Disseminated intravascular coagulation (DIC)(consumptive coagulopathy)

a pathological activation of coagulation (blood clotting) mechanisms that happens in

response to a variety of diseases.

* DIC leads to the formation of small blood clots inside the blood vessels throughout the body* The small clots consume coagulation proteins and platelets, normal coagulation is disrupted and abnormal bleeding occurs from the skin, the gastrointestinal tract, the respiratory tract and surgical wounds. The small clots also disrupt normal blood flow to organs* DIC can occur acutely slower, chronic basis, and may participate in the development of multiple organ failure, which may lead to death.* In acutely ill and shocked with widespread hemorrhage (common bleeding sites are mouth, nose and venipuncture sites), extensive bruising, renal failure and gangrene.

Activation of Platelets

Thrombin activates platelets via protease activated receptors PAR Thrombin cleaves PAR PAR ‘plugs itself in’ Activation of PAR signals intracellular events to begin - Shape change and Granule release

Three Phases of Hemostasis

_ Primary hemostasis_ Formation of a platelet plug_ Vasculature and platelets

_ Secondary hemostatis_ Consolidation of the platelet plug by fibrin_ cascade and inhibitors

_ Fibrinolysis_ Cleanup_ cascade and inhibitors

Acute Phase Response

a group of physiological processes occurring soon after the onset of infection, trauma, inflammatory processes, and some malignant conditions;

increase in acute phase proteins in serum, fever, increased vascular permeability, and metabolic and pathologic changes.

These reactions aim to prevent ongoing tissue damage, isolate and destroy the infective organism and activate the repair processes necessary to restore the host/organism’s normal function

leukocytosis, fever, alterations in the metabolism of many organs as well as changes in the plasma concentrations of various acute-phase proteins made by hepatocytes in the liver

main mediator of the acute phase reaction is IL6 regulated by IL1

Start of Common Pathway Factor 10

Primary Hemostasis Formation of a platelet plug - Vascular system and platelets_ Activation in response to stimuli (Thromboplastin)

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_ Damaged vessel releases_ Epinephrine, collagen, thrombin, ADP, serotonin

_ Adherence to the margins of the lesion - platelets_ Cell membrane receptors _ GpIIb/IIIa, GpIb

_ Release of granule content (caused by GpIIb/IIIa, GpIb)_ Alpha granule _ Dense granule

_ Aggregation into a primary platelet plug

Start of Intrinsic PathwayActivated by

contact activation of factor XII (12)HMWK – high molecular weight kiniageninitiated by components entirely contained within the vasculaturethe contact system-various electronegative surfaces, such as collagen fibers, skin, or sebum

Start of Extrinsic PathwayActivated by

F - III (3) – Tissue ThromboplastinActivated by tissue trauma

factors circulate in the form of inactive precursors called

zymogens

von Willebrand Disease

symptoms include nosebleeds, skin bruises, and hematomas. oral cavity bleeding

excessive menstrual bleeding, easy bruising. Prolonged bleeding after minor trauma to skin or mucous Severe hemorrhage after major surgery delayed bleeding, up to several weeks after surgery. Heavy bleeding is common after tooth extraction or other oral

surgery Bleeding symptoms are often exacerbated by aspirin and oral

contraceptives.

High molecular weight kininogen(HMWK) is also known as what Factor?

Fitzgerald Factoracts as a cofactor for the activation of Factor XI,

prekallikrein, and Hageman factor

Factor (I): Fibrinogen - acted on by thrombin to form fibrin

Factor (II): Prothrombin - Coverts into thrombin

Factor (III): Tissue Thromboplastin - Activates Factor VII when exposed to tissue fluids.

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Factor (IV): Ionized Calcium - Assists in the activation of other factors.

Factor (V): Proaccelerin - Accelerates the conversion of Prothrombin to Thrombin.

Factor (VI) Not assigned

Factor (VII): Stable Factor - activated by Factor IIIEnzyme involved transforming

prothrombin to thrombin

Factor (VIII): Antihemophilic A Passed on from father to son, loss of this factor leads to

inability to clot.essential for the formation of blood thromboplastin

Factor (IX): 2 names and what it does? plasma thromboplastin componentChristmas Factor

Helps activate Factor X

Factor (X): Stuart /Prower enzyme for common pathway activation

of prothrombin

Factor (XI): Plasma Thromboplastin Antecedentprecursor to Plasma Thromboplastin, which is one of the

proteins that alters the shape of blood platelets

Factor (XII): What pathway?Activated when?

Hageman Factor Intrinsic Activated when a foreign substance is in the body.

Factor (XIII): Fibrin Stabilizing Factor(FSF)Helps to further stabilize the clot.

Prekallikren is also known as what Factor? Fletcher Factor

Factor V Leiden: this mutated gene leads to what?

Increase in clot formation which results in a degradation of Factor (V), by Protein C.

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AdhesionPlatelet adhesion is primarily mediated

by binding of

_ the platelet glycoprotein GpIb to subendothelialvon Willebrandfactor(vWF) carries Factor 8

_ it is rapidly followed by release of granule content

Release of granules

_ Dense granules - _ ATP, ADP, serotonin, Calcium_ Alpha granules_ Albumin, fibrinogen, factor V, vWF, HWMK_ Platelet specific - _ β-thromboglobulin, PLT-factor 4, PLT-derived growth factor_ Lysosomal granules - Acid hydrolase

Secondary hemostasis

“Blood coagulation” – coagulation factors Activated by primary hemostasisChange in platelet surface upon aggregation A secondary hemostatic plug is formed The primary platelet plug is reinforced Further bleeding from the lesion is prevented Primary and secondary hemostasis occur simultaneously Interactive mechanism

Physiologic control mechanisms

Blood flow Feedback inhibition Liver clearance Inhibitors

Anti-thrombinsTissue factor pathway inhibitorProtein CProtein S

Types of coagulation factors

ZymogensCirculate in inactive forms Activated to proteasesBecome activated to enzyme formFactors II, VII, IX, X, XI, XII, XIII

Accelerators - ‘Gas pedals’ Factors V and VIII

Others - Fibrinogen (Factor I)

Inhibitors (* Vitamin K dependent) Put the ‘brakes’ on clot formation by

slowing down and regulating the process

Protein S* Protein C*ATIII TFPI

Alpha-2-antiplasminPAI (plasminogen activator factor)

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Fibrinolysis2 actions

_ Keeps blood clots from growing andcausing problems_ Leads to the breakdown of fibrinclots

FibrinolysisPrimary and Secondary

_ 1o - _ Normal clot breakdown _ Occurs naturally_ 2o - _ Clot breakdown due to medications_ tPA, streptokinase, urokinase_ Another disorder _ DIC, Bacterial infections_ Intense exercise, inadequate oxygenation, low blood sugar

3 types of disorders of HemostasisCause bleeding or thrombosis problems

Platelet disordersCoagulation factor disordersFibrinolytic disorders

*Many more deaths occur from thrombosis problems than bleeding

Formation of Platelets

_ Formed in bone marrow_ Begin as megakaryoblasts mature to megakaryocytes_ 4-5 days maturation_ Thrombocytes - _ In circulation 8-12 days_ Normal range: 150,000-450,000 uL_ 10-30 PLTs/HPF

Thrombocytopenia

Most common cause of bleeding_ Range - 50-150 x 109/L

_ Bleeding unexpected_ May imply qualitative disorder

_ 10-50 x 109/L_ Bleeding may follow trauma, surgery, dental_ <10 x 109/L_ Spontaneous bleeding requiring treatment

ThrombocytopeniaCauses

Inherited Marrow hypoplasia

ChemotherapyDrug or ethanol

Immune-mediated consumptionAcute or chronic AITT, neonatal alloimmune,post-transfusion purpura

Non-immune mediated consumption

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TTP, HUS, DIC, HIT, type 2B vWD

Factors that alter automatedplatelet counts

_ False decreases _ Platelet satellitism _ Platelet aggregates_ Giant platelets

_ False increases_ Cell fragments (schistocytes)_ Extreme microcytosis

Chemical that activates platelet receptors seratonin

1. clot stabilizing factor XIII (13)

5. % HCT level where anticoagulant needs adjustment 55%

8. fragments D & E are measured by this test D dimer

10. inactivation of cyclo-oxygenase Aspirin

11. caused by; viruses, antibodies, overactive spleen and unknown etiological agents

ITP (autoimmune)Immune Thrombocytopenic Purpura

Antibodies bind to platelets, shorting platelet lifespan

13. anticoagulant of choice for coag tesing Sodium Citrate

14. intrinsic pathway is activated by Contact Factor

16. plt satellitism, giant plts, and plt clumps all cause falsely "blank" automated platelet counts decreased

18. large molecule that helps platelets bind Von Willibrand Factor

21. Factors X, V, II, and I are in this pathway Common pathway

23. factor VIII deficiency Hemophilia A

27. APTT is the test of choice monitoring "blank" therapy Heprin

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29. Vitamin K antagonist Rat Poison (Coumadin)

28. deficiency causes deep vein thrombosis & hypercoagulabilty

Protein C

31. factor XII deficiency demonstrates no "blank" manifestations

Hemorrahaic

30. Factors II, VII, IX, & X are "blank" dependent Vitamin K

2. factors in this pathway are XII, XI, IX, VIII, X, V, II, & I Intrinsic

3. 170-410 mg/dl is reference range Fibrinogen

4. treatment is Factor IX concentrate Hemophilia B

6. for bleeding time the blood pressure cuff is kept at "blank" mm/Hg

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7. the platelet stores and synthesizes this factor Von Willibrand

9. measured by the FSP test (X, Y, D, & E) Monomers

12. individuals are clinically asymtomatic for most of their life

Factor V Leiden

15. extrinsic pathway is activated by Thromboplastin

17. PT is the test of choice for monitoring "blank" therapy

Coumadin

19. disease state indicated by elevated FSP and D-dimer fragments

DIC

20. non-correction in a mixing study indicates the presence of

inhibitor

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25. 2.0-3.0 is standard therapeutic range INR

26. 27-36 sec is reference range APTT

22. factors in this pathway are VII, X, V, II, & I Extrinsic

24. two weeks is average life span in circulation

Platelet

Thrombus is A Blood clot

Vitamin K coagulation inhibitors Protein C and protein S

plasmin Meat cleaver that chops up clot into smaller pieces

Regulators of the coagulation cascade

Identify substances that are contact activators

G. Describe the physiologic controls of hemostasis

3 actions and 4 inhibitors

• Blood flow • Feedback inhibition• Liver clearance

• Inhibitorso antithrombins o tissue factor pathway inhibitor (TFPI)o protein C o protein S

Primary Hemostasis involves (2) Smaller injuries to blood vessels, platelet plug

Secondary Hemostasis involves (2) Large injuries to blood vessels/tissueInvolves platelets and plasma coagulation

Fibrinolysis - definition Breakdown of fibrin clots caused by enzymes

Man made clot busters for ER TPA, streptokinase, urokinase

Tissue factor (in all cells) does Activates the plasma coag system through F 7

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Coagulation occurs in steps

1. Reactions in 2 pathways activate serine proteaseenzymes and form prothrombin activator(V and X)

2. Prothrombin activator cleaves prothrombin into 2 fragments

3. Thrombin cleaves small peptides to give rise to an insoluble fibrin polymer (also activates F 13 – catalyzes formation of covalent bonds between fibrin molecules, cross-linking)

vWF binds during platelet adhesion to Glycoprotein receptor for platelet surface membrane (GPIb)

Protein C

prevents blood clottinga major physiological anticoagulant.

a vitamin K-dependent serine protease enzyme that is activated by thrombin

Protein S

a vitamin K-dependent plasma glycoprotein synthesized in the endothelium

functions as a cofactor to Protein C in the inactivation of Factors Va and VIIIa

ThrombocytosisPlatelets still may not function right so you still

may have bleeding

_ PLT >450 x 109/L_ Reactive

_ Hemorrhage, surgery, iron def, inflammation_ Myeloproliferative disorders

_ ET, CML, PV_ May demonstrate bleeding or thrombosis

_ PLT >1 x 1010/L - _ Plateletphoresis may be required

Platelets

_ Normal - 2-4 um diameter_ Dense granules _ α-granules

_ Giant - >7um diameter_ MPD, MDS, May-Hegglin anomoly

_ Grey - _ Slightly inlarged_ Bernard-Soulier syndrome_ α-granule deficiency

Qualitative disordersDon’t work right

_ Storage pool deficiency_ Dense granules

_ Chediak-Higashi: albinism

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_ Wiskott-Aldrich: immunodef/severe eczema_ α granule_ Grey PLT syndrome: moderate bleeding_ Acquired - _ MDS, MDP

Metabolic PathwayDefects

_ Platelet release/secretion defects_ “Aspirin like”

_ Cycloxygenase_ Thromboxane synthetase

Qualitative disorders_ Receptor Defects

_ Mucocutaneous bleeding

Disorder Receptor lackingBernard-Soulier GP Ib/VGlanzmann’s thrombasthenia GPIIb/IIIa

Platelet Function “3A’s & 2R’s”_ Activation _ Adhesion _ Aggregation_ Release Reactions

_ Dense body and alpha granulecontents_ Retraction

Aspirin (3 things)blocks cycloxygenase

inhibits platelet activationaffects all platelets in circulation

General: Aspirinlike effect

_ Aspirin _ NSAIDs_ Antibiotics _ Herbs_ Garlic _ Vitamin E

Coumadin stays in system 2 months

Heparin stays in system 4 hours

The PFA-100 R System Assesses how aspirin affects platelet function

_ measures platelet plug formation_ small, whole blood sample (< 1 mL)_ reports a Closure Time in 4-8 minutes._ challenges platelets under highshear flow conditions_ hemodynamic conditions_ sensitivity of 95% following ingestion of a single 325 mg tablet of aspirin

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Platelet AggregometryMeasures aggregation

_ Qualitative function test_ Whole blood or PRP

_ Agonist (activator) is added_ Platelet clumping is recorder

Agonist aspirin/release defect

Membrane defect

storage pool defect

Thrombin normal normal ↓ Arachidonic

acid ↓ normal ↓

ADP, epi, collagen

↓ ↓ ↓

Specimen Collection_ Plasma

Lt Blue Tube9:1 blood to anticoag

_3.2% (0.109 M) Sodium Citrate - Buffered_Allow tube to fill_Short draws

• Excess anticoag • Erratic results_Invert to mix _Room temp_Centrifuge - _2500 x g for 10 min – we want plasma to be platelet free_Freeze rapidly at -70oC, then thaw rapidly too

Results of Hemolysis Activates clotting mechanism

Silicon coated and plastic tubes and pipets Prevent activationGlass and air can activate coag cascade

_ Whole bloodFor PFA 100 assay to replace bleeding time

Sodium Citrate_ Room temp _ No centrifugation

_ Run within 3 hours

_ Hct Adjustment

_ Hct >55%More anticoagulant goes into plasma

C=(1.85 x 103)([100-Hct]*V)C= Sodium Cit in mls V= whole blood in mls

Inhibitors of clotting

AT Heparin CofactorProtein C Protein STFPA Alpha 2 MacroglobulinAlpha 1 Antitrypsin C1 Inactivator

Inhibitors of Fibrinolysis PAI-1 PAI-2 PAI-3Alpha 2 Antiplasmin

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Specimen CollectionAvoid

_Hemolysis_Lipemia – can run on mechanical Fibromenter

(New analyzers have both mechanical and optical detection)_Traumatic draws

Activators of Coagulation Coagulation Factors PhospholipidsCa++

Job of activators and inhibitors To get clotting and fibrinolysis to balance

Activators of Fibrinolysis Factor X11a(12) Factor X1a(11)Kallikrein T-PA U-PA

Anticoagulant Therapy MonitoringMajor reason that we do coag testing in the lab

You want to

_ Oral: Warfarin (coumadin) - _PT with INR_ Standard unfractionated heparin - PTT, Chromogenic anti-

Xa_ LMWH(low molecular weight heparin): Enoxaparin, Tinzaparin - Chromogenic anti-Xa_ Direct thrombin inhibitors: Argatroban, Lepirudin,

Bivalirudin - _PTT

Hemorrhage and Coagulopathy

_ Bleeding - Local tissue injuries (not from coagulopathies)_ Suspect Coagulopathy

_Multiple sites _Spontaneous_Excessive _Recurrent

_ Most are Acquired_Drug interactions _Underlying systemic disorder

ThromboplastinGold standard

ISI 1.0 – most sensitive, most expensiveDon’t use over ISI 2.0

Coagulopathy Missing a factor

Primary hemostasis_Von Willebrand tests

_ Factor VIII activity_vWD Activity (ristocetin cofactor)

_vWF Antigen (Is it there?)Aggregomentry – how platelets respond to different stumuluses

Von Willebrand Disease Deficiency or abnormality of VWF

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Involved in Primary and SecondaryHemostasis

Multimeric protein necessary for PLT adhesionCarrier of factor VIII (8)Ineffective platelet aggregationLevels vary with blood type, common in OAcute phase reactantHuge platelet

Incidence - 1-2% in general population (inherited and acquired) Treatment - Desmopressin (DDAVP) nasal spray – stimulates any stored VWF to go into peripheral blood

ScurvyIncidence on rise

_Marked capillarybleeding_Progressive anemia

_ Vitamin C deficiency

Systemic (mucoscutaneous) Bleeding implies defect in 1o hemostasis

Bleeding Due to Factor Deficiency implies defect in 2o hemostasis

Test Menu _Bleeding Disorders

_Intrinsic - _PTT_Extrinsic - _PT

_Common - _Fibrinogen, _TT, _Reptilase Time (not prolonged by heparin)

Most common factor deficiency Factor 8

PTT↑, PT normal – Missing? 12, 11, 9, 8

PT↑, APPT normal – Missing? 7

PT↑, APPT↑ – Missing? 10, 5, 2, 1

Covalently links loose fibrin into strong clot Factor 13

Test Menu_ Fibrinolysis

_Fibrinogen activity _Reptilase time_TT _Euglobulin clot lysis time_Plasminogen activator inhibitor-1 (PAI-1)

Test Menu_ DIC

_↑D-dimer (quantitative) – for D and E fragmentsSpecific for covalently linked clot

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(many fibin stands in circulation) _↑PT _↑PTT _↑Fibrinogen Schistocytes in blood smear

What you use up when you make a clot 1, 13, 8 , 5, platelets

Thrombosis disorders: implies defects in fibrinolysis

Lupus Anticoagulant _ScreensCan develop antibodies against their factors

APPT↑_LA sensitive PTT (PTT-LA)Have clotting problems because of inhibition in the

fibrinolytic pathway

Mixing study (PTT-LA mixing study)½ patients + ½ normal blood

APPT↑ after mixing = inhibitor Normal results after mixing = factor deficiency

Treatment for DIC Heparin

Factor V Leiden

Defect in Factor V _ Inherited Condition_Seen mostly in northern European populations

_ Results in hypercoagulation and thrombosisFactor V resistant to Proteins C and S

Process does not get slowed down

Factor V Leiden

_Factor V has increased resistance to inactivation by protein C_Normally Protein C along with Protein S binds to the surface of a platelet, which degrades Factors V and VIII, preventing premature clotting_When Factor V Leiden is present, degradation is prevented and clotting occurs inside the veins

Factor V LeidenWho is affected?

6% of populationPregnant women or on Birth Ctrl pills

Causes thrombosis, even death

Symptoms

_ Pain swelling, and discoloration in legs, and/or arms due to clotting in the veins_ Extreme difficulty breathing due to pulmonary clotting_ Severe headaches due to clotting in the brain_ *There is no evidence showing that Factor V Leiden is associated with clots in the heart because it has its effects in the veins and not the arteries

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Increased Risks For Clotting

_Smoking _Overweight_Sitting or standing for long periods of time_Women with Factor V Leiden should not take the contraceptive pill_Pregnancy also increases the risk of thrombosis

Factor V LeidenTreatment

_Treatment may or may not be needed, depending on severity of disease (Homozygous or heterozygous)_If treatment is needed, long term anticoagulants are used_Heparin _Warfarin_Low-molecular-weight heparins

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