AfibrinogenemiaAfibrinogenemia

• Prevalence approx 1:1,000,000• Recessive inheritance

– Most reported cases from consanguineous parents– Parents typically have asymptomatic hypofibrinogenemia

• Genetically heterogeneous (>30 mutations)• May be due to failure of synthesis, intracellular transport or

secretion of fibrinogen• Moderate to severe bleeding (typically less than in severe

hemophilia)– Death from intracranial bleeding in childhood may occur– GI and other mucosal hemorrhage– Menorrhagia– Placental abruption

• Treat with purified fibrinogen concentrate or cryoprecipitate for bleeding, during pregnancy

Inherited dysfibrinogenemiaInherited dysfibrinogenemia

• Prevalance uncertain (most cases asymptomatic)• Usually exhibits dominant inheritance• Most cases due to missense mutations• Mutations may affect fibrin polymerization,

fibrinopeptide cleavage, or fibrin stabilization by FXIIIa

• Variable clinical manifestations (mutation-dependent):– Over 50% asymptomatic– Approx 25% with bleeding tendency (mild to severe)– 20% have a thrombotic tendency (arterial, venous, or both)

• Decreased thrombin-binding (antithrombin effect) of fibrin?• Altered fibrin clot structure?

Acquired dysfibrinogenemiaAcquired dysfibrinogenemia• Usually associated with liver disease

– Cirrhosis– Acute hepatitis– Hepatoma

• Fibrinogen has higher than normal sialic acid content• Diseased liver has increased levels of sialyltransferase and other

enzymes that control oligosaccharide content

JCI 1978;61:535

Diagnosis of dysfibrinogenemiaDiagnosis of dysfibrinogenemia

• Prolonged thrombin & reptilase times– PT, aPTT may be prolonged

• Disparity (>30%) between fibrinogen activity and antigen

• Family testing

• Evaluate for liver disease

Arch Pathol Lab Med 2002;126:499

Other acquired abnormalities of fibrin Other acquired abnormalities of fibrin formationformation

• High fibrinogen• High levels of FDP• Myeloma proteins (IgM > IgA > IgG)• Autoantibodies

Recessively inherited clotting factor Recessively inherited clotting factor deficienciesdeficiencies

• Rare– Exceptions: XI, XII deficiency

• Homozygotes (often consanguineous parents) or compound heterozygotes

• Heterozygous parents usually asymptomatic• Quantitative (“type 1”) deficiency: parallel reduction

in antigen and activity• Qualitative (“type 2”) deficiency: reduced activity

with near-normal antigen• Genetically heterogeneous• Complete deficiency of II, X not described (lethal?)• Mutation usually in gene encoding clotting factor

Exceptions: Combined V, VIII deficiencyCombined deficiency of vitamin K-dependent factors

Combined deficiency of factors V and VIIICombined deficiency of factors V and VIII

• Levels of affected factors 5-20% of normal

• Associated with mutations of LMAN-1 (ERGIC-53) or MCFD2, both of which regulate intracellular trafficking of V and VIII

Deficiency of multiple vitamin-K Deficiency of multiple vitamin-K dependent clotting factorsdependent clotting factors

• Levels of II, VII, IX, X, proteins C and S range from <1% to 30% of normal

• Bleeding symptoms proportional to degree of deficiency

• Usually associated with missense mutations in vitamin K epoxide reductase subunit 1 (VKORC1)

Relative frequencies of recessively Relative frequencies of recessively inherited factor deficienciesinherited factor deficiencies

Blood 2004; 104:1243

Clinical features of recessively inherited Clinical features of recessively inherited factor deficienciesfactor deficiencies

Blood 2004; 104:1243

Patterns of bleeding in recessively Patterns of bleeding in recessively inherited factor deficiency vs hemophiliainherited factor deficiency vs hemophilia

Blood 2004; 104:1243

Treatment of rare clotting factor deficienciesTreatment of rare clotting factor deficiencies

• FFP• Prothrombin complex concentrate (II, VII, IX, X) or

specific factor concentrate (XIII – others available in Europe) when appropriate

• Goal is to maintain “minimal hemostatic levels”• Antifibrinolytic drugs may be helpful in patients with

mucosal hemorrhage• Routine prophylaxis appropriate for F XIII deficiency

(long half-life, low levels adequate for hemostasis)• Otherwise treatment appropriate for active bleeding

or pre-procedure

Factor XIFactor XI

IX

X

Fibrinogen Fibrin

PT

XIa

Xa

V

VIII

XIInjury

TFVIIa

IXaVIIIa

XaVa

ThrombinPropagation

Initiation

Factor XI deficiencyFactor XI deficiency

• Recessively inherited• Most common in individuals of Ashkenazi

Jewish descent– 2 common mutations (one nonsense, one

missense)– Allele frequency as high as 10%, 0.1-0.3%

homozygous– Most affected patients compound heterozygotes

with low but measurable levels of XI activity

• Long aPTT, normal PT– XI activity < 10% in most patients with bleeding

tendency

Factor XI deficiencyFactor XI deficiencyClinical features & treatmentClinical features & treatment

• Variable, generally mild bleeding tendency– Bleeding after trauma & surgery– Spontaneous bleeding uncommon– Bleeding risk does not correlate well with XI level

• Treatment: FFP– 15 ml/kg loading, 3-6 ml/kg q 12-24h– Half life of factor >48 hours– Amicar useful after dental extraction, surgery– rVIIa is effective but expensive; thrombotic

complications reported

Factor XIIIFactor XIII

• Transglutaminase: forms amide bonds between lysine and glutamic acid residues

• Heterotetramer (A2B2) in plasma– A chains made by megakaryocytes and

monocyte/macrophage precursors– Platelet XIII (50% of total XIII) has only A chains– B chains (non-catalytic) made in liver

• Proenzyme activated by thrombin• Crosslinks fibrin

Inherited factor XIII deficiencyInherited factor XIII deficiency

• Autosomal recessive, rare (consanguineous parents)

• Heterozygous woman may have higher incidence of spontaneous abortion

• Most have absent or defective A subunit

• F XIII activity < 1%

Inherited factor XIII deficiencyInherited factor XIII deficiencyClinical features & treatmentClinical features & treatment

• Bleeding begins in infancy (umbilical cord)• Poor wound healing• Intracranial hemorrhage• Oligospermia, infertility• Diagnosis:

– Urea solubility test– Quantitative measurement of XIII activity– Rule out acquired deficiency due to autoantibody

• F XIII concentrates available (long half life, can administer every 4-6 weeks as prophylaxis)

Acquired factor XIII deficiencyAcquired factor XIII deficiency

• Autoantibody-mediated– Very rare

• Most patients elderly• May be drug-induced (isoniazid, other

antibiotics)• Bleeding may be severe• Diagnosis:

– Urea solubility– F XIII activity– Mixing study?

Factor XII deficiencyFactor XII deficiency

• Recessive inheritance, but fairly common

• Markedly prolonged aPTTNo bleeding tendency

• Deficiency of other contact factors (HMWK, prekallikrein) less common, but has similar phenotype

PLATELET DISORDERSPLATELET DISORDERS

Platelet structure 1Platelet structure 1

• Membrane glycoproteins– IIb-IIIa: integrin, cryptic in resting platelet,

after platelet activation binds fibrinogen and other adhesive proteins, necessary for aggregation

– Ib-IX-V: binds VWF, necessary for platelet adhesion at high shear rates

– Ia-IIa: integrin, binds collagen, mediates adhesion at low shear rates and platelet spreading (also acts as receptor)

Platelet structure 2Platelet structure 2

• Membrane receptors– Thrombin receptors (2): cleaved and

activated by thrombin– Thromboxane A2 receptor– ADP receptors (3)– Epinephrine receptor– Serotonin receptor– Cytokine, chemokine receptors– Fc receptor

Platelet structure 3Platelet structure 3

• Membrane phospholipids– Procoagulant lipids (phosphatidyl serine)

sequestered on inner leaflet, transferred to outer leaflet by “scramblase” enzyme with platelet activation

– Arachidonic acid cleaved from inner leaflet by phospholipase, converted to thromboxane by cyclooxygenase & thromboxane synthase

Platelet structure 4Platelet structure 4

• Granules– Dense granules: small molecules involved

in platelet activation (ATP/ADP, serotonin)– Alpha granules: fibrinogen, fibronectin,

thrombospondin, P-selectin, plasminogen, alpha-2 antiplasmin, factor V, PF4, PDGF, TGF-alpha and beta, ECGF

Bernard-Soulier syndromeBernard-Soulier syndrome• Pathophysiology:

– Deficiency of platelet membrane glycoprotein Ib-IX (VWF “receptor”)

– Defective platelet adhesion• Clinical: Moderate to severe bleeding• Inheritance: autosomal recessive• Morphology:

– Giant platelets– Thrombocytopenia (20-100K)

• Diagnosis: – No agglutination with ristocetin, decr thrombin response,

responses to other agonists intact– Morphology– Decreased GP Ib expression

Bernard-Soulier syndromeBernard-Soulier syndrome

Glanzmann thrombastheniaGlanzmann thrombasthenia

• Pathophysiology: – Deficiency of platelet membrane GPIIb-IIIa– Absent platelet aggregation with all agonists;

agglutination by ristocetin intact

• Clinical: Moderate to severe bleeding• Inheritance: autosomal recessive• Morphology: normal• Diagnosis:

– Defective platelet aggregation– Decreased GP IIb-IIIa expression

Gray platelet syndromeGray platelet syndrome

• Pathophysiology: Empty platelet alpha granules • Clinical: Mild bleeding• Inheritance: Autosomal dominant or recessive• Morphology:

– Hypogranular platelets– Giant platelets– Thrombocytopenia (30-100K)– Myelofibrosis in some patients

• Diagnosis– Variably abnormal platelet aggregation (can be normal)– Abnormal platelet appearance on blood smear– Electron microscopy showing absent alpha granules

Gray platelet syndromeGray platelet syndrome

Giant platelet syndromes associated Giant platelet syndromes associated with MYH9 mutationswith MYH9 mutations

1. May-Hegglin anomaly2. Fechtner syndrome3. Sebastian syndrome4. Epstein syndrome

• All associated with mutations in the non-muscle myosin heavy chain gene MYH9

• Thrombocytopenia with giant platelets, but mild bleeding

• Autosomal dominant inheritance• No consistent defects of platelet function detectable

in the clinical laboratory• Diagnosis usually based on clinical picture, family

history, examination of blood smear for neutrophil inclusions

Giant platelet syndromes associated Giant platelet syndromes associated with MYH9 mutationswith MYH9 mutations

Syndrome Neutrophil inclusions

Hereditary nephritis

Deafness

May-Hegglin

Yes No No

Fechtner Yes Yes Yes

Sebastian Yes* No No

Epstein No Yes Yes

*Neutrophil inclusions have different structure from those in May-Hegglin

Neutrophil inclusions in May-Hegglin Neutrophil inclusions in May-Hegglin anomalyanomaly

Wiskott-Aldrich syndromeWiskott-Aldrich syndrome

• Pathophysiology– Mutation in WASP signaling protein– Decreased secretion and aggregation with multiple

agonists; defective T-cell function• Clinical:

– Mild to severe bleeding– Eczema, immunodeficiency

• Inheritance: X-linked• Morphology:

– Thrombocytopenia (20-100K)– Small platelets with few granules

• Diagnosis: Family hx, clinical picture, genetic testing

Wiskott-Aldrich syndromeWiskott-Aldrich syndrome

Hermansky Pudlak syndromeHermansky Pudlak syndrome Chédiak-Higashi syndrome Chédiak-Higashi syndrome

• Pathophysiology: – Platelet dense granule deficiency: decreased aggregation &

secretion with multiple agonists– Defective pigmentation– Defective lysosomal function in other cells

• Clinical:– Mild to moderate bleeding– Oculocutaneous albinism (HPS)– Lysosomal storage disorder with ceroid deposition, lung & GI

disease (HPS)– Immunodeficiency, lymphomas (CHS)

• Inheritance: autosomal recessive• Morphology

– Reduced dense granules– Abnormal neutrophil granules (CHS)

• Diagnosis: clinical picture, neutrophil inclusions (CHS), genetic testing

Chédiak-Higashi, showing neutrophil inclusions

HPS, with oculocutaneousalbinism

Hermansky-Pudlak syndromeHermansky-Pudlak syndrome

Disaggregation after primary aggregation with ADP

Dense granule deficiency Control platelet

Br J Haematol 2007;138:671

Platelet type von Willebrand diseasePlatelet type von Willebrand disease

• Pathophysiology: Gain of function mutation in GP Ib, with enhanced binding to VWF and clearance of largest multimers from blood

• Clinical: Mild to moderate bleeding• Inheritance: Autosomal dominant• Morphology: Normal, but platelet count often low• Diagnosis: Variably low VWF antigen,

disproportionately low ristocetin cofactor activity, loss of largest VWF multimers on electrophoresis, enhanced platelet agglutination by low dose ristocetin (indistinguishable from type 2B VWD)

• Can distinguish from 2B VWD by mixing studies with normal/pt platelets and plasma and low dose ristocetin, or by genetic testing

Treatment of patients with platelet disordersTreatment of patients with platelet disorders

• Platelet transfusions

• DDAVP

• Antifibrinolytic drugs (Amicar)

• rVIIa?

• Treatment decisions must be individualized

VASCULAR DISORDERSVASCULAR DISORDERS

Hereditary Hemorrhagic TelangiectasiaHereditary Hemorrhagic Telangiectasia

• Autosomal dominant inheritance

• Mutation in endoglin gene that controls vascular remodeling– Molecular diagnosis possible

• Multiple small AVMs in skin, mouth, GI tract, lungs

Endoglin and vascular remodelingEndoglin and vascular remodeling

J Thromb Haemost 2010;8:1447

Hereditary hemorrhagic telangiectasiaHereditary hemorrhagic telangiectasia

J Thromb Haemost 2010;8:1447

Hereditary Hemorrhagic TelangiectasiaHereditary Hemorrhagic TelangiectasiaClinical featuresClinical features

• Epistaxis, GI bleeding – may be severe– Severe iron deficiency common

• Pulmonary or CNS bleeding often fatal• Gradual increase in bleeding risk with

age• AVMs enlarge during pregnancy• Risk of brain abscess• Hypoxemia from pulmonary HTN and

R→L shunting in lung

Hereditary Hemorrhagic TelangiectasiaHereditary Hemorrhagic TelangiectasiaTreatmentTreatment

• No consistently effective method for preventing bleeding

• Aggressive iron replacement• Antibiotic prophylaxis for dental work

etc• Screen for CNS lesions → consider

surgical intervention

Ehlers-Danlos syndromeEhlers-Danlos syndrome

• Defective collagen structure– Mutations in genes for various types of collagen

• 9 variants– Type IV (mutation in type III collagen gene) most

likely to cause bleeding

• Bleeding due to weakening of vessel wall → vessel rupture

• Conventional tests of hemostatic integrity normal

Ehlers-Danlos syndromeEhlers-Danlos syndrome

• Thin, weak skin with poor healing– “Cigarette paper” scars

• Bruising

• Hypermobile joints– Spontaneous joint dislocation

• Median survival 48 years in type IV EDS– Death from rupture of large vessels or

colon perforation