all about protein s

8/2/2019 All About Protein s

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Protein C deficiencyHereditary deficiencies occur in 0.14 - 0.5% of general population (the clinically

significant incidence is much lower); >160 mutations exist, either type I (76%, usuallyquantitative) or type II (dysfunctional protein, normal protein levels)

Causes 1-11% of cases of venous thrombosis; these patients are also at risk for warfarin-

induced skin necrosis if treated with warfarin and no heparin until warfarin levels aretherapeutic; this paradoxical clotting is due to a faster fall in natural anticoagulant proteins

than procoagulant proteins in these patients

Heterozygotes have levels 35-65% of normal; first thrombotic event occurs between ages10-50 years; only 30% have thromboembolism, increasing to 75% if coexisting factor V

Leiden

Homozygotes (1 per 500-750K births) with severely decreased levels present as newbornswith DIC and purpura fulminans neonatorum, leading to death unless anticoagulation and

replacement therapy with fresh frozen plasma is started

Must exclude acquired causes of protein C deficiency

• Acquired causes of low protein C levels: clot formation, surgery, liver disease,

warfarin (should be discontinued at least 10 days prior to testing) or Vitamin K antagonist therapy, DIC, vitamin K deficiency, L-asparaginase therapy

• Acquired causes of increased protein C (may mask protein C deficiency):

ischemic heart disease, pregnancy, postmenopausal women, hormone replacement

therapy, oral contraceptives

References: Archives 2002;126:1337

Protein S deficiencyHereditary deficiencies occur in 0.7% of general population; many mutations exist(qualitative or quantitative); much lower prevalence of thrombophilia with clustering in

families; variable penetrance may be due to coexisting risk factors, such as factor V Leiden

Causes 1-9% of cases of venous thrombosis; these patients also at risk for warfarin-inducedskin necrosis if started on warfarin without the addition of heparin until warfarin levels aretherapeutic

Heterozygotes have levels 35-65% of normal; first thrombotic event occurs between ages

10-50 years; 50% risk by age 45Homozygotes with severely decreased levels present as newborns with DIC and purpura

fulminans, leading to death unless anticoagulation and replacement therapy with fresh

frozen plasma is started1. Type I (2/3): low free and total protein S antigen, with decreased APC cofactor

activity

2. Type II (rare): normal free and total protein S antigen, and decreased APC

cofactor activity3. Type III (1/3): normal to low total protein S, low free protein S antigen, and an

elevated fraction of protein S bound to C4B protein

Testing recommended: individual with family history who requests testing, to confirmabnormal protein S result; must interpret with caution

Testing not recommended: during pregnancy or postpartum, during inflammatory,

thrombotic or surgical event; within 30 days of taking warfarin; must delay longer periodsfor vitamin K antagonists (Phenprocoumon)

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Clinical note: acquired protein S deficiency is often seen during pregnancy due to

increased C4b, which may reduce levels to 40% or less


Protein C assays

Deficiencies are either quantitative (type I, reduced amount of normal protein) or qualitative (type II, normal amount of defective protein) Assays are either functional

(measure protein activity) or antigenic (immunoassays that measure quantity, not function)

Perform functional assay first - if decreased, perform antigenic assay; must excludeacquired causes (below)

Low values should be confirmed on a new specimen

Assays should be performed with platelet poor plasma, using sodium citrate collectiontubes

Functional assays are clot-based or chromogenic

1. Clot based functional assays: detects all known type I and II variants; patient’s

protein C is activated by Southern Copperhead venom (Agkistrodon contortrix

contrortrix), which degrades synthetic substrate, factor Va or factor VIIIa with clot based PTT assay; the prolongation of clotting time is proportional to the amount of

factor activity2. PT based assay or amidolytic assays are affected by lupus anticoagulants (raises

protein C result), elevations of factor VIII > 200% (decreases the result), acute

phase reactions, factor V Leiden mutation (decrease the result); cannot perform on patients taking hirudin or argatroban

3. Chromogenic functional assays: not affected by lupus anticoagulants, factor VIII

levels, factor V Leiden or other coagulation abnormalities that interfere with clot- based functional assays; may not detect qualitative deficiencies detected by clot-

based assays; patient’s protein C is activated by snake venom, which cleaves a

synthetic substrate, which releases a chromogenic that is measuredspectrophotometrically

4. Antigenic assays: either ELISA, electroimmunoassay (Laurell rocket method) or

radioimmunoassay; variable levels, so use 3 standard deviations as cutoff

5. ELISA: uses antibody to protein C immobilized to microtiter place; add plasma; addsecondary anti-protein C antibody coupled to an enzyme for colorimetric detection;

use standard curve to determine plasma protein C

6. Laurell rocket antigenic assay: agarose gel has antibody to protein C; plasmasamples are put into wells and electrophoresed; antigen-antibody complexes

precipitate during electrophoresis, and height of precipitin arc is proportional to

plasma protein C, which is compared to standard curve using pooled normal

plasma; may be unable to detect protein C levels < 5%7. Radioimmunoassay: similar to ELISA, but uses single, radiolabeled antibody

Values falsely increased by bivalirudin, lepirudin, argatroban, and fondaparinux (Archives

2004;128:1142), lowered by warfarin (must discontinue for 10 days prior to testing)

Reference range: 70-140% of normal; newborns levels are 20-30% of adult values;

usually rise to near adult levels by age 6 months, but may remain below adult normal levels

until age 10 years

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Indications: necrotic skin in newborns days 1-3 of life (purpura fulminas neonatorum, can

also test parents also for heterozygosity); evaluation of cause of venous thromboembolism

(recommended to use chromogenic protein C assays initially)

Non-indications: screening before oral contraceptives or oral anticoagulants (discontinue

for 10 days, or test family members)

• Acquired causes of low Protein C levels: more common than hereditary

deficiencies - clot formation, surgery, liver disease, warfarin (should be

discontinued at least 10-30 days prior to testing), DIC, vitamin K deficiency,vitamin K antagonist therapy, L-asparaginase therapy; repeat protein C test once

these conditions are no longer present

• Acquired causes of increased Protein C (may mask protein C deficiency):

ischemic heart disease, pregnancy, postmenopausal women, hormone replacement

therapy, oral contraceptives


Protein S assaysDeficiencies are either quantitative (type I, reduced normal protein) or qualitative (type II,

normal amount of defective protein)Assays are either functional (measure protein activity) or antigenic (immunoassays that

measure quantity, not function)

Gold standard to measure free protein S or APC cofactor activity of protein S is consideredthe polyclonal ELISA with or without polyethylene glycol precipitation, although this

procedure has poor reproducibility

Perform functional assay first (detects all types of deficiencies)

Functional assays are clot-based, cannot be performed in patients taking hirudin or argatroban

Methodology: clot based protein S method is based on the addition of activated protein C,which in the presence of protein S, accelerates the inhibition of thrombin-activated factors

VIII and V; the prolongation of clotting time is proportional to the amount of factor S

activity; interference may occur with elevated factor VIII (acute phase reactions or otherwise); values falsely increased by bivalirudin, lepirudin, argatroban, and fondaparinux

(Archives 2004;128:1142), lupus anticoagulants

Reference ranges are in nmol/liter (each lab should establish its own, values in acute

phase plasma are higher):

• Total protein S:- 65% of value in pooled normal human plasma (289-397)

•Free protein S: 71-115

• C4 binding protein beta+: 228-310

• Total C4 binding protein: 257-423

•

Antigenic assays measure free protein S (functionally active form) or total (bound plusfree) protein S - usually 60% of protein S is bound to C4b-binding protein

Free protein S levels in protein S deficient patients are very sensitive to timing, temperature

and dilutional conditions of assays compared to normal individuals

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Acquired causes of low Protein S levels: more common than hereditary deficiencies - clot

formation, surgery, liver disease, warfarin (should be discontinued at least 10 days prior to

testing), nephrotic syndrome, DIC, L-asparaginase therapy, any stimulus to acute phaseresponse (increases C4b binding protein, decreases free protein S), newborns (12-60% of

adult levels, rise to adult levels by 6 months), women (lower than men before menopause,

while taking oral contraceptives, during pregnancy or with hormone replacement therapy),vitamin K antagonist drugs, vitamin K deficiency, elevated factor VIII levels (>200%) in

PTT based functional assays, thrombosis; also nephrotic syndrome, varicella infection,

HIV infection

Classification of deficiencies: all have low functional protein S

• I - also low free and total protein S

• II / IIb - also normal free and total protein S

• III / IIa - low free but normal total protein S

Protein C / Protein S anticoagulant pathwayPathway is a physiologic anticoagulant system to limit blood clot formation (i.e. fibrinogen

to fibrin conversion) to site of vessel injuryMajor anticoagulant systems are protein C and protein S, antithrombin and tissue factor

pathway inhibitor (TFPI, see Extrinsic pathway above)

Protein C and S: vitamin K dependent anticoagulant proteins produced mainly in liver (“C” because was third peak to elute from a diethylaminoethyl affinity column)

Activation: endothelial cell protein C receptor binds thrombin-thrombomodulin complex,

which activates protein C, which binds to free protein S on endothelial or platelet phospholipids surfaces; this protein C / protein S complex degrades factors Va and VIIIa,

which reduces fibrin formation

Activated protein C also indirectly promotes fibrinolysis60-70% of protein S is bound to and inactivated by C4b binding protein, an acute phase

reactant

Clinical note: since C4b increases during pregnancy, the protein S level will routinely fall

below the normal non-pregnant range

Diagram: Protein C / Protein S anticoagulant pathway


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Protein C / Protein S anticoagulant pathwayThrombin-thrombomodulin (TM) complex activates protein C. Activated protein C with its cofactor, free

protein S, degrades factors Va and VIIIa. In addition, when thrombin binds thrombomodulin, thrombin

loses its procoagulant functions. Reprinted with permission from Van Cott and Laposata.1

Thrombomodulin

Intrinsic membrane glycoprotein on luminal surface of endothelial cells that bindsthrombomodulin and facilitates the activation of protein C

C/T dimorphism at nucleotide 1418 is associated with premature myocardial infarction, butno definite association with venous thromboembolism

Drawings: thrombomodulin protein, flowchart of protein C activation

References: BMC Neurology 2004;4:21

Antithrombin

Formerly called antithrombin III

Functions as anticoagulant by inhibiting activated factors II (thrombin), IX, X, XI, XII,kallikrein, plasmin and probably factor VII (all are serine proteases)

Activity is accelerated 1000x by interaction with heparin or heparan sulfate (located on

endothelial cells)

Member of serine protease inhibitor (serpin) gene family on #1q23-25


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Fibrinolysis pathway

Process of degrading the fibrin clot when it is no longer needed

Also prevents extension of clot beyond site of injuryInitiated by tPA (tissue plasminogen activator) or uPA (urokinase-like plasminogen

activator), which convert plasminogen to plasmin in the presence of fibrin by cleaving the

Arg561-Val562 peptide bondPlasmin degrades the fibrin clot and intact fibrinogen to soluble fibrin/fibrinogen

degradation products (FDP)

Plasmin also inactivates factors Va and VIIIa (as does Protein C and Protein S)tPA is produced by endothelial cells; its activation of plasminogen is major mechanism for

lysis of fibrin clots

Recombinant tPA is used to treat myocardial infarction, stroke and some cases of acute

thrombosisuPA is produced by urine and plasma; keeps renal tracts free of blood clots; also is

important for other cell surfaces and initiating nonfibrinolytic activities of plasmin

Excessive fibrinolysis is prevented by plasmin inhibitor (antiplasmin, formerly called

alpha2-antiplasmin) and plasminogen activator inhibitor 1 (PAI-1, inhibits tPA and uPA)PAI-1 is synthesized by hepatocytes and endothelial cells, is present in platelets and

plasma; can bind to fibrin and inhibit plasminogen activators tPA and uPAPAI-1 is an acute phase reactant protein, and may increase 30-50 fold over baseline,

possibly immediately inactivating systemically administered tPA

Homozygous deficiency of plasminogen is associated with ligneous conjunctivitis (rareform of chronic pseudomembranous conjunctivitis), and replacement therapy with

plasminogen is therapeutic

Neither heterozygous plasminogen deficiency (0.5 to 2.0% of patients with thrombosis) nor

tPA deficiency are associated with increased risk of thrombosis

Diagrams: coagulation cascade and fibrinolytic system, diagram


flowchart of protein C activation

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http://depts.washington.edu/labweb/dept/staff/bios/hemostas/images/introfibsys_figure_1.jpg










coagulation cascade and fibrinolytic system, diagram

coagulation cascade

The coagulation cascade is initiated by the exposure of blood to tissue factor (TF). The factor VIIa–TFcomplex then activates both factors IX and X. Factor IXa also converts factor X to Xa in the presence of

factor VIIIa, phospholipid (PL), and calcium (Ca2+). Factor Xa then converts prothrombin (II) to thrombin

(IIa) in the presence of factor Va, PL, and Ca2+. Thrombin then converts fibrinogen to fibrin, which

polymerizes to form the fibrin clot (Fibrin (n)). The generation of thrombin is amplified by the feedback

activation of factors V, VIII, and XI mediated by thrombin itself. In addition, thrombin activates factor

XIII, which stabilizes the fibrin clot. The fibrinolytic system is one of the key regulatory pathways of thecoagulation cascade. Plasminogen is converted to plasmin by tissue-type plasminogen activator (tPA) in the

presence of fibrin. Plasmin then digests the fibrin clot, forming soluble fibrin degradation products (FDPs)

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Acquired bleeding disorders

Liver dysfunction

Liver is site of production of most coagulation factors, but response of each factor to liver

disease is variable due to differences in biologic half lives and acute phase reactionsPT usually prolonged first, then PTT

• Factor VII: shortest biologic half life, often affected earliest with largest decrease

in serum level Clinical note: Factor VII also decreases earliest with warfarintreatment

• Factor VIII: may be normal or elevated due to acute phase reaction

• Factors XI and XII: long biologic half lives, may be normal until liver disease isadvanced

Proteinuria

Patients with nephrotic syndrome may have decreased factors XI and XII

Hereditary bleeding disordersFactor XIII deficiency

Rare

Autosomal inheritance

Need 5-50% for surgical hemostasis, biologic half life is 9-12 daysAlthough fibrin clots form, they are weak and subsequently lyse

Normal PT and PTT

Patients commonly present with history of delayed bleeding; often associated with

bruising, epistaxis, menorrhagia, GI/GU bleeding, umbilical stump bleeding, miscarriage,intracranial hemorrhage, poor wound healing; also bleeding after surgery, trauma, dental

procedures, pregnancy or circumcision

Testing recommended if delayed bleeding, umbilical stump bleeding, or miscarriages (withnormal PT and PTT)

Diagnosis: usually with “urea clot lysis” assay, although specific factor assay is available

in specialized labs50% of population has Val134Leu polymorphism, which may protect against deep venous

thrombosis, but predispose to intracranial hemorrhage

Acquired causes of factor XIII deficiency: liver disease, DIC, Crohn’s disease, ulcerative

colitis, Henoch-Schonlein purpura, leukemia, myelodysplasia, myeloproliferative disorders

Treatment: 500 ml plasma or 1 bag cryoprecipitate / 10 kg every 3 weeks

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von Willebrand’s diseaseMost common hereditary bleeding disorder, affecting 1-2% of population, no gender

preferenceOften mild and undiagnosed; may be masked by acute phase reactions

Unmarked vWD is still a common underlying cause of hysterectomy; test women with

menorrhagia during first few days of periodDue to quantitative or qualitative deficiencies of von Willebrand factor (vWF), found on

#12

Symptoms are similar to a platelet function defect (epistaxis, easy bruising, bleeding,menorrhagia)

vWF is synthesized by (a) endothelial cells, stored in Weibel-Palade bodies, secreted into plasma and subendothelium and (b) megakaryocytes, present in platelets in alpha granules

vWF is large polypeptide that polymerizes to form multimers of up to 100 subunits

Plays a role in platelet plug and fibrin clot, both essential to hemostasis at site of

endothelial injury, particularly in high flow vessels

vWF mediates platelet adhesion to endothelium (and formation of platelet plug) by servingas a bridge between them - binds to GPIb glycoprotein on platelet surface and to exposed

subendothelium at site of endothelial injuryvWF supports coagulation (fibrin clot) by serving as protective carrier protein for factor

VIII; without vWF, factor VIII has shorter half life and its plasma levels are lower

Note: type O patients have lower levels of vWF, type AB patients have highest levels;levels increase with age and with acute phase reactions

Treatment:

DDAVP (desmopressin) temporarily increases vWF and factor VIII levels 2-3x; for the patients that don’t respond, give vWF-containing factor VIII concentrates

Testing:

Factor VIIII activity, vWF antigen, vWF activity (often done by “ristocetin

cofactor” assay); possible additional tests include vWF multimer size

determination and blood type (vWF is significantly decreased in type O patients)

Repeat testing is often required because vWF and factor VIII become

elevated during minor illnesses, injury, stress, pregnancy, estrogen use,other acute phase reactions or in newborns

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Diagrams: binding of von Willebrand factor

Subtypes

Clinical note: As a general rule in coagulation, type I deficiencies refer to a decrease in the

absolute amount of a normal factor and type II deficiencies indicate a defective protein thatmay be present in normal amounts

Type 1 (70-80%): most common, autosomal dominant, partial quantitative deficiency of

vWF but normal function; causes mild/moderate bleeding disorder

Low factor VIII, vWF antigen and ristocetin cofactor; low/normal ristocetin induced platelet aggregation, normal or all sizes decreased in multimer analysis, mean ristocetin

cofactor/vWF antigen ratio is 1.0; normal platelet count

Type 2 (15-20%): qualitative deficiency of vWF, variable quantitative deficiency, usually

mild/moderate bleeding disorder, but may be severe

• Type 2A: most common type 2 subtype; autosomal dominant; low/normal factor VIII activity and vWF; relative reduction of intermediate and high molecular

weight multimers due to in vivo proteolytic degradation or defective multimer

assembly and secretion; markedly reduced ristocetin cofactor, low ristocetininduced platelet aggregation; platelet vWF has similar abnormalities as plasma

vWF; mean ristocetin cofactor/vWF antigen ratio is 0.3; normal platelet count

• Type 2B: autosomal dominant, hemostatic defect due to intermittent

thrombocytopenia and qualititatively abnormal vWF, with increased binding of

vWF to GPIb (platelet vWF receptor), causing faster clearing of vWF coated

platelets from the bloodstream; the platelet count drops further during pregnancy,surgery, DDAVP therapy; have low/normal factor VIII activity and vWF; reduction

of high molecular weight multimers but increase in low molecular weightfragments; reduced ristocetin cofactor but increased ristocetin induced platelet

aggregation; mean ristocetin cofactor/vWF antigen ratio is 0.6

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• Type 2C: autosomal recessive; reduction of high molecular weight multimers,

increase in small multimers and qualititatively abnormal individual multimers;reduced ristocetin cofactor activity out of proportion to reductions in vWF

• Type 2M: rare; autosomal dominant, decreased platelet directed function NOT due

to a decrease of high molecular weight multimers, but otherwise similar to type 2A(may be due to mutation that impairs vWF and GPIb binding); low/normal factor

VIII and vWF, normal multimer analysis, but very low ristocetin cofactor and

low/normal ristocetin induced platelet aggregation; mean ristocetin cofactor/vWFantigen ratio is < 1.0

• Type 2N: rare, autosomal recessive; markedly reduced affinity of vWF for factor VIII, causes reduction of factor VIII levels to 5% of reference range; other vWF lab

tests are normal; often misdiagnosed as hemophilia A (which is X linked recessive),

but males and females in type 2N are equally affected; assay that measures bindingof factor VIII to vWF is available in specialized laboratories

Type 3: very rare; autosomal recessive, often associated with consanguinity; most

severe clinical bleeding; homozygous patients have marked deficiencies of plasmavWF and factor VIII activity, no vWF in platelets and endothelial cells, no secondary

transfusion response, no response to DDAVP; also undetectable ristocetin cofactor, low

ristocetin induced platelet aggregation, all multimer sizes are absent

Platelet type or pseudo von Willebrand’s disease: rare disorder of mutation in GPIb

(not vWF gene), causing increased binding of vWF to GPIb, with similar clinicalfindings as type 2B

Molecular basis of von Willebrand’s diseaseType 1: mutations throughout the gene, not well characterized

Type 2A: mutation in proteolysis site (most common type 2A mutation)

Type 2A: loss of propeptide, required for multimer formation from dimers

Type 2A: mutation in C-terminus, required for dimer formation from monomersType 2B: mutation in GPIb binding site, causing increased binding of vWF to GPIb

Type 2M: mutation in GPIb binding site, causing decreased binding of vWF to GPIb

Type 2N: mutation in N-terminis (factor VIII binding site), leading to decreased binding of vWF to factor VIII

Type 3: mutations throughout the gene, not well characterized

Common pathwayInvolves fibrinogen (factor I), factors II (prothrombin), V, X

Thrombin converts soluble fibrinogen to insoluble fibrin; releases fibrinopeptides A and B;

remaining fibrin monomers polymerize to form fibrin; thrombin also binds to antithrombin,which inhibits thrombin to prevent excessive clotting

Thrombin may also activate factor XI (part of intrinsic pathway), factors V, VIII, XIII, XI

and plateletsFactor XIII cross links fibrin to increase stability of fibrin clot

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CoagulationAcquired bleeding disorders : Disseminated intravascular coagulation (DIC)

Definition and pathophysiology

● A common acquired syndrome arising from various causes (see etiology below), characterized by

massive, sustained and excessive activation of coagulation with the eventual inundation(overwhelming) of the anticoagulant and fibrinolytic systems● Leads to disseminated microthrombi and tissue ischemia; consumption of platelets, coagulationfactors and natural anticoagulants; and variable bleeding● Activation of inflammatory pathways via cytokines also plays a role● Ultimately free, circulating, unopposed thrombin and plasmin are generated (the two key agentsresponsible for DIC) which then leads to:- Activation and consumption of platelets, coagulation factors, fibrinogen and fibrin- Consumption and depletion of anticoagulant proteins (protein C, protein S and antithrombin)- Generation of D-dimers and fibrin degradation productsn- Formation of microthrombi leading to tissue ischemia (large thrombi can also be formed,particularly in cancer patients)- Schistocytes (fragmented red blood cells) are formed as red blood cells are severed flowing

through fibrin strands (microthrombi within vasculature)- Variable bleeding

Diagrams Key event in DIC

Epidemiology● 1% of hospitalized patients are estimated to develop DIC● 20% of patients with Acute Respiratory Distress Syndrome (ARDS) develop DIC and 20% of patients with DIC develop ARDS● 20% of patients with gram-negative sepsis develop DIC

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Other causes● Infection/sepsis: most common cause; includes bacterial, viral, fungal, rickettsial and protozoalorganisms● Tissue damage: trauma, burns● Malignancy: solid and hematologic● Obstetric complications: abruptio placentae, retained dead fetus syndrome,

preeclampsia/eclampsia, amniotic fluid embolism, acute fatty liver of pregnancy, septic abortion● Miscellaneous: near drowning, fat embolism, snake bites, aortic aneurism, acute hemolytictransfusion reaction, adult respiratory distress syndrome, giant hemangioma, homozygous proteinC deficiency)

Clinical features ● Patients can have either bleeding or thrombosis or both● Septic patients are more likely to have thrombosis than bleeding● If severe and prolonged, will eventually lead to multiorgan dysfunction/failure● Causes of DIC can be acute (meningococcemia) or chronic (retained dead fetus), localized(abdominal aortic aneurysm) or systemic (acute promyelocytic leukemia)● Chronic causes of DIC are typically malignancy, liver disease, retained dead fetus syndrome,abdominal aortic aneurysm, giant hemangioma and head trauma● Bleeding can present as surgical site, venipuncture site or mucocutaneous bleeding (mostcommon); gastrointestinal bleeding, CNS bleeding, hematuria or ecchymoses● Thrombosis can present as purpura fulminans (manifestation of subdermal microthrombi with skinnecrosis); cold, pulseless limb; sudden loss of vision; oliguria; mental status changes, seizures,behavioral changes or adrenal insufficiency

Laboratory ● Prolonged PT and PTT● Elevated D-dimers (Am J Clin Pathol 2004;122:178) and other fibrin degradation products (but D-dimer may be falsely positive in HIV+ Castleman’s disease due to interference from monoclonalgammopathy (Arch Pathol Lab Med 2004;128:328)● Fall in platelet count (usually not lower than 30,000-40,000 x 109/L)● Drop in fibrinogen● Presence of schistocytes on peripheral blood smear (not specific for DIC)● With chronic causes, fibrinogen and platelets may actually be elevated as acute phase reactants● All coagulation factors may be variably decreased due to factor activation and consumption● Multiorgan dysfunction may manifest as elevated cardiac enzymes or elevated BUN/creatinine● Baseline coagulation studies and serial follow-up are needed to follow the trends

Prognostic factors ● One multicenter study of critically ill patients with DIC found that the 28 day mortality was 21.9%,which was significantly higher than non-DIC patients (11.2%) (Crit Care Med 2008;36:145)● Another study found the mortality rate was significantly higher in sepsis patients than traumapatients (Thromb Haemost 2008;100:1099)

Treatment ● Treat underlying disease

● Keep fibrinogen levels above 100 mg/dL with cryoprecipitate or fresh frozen plasma● Monitor PT, PTT, platelet count, fibrinogen and possibly antithrombin levels● If bleeding predominates, replace coagulation factors and fibrinogen with fresh frozen plasma(FFP) and cryoprecipitate; consider plasmapheresis, platelet-transfusions and immunoabsorption● If platelet count is lower than 50,000 x 109/L with active bleeding, or lower than 10,000 x 109/L,give platelet infusion● If thrombosis predominates (chronic DIC), heparinization should be considered

Differential diagnosis

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● Severe hepatic cirrhosis● Dilutional coagulopathy (but may coexist with DIC)● HELLP syndrome (H-hemolysis; EL-elevated liver enzymes; LP-low platelets); can alsodegenerate to DIC● TTP/HUS

Thromb Haemost. 2001 Nov;86(5):1144-7.http://www.ncbi.nlm.nih.gov/pubmed/11816698

Analytical considerations for free protein S assays in protein S

deficiency.Persson KE, Hillarp A, Dahlbäck B.

Department of Clinical Chemistry, Lund University, University Hospital, Malmö, [email protected]

Abstract

Protein S is an anticoagulant protein that circulates in plasma in complex with C4b-binding

protein (C4BP) or in free form. Deficiency of protein S increases the risk of venousthrombosis. Measurement of free protein S, as compared to total levels, has been shown to

be superior for prediction of protein S deficiency. We studied the effects of different

handling protocols for an immuno- and a ligand (C4BP)-based assay for free protein S.When the assay was performed at 37 degrees C, the levels of free protein S in plasma from

protein S deficient patients were approximately twice those obtained at room temperature.

The reason for this phenomenon was that plasmas from protein S deficient patientsexhibited a time-, temperature-, and dilution-dependent increase in free protein S, which

was more pronounced than corresponding dilution of the normal plasma that was used tocreate the standard curve. These findings demonstrate the importance of assay procedure

and sample handling in assays for free protein S

Indian Journal of Pediatrics

Volume 70, Number 11, 903-907, DOI: 10.1007/BF02730597

Neonatal thrombosis

Abstract

Neonatal thrombosis is a serious event that can cause mortality or result in severe

morbidity and disability. The most important risk factor for the development of thrombosis during the neonatal period is the presence of an indwelling central line andconsequently the vessels involved tend to be those most frequently used for catheterization. Other documented risk factors for the development of neonatalthrombosis include asphyxia, septicemia, dehydration, maternal diabetes and cardiacdisease. Main laboratory findings for the diagnosis of hypercoagulable states, includeshortened aPTT, decreased levels of inhibitors (AT III, Protein C and Protein S),increased resistance to activated protein C, defective fibrinolysis (basal and after stimuli), increased levels of clotting factors (fibrinogen, factor VII, factor VIII, etc.),

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http://www.ncbi.nlm.nih.gov/pubmed?term=%22Persson%20KE%22%5BAuthor%5D

http://www.ncbi.nlm.nih.gov/pubmed?term=%22Hillarp%20A%22%5BAuthor%5D

http://www.ncbi.nlm.nih.gov/pubmed?term=%22Dahlb%C3%A4ck%20B%22%5BAuthor%5D

http://www.springerlink.com/content/0019-5456/

http://www.springerlink.com/content/0019-5456/70/11/

http://www.ncbi.nlm.nih.gov/pubmed?term=%22Persson%20KE%22%5BAuthor%5D

http://www.ncbi.nlm.nih.gov/pubmed?term=%22Hillarp%20A%22%5BAuthor%5D

http://www.ncbi.nlm.nih.gov/pubmed?term=%22Dahlb%C3%A4ck%20B%22%5BAuthor%5D

http://www.springerlink.com/content/0019-5456/

http://www.springerlink.com/content/0019-5456/70/11/



increased and/or hyperactive platelets, increased whole blood and/or plasma viscosity, Antiphospholipid antibodies and presence of prothrombotic molecular defects like FVLeiden, P20210 and MTHFR. Approximately 4% and 2% respectively of Caucasians areheterozygous for these gene defects. Their causative role in neonatal thrombosis isunknown but they may have a contributory role in the pathogenesis of thrombosis inneonates.

Hereditary thrombophilia due to congenital protein S deficiency

Summary

Congenital protein S deficiency is an inherited coagulation disorder characterized byrecurrent venous thrombosis symptoms due to reduced synthesis and/or activity levelsof protein S. Prevalence of partial protein S deficiency (heterozygous individuals) isestimated at 0.16-0.21% in the general population. Prevalence of severe protein Sdeficiency (homozygous or compound heterozygous individuals) is unknown but isprobably comparable to that of severe protein C deficiency which is estimated at

1/500,000. Men and women are equally affected. In severe protein S deficiency, thedisease manifests several hours to days after birth, with purpura fulminans (see thisterm) or massive venous thrombosis. Purpura fulminans is a life-threatening conditioninvolving severe clotting throughout the body and causing necrosis of tissues. Severeretinopathy of prematurity (ROP) (see this term) may also occur. Heterozygous patientsare usually asymptomatic until adulthood. Thrombotic episodes are mainly provoked byother risk factors such as surgery, pregnancy or immobilization. Deep vein thrombosis of the lower limbs with or without pulmonary embolism is the most common manifestationof the disease. Arterial thrombosis may also occur. Protein S deficiency is caused bymutations in the PROS1 gene (3q11-q11.2). Transmission is autosomal recessive.Diagnosis is based on the measurement of protein S antigen levels (total protein S or free protein S) and anticoagulant activity. There are three biological forms. Type I and

type III are quantitative deficiencies with low free antigen levels (with normal total proteinS levels in type III and decreased total protein S levels in type I deficiency). Type II is aqualitative deficiency with normal total and free protein S levels. Molecular testing isavailable, but is unnecessary for diagnosis. Differential diagnoses include other inheritedthrombophilias including antithrombin and protein C deficiencies (see these terms).

Antenatal diagnosis is feasible for families with affected children and is based on theidentification of the causal mutation on DNA obtained by chorionic villus sampling.

Administration of fresh frozen plasma may be required for the initial treatment of neonatal purpura fulminans. Surgical procedures may be required for excision of thrombotic lesions. Patients with thromboses are treated with anticoagulant therapy(heparin, wafarin). Attention should be paid to the risk of coumarin-induced skinnecrosis. Preventive treatment is indicated in cases with strong positive family history of thrombotic diseases, during the peripartum period or perioperatively. Prognosis is severein homozygous or compound heterozygous patients. Prognosis is good for heterozygouspatients. With adequate treatment and monitoring, the risk of thromboembolic disease ismarkedly reduced. Mortality may result from pulmonary embolism. *Author: Prof. J.Goudemand (November 2009)

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