Hemostasis SOLO 4

Secondary hemostasis disorders

Factor I

• fibrinogen is a large glycoprotein produced in the liver

• acute phase reactant (increased with trauma, preg, inflamm)

• participates throughout coagulation

  • involved in plt aggregation via GPIIa/IIIb

  • acts as the substrate to create a fibrin clot

  • fibrinogen is acted upon by thrombin and plasmin depending on the situation

• deficiencies of fibrinogen levels are congenital (usually recessive)

  • hypofibrinogenemia

  • afibrinogenemia

• dysfibrinogenemia can be acquired due to liver disease or autoimmune disease

• symptoms can vary depending on the severity of the case

  • hypofibrinogenemia and dysfibrinogenemia may go undeteccted for years until a severe wound when there is a noticeable delay in blood clotting

  • afibrinogenemia will usually present earlier in life

  • umbilical cord stump bleeding

  • testin to confirm abnormality

    • PT/INR, aPTT, fibrinogen, thrombin time, reptilase time

  • treatment

    • transfusion of FFP and cryo

Factor II

• most abundant and longest half likfe of all factors of the prothrombin group

• it is converted to thrombin by the action of FXa, accelerated by FVa

• hypoprothrombinemia can be congenital or acquired

  • congenital: autosomal recessive

  • acquired: vitamin K deficiency or anticoagulant therapy

  • deficiencies delay thrombin generation causing hemorrhagic symptoms

• dysprothrombinemia is generally congenital

  • FII concentration is normal, a structural defect impairs its ability to activate and cleave fibrinogen

  • activity assays (abnormal) and antigen assays (normal) ma identify

• treatment is FFP

Liver disease causes a decrease in what factors?

• all factors made in the liver

• include Factors I, II, V, XI, and X

• FV and FVII differentiate liver disease from vitamin K deficiency

amyloidosis

• a rare disease that occurs when a protein called amyloid builds up in organs, which can make the organs nor work properly

• amyloid fibrils in the vasculature can selectively adsorb FX leading to an acquired FX deficiency

• prolonged PT/INR and aPTT which corrects with mixing studies

Prothrombin mutation (G20210A)

• single point mutation in prothrombin gene on chromosome 11

• associated with elevated prothrombin concentration and increased risk of thrombosis

• second most common cause of inherited thrombophilia

• diagnosis is confirmed with testing in the genetics department using PCR in an EDTA tube

Factor V

• along with FVIII, proaccelerin enhances the enzymatic action of the coag cascade

• heat labile, short half life, synthesized in liver, stored in plt granules

• congenital deficiency called parahemophilia

• acquired deficiency seen in patients with specific antibodies, liver disease, carcinoma, tuberculosis, or DIC

• lab testing shows an increased PT/INR and aPTT

• treatment with FFP

Factor V leiden Mutation

• normally protein C will inactivate FV in the regulation of hemostasis

• however, in some patients there is a mutation present in the gene for FV at the site where protein C binds

• this inability of protein C to bind to FV allows continued coagulation and increases the risk of thrombosis

• the mutation is seen in 90% of patients with activated protein C resistance

• 20-60% of patients with FV Leiden Mutation will present with venous thrombosis

• testing to confirm is performed in genetics via PCR to detect the mutation in an EDTA tube

Factor VII

• proconvertin is a plasma protein of the extrinsic pathway

• interacts with calcium and tissue factor during injury of the blood vessel

• vitamin K dependent

• congenital deficiency due to autosomal recessive inheritance

• acquired due to oral anticoagulant therapy (OAT) and liver disease

• lab testing will show increased PT/INR but normal aPTT

• treat with FFP or vitamink K

Factor VIII

• antihemophilic factor; heat labile; short half life; acute phase reactant

• circulates in the plasma as a complex with vWF

  • complex helps to transport FVIII to the site of injury

• activated by separating FVIII from vWF by thrombin (FVIII:C)

  • aids in the conversion of FX to FXa with PF3 and calcium

• regulated by APC to prevent ongoing coagulation and thrombosis

• elevated FVIII:C is linked to a hypercoagulable state

• deficiency leads to bleeding disorders

Hemophilia A

• FVIII deficiency

• most common hereditary coagulation disorder

• the gene for FVIII lies on the X chromosome

  • all daughters of hemophilic men are carriers and all sons are normal

  • although men who inherit it will get hemophilia, women carriers can expeerience mild bleeding tendencies due to X-inactivation

• majority of people have a deficiency while 5-10% have a dysfunctional form of FVIII:C

• vWF is not affeced by this disease

• small portion of patients will develop antibodies/inhibitors to FVIII:C

• symptoms include hemarthrosis, hematuria, intracranial bleeding, hematomas, and spontaneous hemorrhage

• mild hemophili: >5% FVIII”C activity

• severe hemophilia: <1% FVIII:C activity

  • spontaneous bleeding occurs at <1% FVIII levels

• lab shows prolonged aPTT and mixing study is corrected

  • apTT as a screen is relatively insensitive in detecting FVII deficiency and will be normal unless the patietn has <30% of FVIII levels

• normal PT/INR, PFA, plt aggregation

• treatment with FFP, cryo, novoeight

Factor IX

• christmas factor, plasma thromboplastin component

• single chain glycoprotein

• vitamin K dependent

• intrinsic pathway factor

  • activated by FXIa and calcium

  • also activated by tissue factor

Hemophilia B

• FIX deficiency

• sex-linked recessive bleeding disorder

• can be acquired though OAT, vit K deficiency, or liver disease

• mild hemophilia: 2-25% activity

• severe hemophilia: <1% activity

• lab shows prolonhed aPTT and corrected mixing study

• normal PT/INR, PFA, plt aggregation

• treatment with FFP or BeneFIX

Factor X

• Stuart-Prower factor

• beginning of common pathway

  • intrinsic pathway: activated by FIX

  • extrinsic pathway: activated by tissue factor tenase complex

• FX assay is not the same as anti-XA test

• congenital deficiency due to autsomal-recessive trait

• symptoms vary from easy bruising, epitaxism to menorrhagia and hemarthrosis

• diagnosis can be obtained by family analysis and proper labs

• labs show prolonged PT and aPTT, corrected with mixing study

• treatment is FFP

Factor XI

• plasma thromboplastin antecedent

• contact factor of involved in the early stages of the intrinsic pathway

• circulates as a complex with HMWK

• after contact with a negatively charged surface, become FXIa

Hemophilia C

• FXI deficiency

• autosomal recessive trait

• heterozygotes have some degree of abnormal bleeding

• FXI is the only factor of the contact system in which a deficiency may cause bleeding

  • bleeding does not correlate well with factor levels in FXI deficiencies

• symptoms are similar to other hemophilias, except bleeding in joints usually doesn’t occur

• normal PT/INR, PFA, TT

• prolonged aPTT that corrects with mixing study

• treat with FFP

• FXI supplements or boosts the activation of FIX, so deficiencies of FXI are less severe clinically than deficiencies of FIX or VIII

Factor XII

• Hageman factor

• single chain beta-globulin

• member of the contact system

• initiates the intrinsic pathwya by converting PK into kallikrein and FXI to become FXIa

• activated in vitro by contact with glass, celite, kaolin, ellagic acid

• activated in vivo by contact with collagen and C1 esterase (cleaves FXII)

• involved in the activation of plasminogen for fibrinolysis

Factor XIII

• fibrin stabilizing factor

• circulates with fibrinogen in the bloodstream

• is a heterotetramer consisting of 2a and 2B linear chains

• activated by thrombin

• involved in the final stage of coagulation where cross-links between D subunits of firbin are made

• strengthens and make the cross insoluble

• requires calcium to function

Factor XIII deficiency

• congential deficiency autosome recessive trait

• acquired deficiency reported with certain leukemia, DIC and severe liver disease

• deficiency is characterized by moderate to severe hemorrhages; inital bleeding is stopped, but then a sudden recurrence within 36 hours is likely

  • patients intially clot but later start to bleed

  • slow, delayed but progressive bleeding occurs accompanied by poor wound healing and slowly resolving hematomas

• often diagnosed at birth with umbilical stump bleeding

• lab tests show normal screens

• a 5M urea test must be performed to confirm FXIII deficiency; dissolution of a clot within 24 hours of being placed in 5M urea solution will confirm the deficiency

• treat with FFP or cryo

Prekalikrein

• fletcher factor

• substrate for FXII to form kalikrein and FIXa

• approx. 75% of PK is bound to HMWK and released during contact activation

• kalikrein is the most potent plasminogen activator in the contact phase of coagulation

• PK deficiency

  • inherited as both autosomal-dominant and autosomal recessive traits

  • not associated with clinical bleeding

  • patients may have thrombotic event throughout lifeitme

  • prolonged aPTT

HMWK

• fitzgerald factor

• single chain glycoprotein

• required for contact activation of the intrinsic pathway

• HMWK deficiency

  • autosomal recessive

  • no clinical bleeding assocatied, but increased risk of thrombosis

  • aPTT will be prolonged and corrected with mixing study

  • no treatment neccessary

Defects that lead to impairment of the coagulation system

• decreased factor synthesis

• intereference of abnormal molecules that interfere with coagulation pathway

• loss, consumption, or inactivation of factors

• inactivation of factors by inhibitors or antibodies

Acquired causes of factor deficiencies

• vitamin K deficiency

• liver/renal disease

• hemorrhage

• contaumptive coagulaopathy (DIC)

• anti-coagulant therapy

• inflammatory disorders

• drug therapy

Thrombosis

• hypercoagulability is an alteration of the coagulation mechanism that predisposes a person to thrombosis

• thrombosis is one of the most common causes of death in the US

  • 10-30% of people with VTE will die within one month of diagnosis

• half of people with DVT will have long term complications such as swelling, apin, discoloration and scaling

• lab screening includes D-Dimer

Venous thrombosis

• caused by fibrin and RBCs

• Deep vein thrombosis

  • a condition in which blood clots form in veins located deep inside the body, usually in the thigh or lower legs

  • can cause pain and swelling in the area

arterial thrombosis

• caused by WBCs and PLTs

• activated platelets, monocytes, and macrophages become embedded within fatty plaque combining with vWF to form arterial platelet plugs, resulting in white thrombi

• pulmonary embolism

  • a condition where a blood clot travels form another part of the body, usually the legs (DVT), to the lungs and blocks one or more pulmonary arteries

inherited thrombophilia

• group of congenital hematologic disorders in which a genetic defect can be identified that increased the risk for thrombosis

• approx. 5-8% of the US has one of several genetic risk factors for inherited thrombophilia

Causes of VTEs

• altered blood flow

  • disease blood vessels

  • post-op patients

  • sedentary patients

  • prolonged sitting

• abnormalities of hemostasis

  • disturbances in coagulation inhibitors

  • disturbances in fibrinolysis

• specific causes

  • FV leiden mutation

  • PT G20210A mutation

  • protein C/S deficiency

  • ATIII mutation

  • FXII deficiency

  • dysfibrinogenemia

  • microangiopathic thrombocytopenia

  • anti-phospholipid antibodies

  • hyperhomocysteinemia

  • secondary to contraceptives, nephrotic syndrome, medication, major traume, or pregnancy

Pregnancy associated thrombosis

• increased factor levels

  • FVII, FVIII, FX, vWF

• decreased protein S

• greatly increase PAI-1

• increased PAI-2, which is produced by the placenta

Pro-thrombotic platelet action

• PF3 found on the membrane which facilitates factor activation

• shape change aids in the formation of aggregates along with firbinogen and GPIIb/IIIa

• when activated, platelets release granules that promote coagulation

anti-thrombotic platelet action

• while in an inactive state, the glycocalyx is positively charged which prevents interaction with endothelial cells and erythrocytes

• platelet activation is reversible as long as the acting stimuli isn’t potent enough

pro-thrombotic endothelium action

• upon damage, endothelial cells release arious components:

  • vWF which is an adhesion protein for platelets

  • tissue factor which initiates the extrinsic pathway

• exposure of collagen initiates the intrinsic pathway

• endothelial cells produce and secrete PAI-1, which suppresses fibrinolysis

Anti-thrombotic endothelium action

• intact endothelial cells prevent platelet adhesion

• healthy and intact endothelial cells release components tha tinhibit coagulation

  • prostacyclin (PGI2) and nitric oxide which are powerful inhibitors to platelet agreegation and also act as vasodilators

  • t-PA and u-PA both activate plasminogen to cleave any excess fibrinogen or fibrin in the vicinity

• surface proteins such as thrombomodulin and heparan sulfate play a part in maintaining an anti-thrombotic environment

pro-thrombotic thrombin action

• cleaves fibrinogen to form fibrin

• converts FXIII into the active transglutamase, which cross-links fibrin with covalent bonds that render the fibrin insoluble

• activated FV, VIII, IX, XIII

• binds and activates nearby platelets to initiate shape change, aggregation, and possibly granule release

anti-thrombotic thrombin action

• binds to thrombomodulin on the surface of endothelium which goes on the activate the protein C system

• contributes to fibrinolysis by inhibiting PAI-1

Pro-thrombotic plasma protein action

• contains coagulation factors necessary for the cascade to take place

anti-thrombotic plasma protein actions

• contains a series of serpins (serine protease inhibitors) that oppose the generation of thrombin by suppressing activated proteases as a result of the cascade

  • ATII and heparin cofactor II

  • protein C and S

  • TFPI (tissue factor pathway inhibitor)

specific inhibitors

• certain patients create immunoglobulins to specific coagulation factors

• some people can develop inhibitors if they have a deficiency, but there are cases where factor levels are insufficient

• patients with specific inhibitors can experience hemorrhagic episodes

• FVIII inhibitors are the most common specific factor inhibitor

  • an antibody against FVIII presents as an increased aPTT and a normal PT/INR

  • FVIII inhibitors will develop in 10-15% of patients with hemophilia A

  • pregnant women are also reported to develop this inhibitor during pregnancy

  • patients with underlying immunological disorders have an increased risk of developing this autoimmune disease

antiphospholipid syndrome

• non-specific inhibitor

• a prothrombotic disorder that can present as recurrent thombotic events and/or a history of miscarriages

• patients have a non-specific anti-phospholipid antibodies present in their circulation

• these antibodies promote thrombosis by interacting with various phospholipids in the bloodstream to stimulate coagulation

lab testing for inhibitors

• Bethesda assay/titer

• involves incubation of normal pooled plasma with varying dilutions of patient plasma for two hours, then measuring factor activity

anti-phospholipid antibodies can include:

• lupus anticoagulant

  • most often associated with thrombosis, but may experience bleeding due to presence of antibodies to FII

• anti-cardiolipin antibodies

  • first noted in high incidence of fals epositive for syphilis due to the fact that thee test uses cardiolipin as a reagent

• anti-beta2-glycoprotein 1 antibodies (anti-apolipoprotein H)

  • interferes with FXa inhibition

Lupus anitcoagulants (LAC)

• factor assays are normal with higher dilutions

  • dilutions are performed in factor assays and should agree within 10% f each other, if not they are suggestive of an inhibitor

  • as the patient sample with the inhibitor is diluted, it dilutes out the inhibitor and the factor activity increases

• PT/INR normal

• PTT increased

• mixing study will NOT correct the PTT

• dilute Russell’s Viper Venom time (dRVVT) is sensitive to the presence of LA

  • assessment of the time for blood to clot in the presence of a diluted amount of venom from Russell’s viper (Daboia russelii), a highly venomous snake

Circulating inhibitors will lead to the following lab results

• normal PT/INR

• abnormal aPTT

• mixing study does not correct

• normal factor assay

Unfractionated heparin overview

• UFH is widely used anticoagulant administered IV or subcutaneously given to prevent thrombosis, keep IV lines open, and used in heart-lung bypass machines

• heparin is an unbranched polysaccharide that is heavily sulfates, making it anionic

• commercial preps are made by extracting these molecules form bovin lungs or porcine intestines

• AT binds FXIIa, XIa, IXa, X, and thormbin; heparin acts by accelerating the rate at which AT is capable of binding, irreveribly inhibiting it actvitiy

UFH monitoring

• dose is determined by weight of patient and given as a bolus dose followed by continuous IV infusion

• monitoring UFH should beign after equilibrium is reached usually 6 or more hours after starting or changing therapy

• once in therapeutic range, daily monitoring is sufficient

• heparin has a veyr narrow therapeutic range

• the goal is to prevent thrombosis without causing hemorrhage

• heparin overdose is treated with protamine sulfate

• lab needs to inform physicians of heprain therapeutic range

Lab strategies to monitor heparin

• aPTT

  • moniors effect of unfractionated heparin on patient’s coagulation system

  • general rule is the therapeutic range is 1.5-2.5 time the upper limit of aPTT reference range

  • PTT ref range- 22.2-36.1 secs

• Anti-Xa assay

  • measuress concentration of heparin in blood

• PT/INR can also become elevated if given too much heparin due to the dilution effect

problems with heprain monitoring

• variation in durg

• variation in resopnse of patient

• variation in reponse of reagent

• variation in instrumentation

low moelcular weigh heparin (LMWH) overview

• LMWH is prepared from UFH by fractionation or depolymerization

• produces heparins of lower, more uniform MW

• function by the same mechanism as UFH, so it is not recommended for the treatment of HIT

• types include:

  • Louvenox, fragmin, arixtra

• administered subcutaneously and many don’t require monitoring

  • patients with renal failure, low/high weight, peds, OB pts, and others require mointoring with anti-Xa assay

• patients do not present with prolonged aPTT

• acts only on FXa (not FIIa)

Direct thrombin inhibitors

• alternative anticoagulants to heparin for HIT patients

• are either isolateed from the leech (Hirudo medicinalis) or are recombinant forms of the same protein (lepirudin, agratroban, dabigatran)

• have a potent inhibitory effect on thrombin

• aPTT target is 1.5-2.5 times mean of normal range

• examples of DTIs

  • Hirudin, leprudin, refludin, argatroban, dabigatran

Heparinoids

• glycosaminoglycans that are derivatives of heparin

• alternative anticoagulants to heprain for HIT patients

• include oligosaccharides and sulfated polysaccharides of platn, animal, or synthetic origin

• danaparoid is a mixture of the glycoaminoglycans, heparan sufate, dermatan sulfate, and chondroitin sulfate

• catalyzes the inactivation of FXa

• can be monitored with anti-Xa assay

Warfarin

• oral anticoagulant that acts by interfering with the recycling of Vitamin K after its carboxylation functions

• the first factor to be decreased is FVII because of its short half life

• takes 5-7 days of warfarin therapy before a stable anticoagulant effect is achieved

• eliminated hepatically and works by inhibiting vitamin K dependent factors

• has a narrow therapeutic range and has less than optimal tests and reagents to monitor it

• Vitamin K is given to treat warfarin overdose

• PT/INR is used to monitor

International Normalized Ratio (INR)

• on warfarin- 2.0-3.0

• with cardiac valves- 2.5-3.5

• INR= (patient’s PT/ geometric mean PT)^ISI

Direct Oral Anticoagulants (DOACs)

• DOACs work by inhibiting FXa or FIIa as well as coplex-bound activated factors and fibrin-bound thrombin

• benefits include no routine monitoring or dietary restrictions, as well as fewer bleeding episodes

• types include:

  • dabigatran (avoid in renal disease, half life increased)

  • rivaroxaban, apixaban, edoxaban

Aspirin

• can be used prophylactically for stroke and MI prevention

• the acetylation of aspirin inactivated cyclooxygenase which blocks thromboxane A2 production and causes impaired platelet function

• TXA2 is necessary for normal plt aggregation

• inhibits prostaglandin synthesis- tissue injury releases prostaglandins that signal plts to form a blood clot at the site of injury in order to stop bleeding

• causes prolonged PFA

Plavix (clopidogrel) and ticlopidine

• inhibits plt aggregation at ADP binding site

• causes prolonged PFA and a decreased plt aggregation with ADP agonist

Reopro, Integrilin, Tirofian, Absiximab, and Aggrestat

anticoagulants directs against platelets and are GPIIb/IIIa inhibitors