of platelet adhesion. Include in the drawing subendothelial collagen, von Willebrand factor, and glycoprotein Ib. Explain why platelet adhesion to blood vessels does not occur under normal circumstances.
Platelet adhesion to blood vessels does not occur under normal circumstances. The endothelial cells of intact vessels prevent blood coagulation by secretion of a heparin-like molecule and through expression of thrombomodulin, which when bound to thrombin activates protein C and S (inhibit factor Va and VIIIa). Intact endothelial cells prevent platelet aggregation by the secretion of nitric oxide and prostacyclin, inhibitors of platelet activation.
Similarly, construct a simple diagram that shows the process of platelet aggregation; include the release reaction (ADP), thromboxane synthesis, ADP and thromboxane receptors, glycoprotein IIb/IIIa, and fibrinogen.
List and describe three mechanisms that could lead to thrombocytopenia.
- decreased platelet production (due to aplastic anemia, myelodysplasia, and leukemia, infection, severe B12/folate deficiency)
- increased platelet destruction or consumption (immune thrombocytopenic purpura - ITP, autoimmunity against platelets; DIC)
- sequestration of platelets in the spleen.
A normal platelet count is between 150,000 and 400,000/uL. Spontaneous hemorrhage and increased risk of hemorrhage with trauma or surgery may be seen with platelet counts <50,000/uL, and with platelet counts less than 10-20,000/uL, life-threatening spontaneous hemorrhage, such as spontaneous intracranial hemorrhage, can be seen.
Identify three methods of treating ITP and the mechanism by which they increase the platelet count.
Acute ITP (more common in kids) may spontaneously resolve or can be treated with steroids.
Chronic ITP (more common in adults) almost always requires treatment. The most commonly used treatment options include corticosteroids, intravenous immunoglobulin (IVIG), and splenectomy. Steroids work by dampening proliferation of the B cell clone making the autoantibody. An effect is usually seen within 7 to 10 days of starting treatment. IVIG acts by blocking splenic Fc receptors to prevent their binding to antibody-coated platelets, with an effect being seen within 1 to 2 days. Splenectomy works by removing the site of autoantibody-induced platelet removal and leads to lasting responses in 60 to 70% of patients.
Describe the molecular defect, typical clinical course, and general approach to treatment for a patient with Von Willebrand Disease.
Molecular Defect: Abnormality in platelet / endothelial interaction
Clinical Course: Mucosal bleeding, nose bleeds, GI bleeds, Menorrhagia, bleeding after surgery if no correction.
Diagnosis: PFA is a screening test that would confirm a bleeding disorder/functionality of VWF protein, can directly measure the level of Factor VIII, Von Willlebrand Antigen measures amount of Von Willebrand protein, can measure Von Willebrand activity via donor platelets
Treatment:
- Avoid aspirin, NDSAIDS, and other platelet inhibiting drugs
- prophylaxis tx not generallly required
- DDAVP (arginine vasopressin) enhances already synthesized VWF release from endothelial stores (only effective in type 1 VWF disease)
- Replacement therapy
List important questions to ask when obtaining a bleeding history in a patient with excessive bleeding.
A detailed history of the type, frequency, and amount of bleeding is essential when evaluating a patient for a suspected bleeding disorder. Some patients may consider an appropriate amount of bruising or bleeding to be excessive, making the evaluation even more challenging.
Questions to address include:
- Does the patient display excessive, prolonged, recurrent, or delayed bleeding?
- Has the patient ever had the opportunity to bleed excessively (physical trauma, skin lacerations, surgery)?
- Is there a family history of significant bleeding?
Other:
- Is the bleeding real?
- Is it platelet type or coagulation factor type bleeding?
- Is it acquired or congenital?
- What tests do I order and how do I interpret them?
List important laboratory studies to obtain when evaluating a patient with excessive bleeding.
Basic screening tests when evaluating excessive bleeding can include:
• Platelet count and blood smear to evaluate for thrombocytopenia or other hematologic abnormalities
• Bleeding time or platelet function analyzer (PFA-100) to evaluate primary hemostasis
• APTT as a screening test for the intrinsic coagulation pathway
• PT/INR as a screening test for the extrinsic coagulation pathway
• Thrombin clotting time (TCT) to evaluate for fibrinogen defects, the presence of fibrin split products, or heparin effects
• Fibrinogen level
Other, more advanced tests
- Appropriate factor levels to rule out factor deficiency
- 1:1 mixing of normal plasma with patient plasma to evaluate for a lupus anticoagulant or factor inhibitor can then be done.
List some of the major congenital or acquired disease states causing bleeding and/or clotting.
Common: Hempophilia A (Factor VIII deficiency) and B (factor IX deficiency) and C (factor XI deficiency) Von Willerbrand disease Less Common: Factor VII deficiency Hypo or dysfibrinogenemia Rare: Factore XIII, V, X, and II (very rare, not likely compatible with life)
Explain what PT/INR is, and what they are testing. Provide a ddx of an abnormal PT/INR. Provide ddx of abnormal findings.
Protime/International Normalized Ratio Calcium and thromboplastin are added to citrated plasma and time to clotting is measured in seconds. Thromboplastin varies by manufacturer and time in seconds is normalized by adjusting for known potency (INR=1 is normal time)
Prolonged with II, VII, V, X and fibrinogen deficiencies. Prolonged with vitamin K deficiency and liver disease.
Explain the key factor in determining how long someone should be anticoagulated for a venous thrombosis.
Key in the decision-making process is determining if the DVT occurred due to transient risk factors (such as surgery, temporary immobilization, trauma, or pregnancy) or if the patient has an underlying hypercoagulable disorder or ongoing risk factors that require longer treatment.
