Term
| What proteins and platelet receptors are involved in platelet adhesion to damaged endothelium? |
|
Definition
| Von willebrand factor on subendothelial collagen binds to platelets (glycoprotein Ib) |
|
|
Term
| What proteins and platelet receptors are involved in platelet aggregation following adherence or activation by agonists (e.g. ADP, thrombin)? |
|
Definition
| Platelet receptor (glycoprotein Ib-IIa) binds to fibrinogen which binds to other glycoprotein Ib-IIa receptors, effectively aggregating platelets together |
|
|
Term
| What platelet membrane phospholipids are involved in the coagulation cascade and how are they made available to coagulation factors when needed? |
|
Definition
| when stimulated the anionic phospholipids involved in coagulation (e.g. phosphatidylserine or platelet factor 3) move from the inner membrane to the outer member. |
|
|
Term
| Ho to platelets participate in wound closure/clot retraction? |
|
Definition
| via glycoprotein Ib-IIa and fibrinogen bridges and platelet actin and myosin contractile fibers |
|
|
Term
| What is the normal BMBT time in dogs? |
|
Definition
usually < 4 min
**the test is relatively insensitive, but will be prolonged with moderate to marked defects in primary hemostasis. Can also be used as a screening test for vWD in predisposed breeds and should be prolonged with vWF: Ag < 20%, but may also be prolonged with greater values |
|
|
Term
| How can anemia prolong BMBT? |
|
Definition
Decreased HCT allows platelets to circulate more centrally so that they are further away from the vessel wall.
Fewer erythrocytes result in decreased erythrocytes ADP, which is a platelet agonist |
|
|
Term
| What anti platelet drug can prolong BMBT (but not necessarily out of reference interval)? |
|
Definition
aspirin
Other anti platelet drugs typically do not prolong BMBT |
|
|
Term
| What substance, other than platelets can affect BMBT? |
|
Definition
| fibrinogen. Patient's with afibrinogenemia can have prolonged BMBT, presumably due to impaired platelet aggregation given the absence of fibrinogen |
|
|
Term
| Why might you do a clot retraction test? |
|
Definition
| To assess for thrombopathias (namely glanzmann's thrombasthenia- deficiency in Gp IIbIIIa, impaired platelet fibrinogen binding). <--- affected individuals will have impaired clot retraction |
|
|
Term
| In what species to circulating platelets contain a large amount of vWF? |
|
Definition
cats
as opposed to dog platelets, that contain very little vWF |
|
|
Term
| What cells produce the most vWF and how is the vWF stored? |
|
Definition
most vWF is produced by endothelial cells and either constitutively secreted or stored (Weibel-palade body) in endothelial cells.
Megakaryocytes and platelets also contain a large percentage of circulating vWF (except in dogs) |
|
|
Term
| What is the carrier molecule for coagulation factor VIII? |
|
Definition
vWF (via non-covalent complexes)
** so animals with vWD may have slightly prolonged PTT due to decreased VIII activity (however, PTT is usually WRI even in dogs with Type 3 vWD i.e. NO vWF) |
|
|
Term
| What is the most common hereditary bleeding disorder in dogs? |
|
Definition
Von Willebrand disease
Secondary vWD has been reported in people with hypothyroidism however there is not strong evidence that tis occurs in veterinary species |
|
|
Term
| What are the three types of vWD and what dogs breeds are they associated with? |
|
Definition
Type 1: most common, occurs in all breeds including dobermans. all vWF multimers are present but at decreased concentrations. The clinical severity varies.
Type 2: rare, reported in German shorthair and wirehair pointers and horses. Disproportionate decrease in large vWF multimers --> severely affected.
Type 3: rare, reported in Chesapeake bay retrievers, dutch kookier, scottish terriers, and shetland sheepdogs. Absence of ALL vWF multimers --> severely affected |
|
|
Term
| How does sample storage affect vWF:Ag? |
|
Definition
vWF:Ag values significantly increase in both whole blood (by 24h) and plasma (by 48h) when stored at room temp.
No change was noted in refrigerated samples. Rec plasma is quickly collected after blood collection and frozen for best results. |
|
|
Term
| What is the primary method used for vWF:Ag assays? |
|
Definition
quantitiative ELISA with species specific antibodies to vWF
values <50% are considered decreased |
|
|
Term
| How can type 1 and type 2/3 vWD be differentiated? |
|
Definition
Multimeric analysis (immunoelectrophoresis)
vWF multimers are separated by agarose electrophoresis, if type 1 vWD high molecular weight multimers are present vs. Type 2/3 high molecular weight multimers are absent |
|
|
Term
| Describe the two functional assays that can be used to asses for vWF |
|
Definition
Botrocetin cofactor assay- the rate of platelet agglutination in the presence of botrocetin and vWF correlates well with the amount of plasma vWF:Ag (except with Type 2 vWD, because the large vWF multimers are most functional)
vWF collagen binding activity- ELISA (dogs only) that measures vWF that can bind to collagen (i.e. functional vWF). Since large vWF multimers are most functional the vWF:CBA may be increased in dogs with Type 2 vWD due to decreased large multimers and therefore a greater decrease in functional vWF than in vWF ag |
|
|
Term
| What non-genetic test can be used to assess dogs for carrier state of affected with vWD? |
|
Definition
vWF : Ag ratio
if vWF : Ag is < 50% these dogs are carriers AND at risk for clinical disease (the lower the ratio the greater the risk, affected are usually <35%)
if vWF : Ag is 50-69% these dogs are unlikely to be clinically affected but may be carriers. |
|
|
Term
| What physiologic conditions can affected plasma vWF? |
|
Definition
strenuous exercise (dogs and horses)
pregnancy/partuition (dogs)
epinephrine |
|
|
Term
| What substance can be administered to dogs to increase plasma vWF? |
|
Definition
| vasopressin (DDAVP), probably increased release of vWF from endothelial cell stores (Weibel–Palade bodies) |
|
|
Term
| What pathologic states may be associated with increased vWF? |
|
Definition
endotoxemia
azotemia
liver disease
other illnesses |
|
|
Term
| What actives the extrinsic coagulation pathway? |
|
Definition
| Tissue factor (expressed on vascular smooth muscle, activated monocytes, and other extravasated cells) |
|
|
Term
| What actives the intrinsic (surface induced) coagulation pathway? |
|
Definition
Exposed sub-endothelial collagen, activated platelets (provide a negatively charged surface for enzymatic reactions to occur)
+ high molecular weight kininogen and prokallikrein +
factor XII (12) |
|
|
Term
| What inhibits the intrinsic/surface induced coagulation pathway? |
|
Definition
| Antithrombin, proteins C, S, Z |
|
|
Term
| What coagulation factor products feed back into the coagulation cascades to amplify the response? |
|
Definition
Factor 7a-TF (extrinsic pathway): also activates factor 9 (intrinsic)
Factor 2a (aka thrombin, combined pathway): activates factors 11 and 8 (intrinsic), and factor 5 (common) |
|
|
Term
| What factor cross links fibrin multimers (i.e. stabilizes fibrin)? |
|
Definition
|
|
Term
| Which of the coagulation factors are vitamin K dependent? |
|
Definition
|
|
Term
| Which coagulation factor genes are located on the X chromosome? |
|
Definition
factor 9 (intrinsic/surface associated)
factor 8 (cofactor, facilitates conversion of factor 9 --> factor 10) |
|
|
Term
| What is the plasma half life of most coagulation factors? |
|
Definition
1-2 days
**factor 7 has the shortest half life, < 5h in dogs |
|
|
Term
| What coagulation factor is the most important for maintaining hemostasis in vivo? |
|
Definition
| Tissue factor (extrinsic pathway) |
|
|
Term
| What coagulation factors act as cofactors by facilitating surface attachment and localization of coagulation factors for enzymatic reactions to proceed? |
|
Definition
Factor 8 (facilitates conversion of factor 9 --> 10)
Factor 5 (faciliates conversion of factor 10 --> factor 2 (prothrombin)) |
|
|
Term
| Which coagulation factor is a positive acute phase protein? |
|
Definition
Factor 8
**factor 8 circulates in a noncovalent complex with vWF and has a shorter half life if there is less vWF. vWF also increases in response to epinephrin, exercise and other diseases (e.g. endotoxemia) |
|
|
Term
| What enzyme is responsible for converting Vit K (from diet or intestinal bacteria) into active Vit K for participating in Coag factor (2, 7, 9, 10) activation? |
|
Definition
Vit K reductase
**reduced Vit K is a cofacotor for Vit K-dependent carboxylase, which activates coagulation factors 2, 7, 9, 10 |
|
|
Term
| What biochemical modification of Vit K dependent cofactors (2, 7, 9, 10) is necessary for activation of these factors? |
|
Definition
gamma-carboxylation of glutamic acid residues (occurs it vitamin K dependent carboxylase)
** this post-translational modification allows these cofactors to bind fCa (which is necessary for engagement with phospholipid membranes) |
|
|
Term
| What is the major inhibitor of coagulation factors? |
|
Definition
Antithrombin (aka antithrombin III) binds, inactivates, and removes most coagulation enzymes from circulation
**action of antithrombin is markedly enhanced by heparin |
|
|
Term
| What coagulation factors are inactivated by protein C? |
|
Definition
factor 5, factor 8
** active protein C also promotes fibrinolysis by inhibiting plasminogen activator inhibitor |
|
|
Term
| How is protein C (anti-coagulant) activated? |
|
Definition
by thrombin (esp. when bound to thrombomodulin on endothelial cell surface)
*activation of Protein C also requires a cofactor (Protein S) |
|
|
Term
| How does heparin enhance antithrombin activity? |
|
Definition
heparin binds to anti-thrombin inducing a conformational change that increases its affinity for thrombin. Thrombin-antithrombin complexes dissociate from heparin and are cleared from circulation by hepatocytes.
**this does not occur when coagulation enzymes are bound to fibrin or platelets, as such localizes and controlled coagulation can proceed but is limited to the site of endothelial injury. |
|
|
Term
| What is the ideal ratio of plasma/whole blood to citrate for samples intended for coagulation testing? |
|
Definition
1 (blood/plasma) : 9 (trisodium citrate)
**the concentration of citrate used can also significantly affect coagulation parameters, the magnitude of which is not known for most veterinary species. It is important to make sure the concentration of citrate is kept consistent with that used to generate reference intervals. |
|
|
Term
| What has a greater effect on coagulation test results with regards to citrated blood, anemia or erythrocytosis? |
|
Definition
Erythrocytosis.
**with erythrocytosis there is less plasma to citrate when a standard volume of blood is collected, the excess citrate binds too much of the Ca added during the testing resulting in falsely prolonged results. The effect of anemia is theoretically opposite, however, doesn't appear to be clinically significant. |
|
|
Term
| Describe the methodology of electromechanical coagulation analyzers |
|
Definition
Electrical conductance: increased electrical conductance is detected between a stationary and moving electrode as fibrin forms
Electromagnetic viscosity: analyzers detect decreased movement of a small metal ball suspended in the sample as it oscillates in an electromagnetic field |
|
|
Term
| Describe the methodology of photo optical coagulation analyzers. What are the benefits and limitations of this methadology compared to electromechanical methods? |
|
Definition
| Photo optical methods detect changes in light transmission or scatter as fibrin forms. Compared to electromechanical methods, these methods are more accurate at low fibrinogen concentrations but are more susceptible to interferences (e.g. icterus, lipemia, hemolysis) |
|
|
Term
| What arms of the coagulation cascade are assessed by the Lee-White method and the ACT? |
|
Definition
intrinsic (surface activated) and common pathways
** essentially how long does it take for blood to form a clot when placed in a glass tube (Lee-White) or a glass tube containing diatomaceous earth (ACT). Both are relatively insensitive and the Lee-White method is rarely used |
|
|
Term
| Describe a situation where ACT may be prolonged despite normal secondary hemostasis |
|
Definition
Severe thrombocytopenia (reduced phospholipid availability for coagulation factors)
**in actuality ACT is often normal even with severe thrombocytopenia. Mild to moderate thrombocytopenia is not expected to affect ACT |
|
|
Term
| How sensitive is PTT for detecting decreased activity of coagulation factors (intrinsic, surface activated pathway)? |
|
Definition
insensitive, 70% of coag factor activity must be lost before PTT is prolonged
**however, if activity of multiple factors is impaired a milder reduction in individual factor activities can result in a prolonged PTT |
|
|
Term
| Deficiencies in which coagulation factors are not associated with clinical bleeding, but do cause prolonged PTT? |
|
Definition
factor 12, prekallikrein
**Kallikrein is a cofactor for activation of factor 12 |
|
|
Term
| T/F PTT is a good way to monitor animals with heparin therapy |
|
Definition
False
target prolongations in PTT developed for humans with heparin therapy do not correlate with appropriate anticoagulation in animals (PTT is less prolonged in dogs with a given plasma heparin activity, possibly due to greater factor 5 and 7 activity). Differences in methodology are likely the result of differences in the sensitivity of various reagents to heparin. |
|
|
Term
| What is the major difference in the reagent for PTT vs. PT? |
|
Definition
The reagent for PT contains tissue factor (and hence the PT tests the extrinsic and common pathways) and heparin inactivators
**Both reagents contain calcium to overcome the anti-coagulant effect of citrate |
|
|
Term
| What coagulation test is best suited to evaluated a patient for hypofibrinogenemia or dysfibrinogenemia (polymerization of fibrin is impaired because of abnormal fibrinogen molecules)? |
|
Definition
Thrombin time (aka Thrombin clotting time)
In this test thrombin (factor 2a) is added to the patient plasma, which should cleave fibrinogen (factor 1) to fibrin |
|
|
Term
| What diseases are associated with dysfibrinogenemia? |
|
Definition
hereditary (rare)
liver diseases may be associated with an acquired dysfibrinogenemia |
|
|
Term
| What is the difference in methodology between total thrombin clot time and thrombin time(clauss)? |
|
Definition
| Thrombin time(clauss) is the method used to measured [fibrinogen], in which the patient plasma sample is diluted prior to the addition of thrombin, so that patient [fibrinogen] is the rate limiting step. [Fibrinogen] in the sample is estimated from comparison of the TT(clauss) to a standard curve. |
|
|
Term
| Describe the assays used to detect most specific factor deficiencies in dogs and cats. |
|
Definition
1:1 dilution of plasma from the patient and plasma from a known factor-deficient animal are made, if the PT or PTT (depending on the factor) remains prolonged then the patient plasma is also missing the factor.
Results (in seconds) are compared to a reference curve and converted to % activity, relative to the reference plasma pool |
|
|
Term
| Describe the assays used to detect most specific factor deficiencies in dogs and cats. |
|
Definition
1:1 dilution of plasma from the patient and plasma from a known factor-deficient animal are made, if the PT or PTT (depending on the factor) remains prolonged then the patient plasma is also missing the factor.
Results (in seconds) are compared to a reference curve and converted to % activity, relative to the reference plasma pool |
|
|
Term
| For what coagulation factors is there a chromogenic assay to detect specific factor deficiencies? |
|
Definition
|
|
Term
| What coagulation test can be used to specifically assess the common pathway? |
|
Definition
Russell's viper venom time
**not widely offered. The venom directly activates factor 10. |
|
|
Term
| How does Antithrombin activity differ in neonates compared to adults? |
|
Definition
| decreased Antithrombin activity in neonates (esp. foals) |
|
|
Term
| What hormone is associated with decreased antithrombin synthesis? |
|
Definition
|
|
Term
| In what species may antithrombin be an acute phase response protein? |
|
Definition
cats and rabbits
**sepsis however, has been associated with decreased antithrombin activity and procoagulant state |
|
|
Term
| What does increased plasma thrombin-antithrombin complexes [TAT] indicate? |
|
Definition
| Increased generation of thrombin which is associated with thromboembolic disease or hypercoagulable states |
|
|
Term
| What does decreased a plasma protein C concentration indicate? |
|
Definition
| hypercoagulable state due to decreased inactivation of factors 5 and 8, and decreased fibrinolysis |
|
|
Term
| What is the primary plasmin inhibitor in the blood? |
|
Definition
|
|
Term
| How does protein C affect plasmin activity? |
|
Definition
| Protein C enhances plasmin activity (and therefore fibrinolysis) by inactivating plasminogen activator inhibitor (PAI) |
|
|
Term
| What is the methodology used for FDP detection? |
|
Definition
Latex agglutination immunoassays (with anti-human FDP ab)
**at least partial cross reactivity has been shown in dogs but the full extent of cross reactivity within veterinary species is not known |
|
|
Term
| How does increased [FDP] potentially affect other coagulation assays? |
|
Definition
prolonged values for tests with clot end points (PT,PTT, TCT, ACT, lee-white clotting time)
*FDPs competed with fibrinogen for active site of thrombin (inhibit conversion of fibrinogen to fibrin) and compete with fibrinogen for platelet binding sites (GpIIIbIIa) inhibit platelet aggregation). FDPs may also disrupt normal polymerization of fibrin. |
|
|
Term
| Deficiency in which coagulation factor is associated with hemophilia A? Hemophilia B? |
|
Definition
| Factor 8 (hemophilia A), factor 9 (hemophilia B) |
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