1 Comment

CHAPTER 127 HEREDITARY THROMBOPHILIA

CHAPTER 127 HEREDITARY THROMBOPHILIA
Williams Hematology

CHAPTER 127 HEREDITARY THROMBOPHILIA

SCOTT H. GOODNIGHT
JOHN H. GRIFFIN

Introduction
Major Hereditary Defects

Hereditary Resistance to Activated Protein C

Prothrombin G20210A Gene Polymorphism

Hyperhomocysteinemia

Protein C Deficiency

Protein S Deficiency

Antithrombin Deficiency

High Levels of Factor VIII and Other Coagulation Factors

Hereditary Thrombotic Dysfibrinogenemia

Other Potential Thrombophilic Disorders
Diagnosis of Thrombophilia
Therapy of Thrombophilia
Thrombophilia and Pregnancy, Oral Contraceptives, and Hormone Replacement Therapy

Management of Pregnancy in Thrombophilic Women

Pregnancy Loss due to Thrombophilia

Estrogens: Oral Contraceptives and Hormone Replacement Therapy
Chapter References

Hereditary thrombophilia is defined as a genetically determined increased likelihood of thrombosis. An emerging paradigm suggests that thromboembolism is a multicausal disease involving one or more genetic defects in conjunction with acquired risk factors such as inactivity, trauma, malignancy, inflammation, pregnancy, oral contraceptive use, or autoimmune disease. The three most common hereditary defects (found in a substantial proportion of patients presenting with venous thrombosis) include activated protein C resistance caused by replacement of Arg506 by Gln in the factor V gene (factor V Leiden), a prothrombin polymorphism (G20210A) that causes elevated plasma prothrombin levels, and hyperhomocysteinemia. Additional genetic abnormalities include deficiencies of the anticoagulant factors protein C, protein S, or antithrombin. The majority of these thrombophilic defects either enhance procoagulant reactions or hamper anticoagulant mechanisms and thus cause a prothrombotic state due to hypercoagulability of the blood. Venous thrombosis is the most common manifestation of hereditary thrombophilia, although a minority of patients, particularly those with other vascular risk factors, may develop arterial thrombi as well. Less usual presentations of venous thromboembolic disease include abdominal and cerebral vein thrombosis, along with pregnancy loss or other complications due to placental vascular insufficiency and thrombosis. Laboratory assays are now widely available to identify the great majority of patients with thrombophilia. Knowledge of these disorders affects patient management, including the duration of anticoagulant treatment, the use of clotting factor replacement therapy, the need for prophylactic antithrombotic agents, and counseling involving the relative risks of pregnancy and use of oral contraceptives or hormone replacement.

Acronyms and abbreviations that appear in this chapter include: APC, activated protein C; C4BP, C4b-binding protein; DVT, deep vein thrombosis; MTHFR, methylenetetrahydrofolate reductase; PAI-1, plasminogen activator inhibitor 1; PCR, polymerase chain reaction.

INTRODUCTION
Hereditary thrombophilia is defined as a genetically determined increased risk of thrombosis. According to Virchow’s classic (and still useful) triad, risk factors for thrombosis may involve abnormalities in the vessel wall (see Chap. 114), rheology, and/or blood components. Identification of defects in specific blood components, especially plasma factors, has provided molecular insights into the pathogenesis of thrombophilia. The major defects associated with familial thrombophilia are listed in Table 127-1. The first description of hereditary thrombophilia caused by a deficiency of an anticoagulant protein was by Egeberg in 1965.1 Members of the family described in the report suffered from recurrent venous thrombosis, and the disorder was inherited in an autosomal dominant pattern. The plasma of affected family members had reduced amounts of an inhibitor to thrombin, antithrombin III. In 1976, Stenflo and coworkers2 purified and characterized an anticoagulant factor, protein C, from bovine plasma, and subsequently the first patients with hereditary protein C deficiency and thrombosis were described by Griffin and colleagues.3 Three years later protein S deficiency was reported in several families with thrombosis by Schwarz and coworkers4 and Comp and coworkers.5,6 In 1993 Dahlback and coworkers reported three families with venous thrombosis associated with hereditary resistance to activated protein C (APC),7,8 and in 1994 the underlying genetic defect was simultaneously reported by th ree laboratories to involve the factor V mutation of Arg506 to Gln, a defect now often referred to as factor V Leiden.9,10 and 11 At about the same time, mild to moderate hyperhomocysteinemia was also recognized as a risk factor for venous thrombosis,12 although a predisposition to arterial vascular disease due to elevated levels of homocysteine had been known for some time.13 In 1996, a mutation in the 3′-untranslated region of prothrombin was identified and linked to familial venous thromboembolism by Poort and colleagues.14 Many important observations have come from the Leiden Thrombophilia Study of 300 to 500 Dutch consecutive patients presenting with a first episode of venous thrombosis.15,16 With the imminent characterization of about 100,000 human genes, one can anticipate a steady stream of advances in the identification of more genetic defects responsible for hereditary thrombophilia.

TABLE 127-1 MAJOR HEREDITARY DEFECTS ASSOCIATED WITH INCREASED RISK OF VENOUS THROMBOSIS

Specific hereditary thrombophilias can now be identified in 30 to 50 percent of patients presenting with a first episode of venous thromboembolism, with even higher percentages found in subjects with recurrent thrombosis. Patients with hereditary thrombophilia may have more than one hereditary thrombophilia and associated acquired abnormalities such as antiphospholipid antibodies, malignancy, myeloproliferative diseases, or inflammatory disorders (Fig. 127-1). Hereditary prothrombotic states are usually associated with venous rather than arterial thrombosis; however, in association with other risk factors such as smoking or diabetes, recent data suggest that up to 10 percent of arterial thromboses are associated with hereditary thrombophilia.17,18

FIGURE 127-1 Paradigm for genetic contribution to venous thrombosis. Clinically significant venous thrombosis most often follows from the simultaneous presence of an acquired risk factor for thrombosis and one or more genetic factors that convey thrombotic risk. The presence of two genetic factors (i.e., gene-gene interaction) greatly increases the likelihood of thrombosis. Mild genetic risk factors include APC resistance with or without factor V Leiden; the prothrombin G20210A polymorphism causing elevated plasma prothrombin levels; and heterozygous deficiency of protein C, protein S, or antithrombin. Hyperhomocysteinemia is a mild risk factor. Elevated levels (more than 150 percent of normal) of various coagulation factors, including factors VIII, XI, and IX and fibrinogen also appear to be mild risk factors for venous thrombosis. Venous thrombosis patients frequently have two or more genetic risk factors. See Schafer,351 Bertina,15 Rosendaal,16,78 and van Boven.279 Taken from Schafer with permission.351

Venous thrombosis and its complications are important and common medical problems, estimated to occur at a rate of 1.2 events/1000 population per year.19 Thus, for the population of the United States, there will be approximately 201,000 new cases of venous thromboembolism per year, of which 107,000 will be deep venous thrombosis and 94,000 will be pulmonary embolism.
In this chapter, we will discuss the pathogenesis and unique clinical features of each of the more common hereditary thrombophilias (see Table 127-1). Thereafter an approach to the diagnosis and subsequent treatment of these thrombophilic patients will be presented.
MAJOR HEREDITARY DEFECTS
HEREDITARY RESISTANCE TO ACTIVATED PROTEIN C
DEFINITION AND HISTORY
The term activated protein C (APC) resistance is defined as an abnormally reduced anticoagulant response of a subject’s plasma to APC on the basis of in vitro testing. A “normal” range for response to APC is established for the various coagulation or other related assay conditions used to assess response to APC.7,20,21 In 1989, an abnormally poor response to APC was described for several individual patients with venous thrombosis when it was shown that partially purified antibody fractions interfered with expression of APC activity.22,23 and 24 In 1991, familial APC resistance was first reported in one kindred and was ascribed to an APC-resistant factor VIII defect.25 Apparently this mechanism could not be confirmed, and in 1993, the description of three unrelated families presenting with venous thrombosis associated with APC resistance without any identifiable defect stimulated an intensive search for genetic and molecular mechanistic explanations.7 Three laboratories independently reported in May of 1994 that a single genetic defect was associated with APC resistance, involving replacement of G by A at nucleotide 1691 in exon 10 of the factor V gene which causes the amino acid replacement of Arg506 by Gln.9,10 and 11. Further analysis indicated genetic linkage between APC resistance and the factor V gene.26,27 In the published literature and in this chapter, this defect is variously termed Gln506-factor V, Q506-factor V, or factor V Leiden.
ETIOLOGY AND PATHOGENESIS
APC resistance can be caused by heterogeneous molecular defects, although replacement of Arg506 by Gln in factor V is responsible for APC resistance in the great majority of patients. APC resistance is a laboratory phenotype, whereas Gln506-factor V is a genotype, and the term APC resistance should not be used as a synonym for factor V Leiden. APC resistance caused by defects other than factor V Leiden is associated with increased risk of venous thrombosis28,29 or ischemic stroke.30,31
Theoretically, any genetic abnormality of a protein C pathway component that interferes with the expression of APC activity can cause APC resistance as could acquired abnormalities such as antibodies against protein C pathway components.32,33 Although the causes of many cases of acquired APC resistance are unknown, the majority (more than 90 percent) of hereditary APC resistant subjects have the same genetic abnormality, factor V at G1691A (Arg506Gln), which arose in a single Caucasian founder some 21,000 to 34,000 years ago.34 The molecular mechanism for APC resistance in such probands involves resistance of Gln506-factor Va to proteolytic inactivation by APC,9,35,36 with kinetic studies showing that the Gln506-variant is inactivated 10 times slower than normal Arg506-factor Va.36,37,38 and 39 Gln506-factor Va, whether activated by thrombin or factor Xa, is partially but not entirely resistant to APC, implying that inactivation of Gln506-factor Va by APC can occur in vivo, albeit at a reduced rate. Explanation for only a partial resistance to APC derives from the fact that cleavage of factor Va by APC at Arg306 also occurs, causing complete loss of factor Va activity, although this cleavage is slower than that at the Arg506 site.36,37,38,39 and 40 This finding helps explain why APC resistance due to Gln506-factor V is a rather mild risk factor for venous thrombosis and why a combination of genetic risk factors or a combination of a genetic risk factor plus acquired risk factors for venous thrombosis is found in a significant fraction of symptomatic patients (see below). A nother possibility to help explain the mild risk of venous thrombosis associated with Gln506-factor V is that factor Va may be inactivated in vivo by proteases other than APC that cleave at sites other than residue 506.
There are additional potential molecular defects that might contribute to thrombosis in hereditary APC resistance. In purified clotting factor reaction mixtures, factor V enhances inactivation of factor VIIIa by APC in the presence of protein S,41 and APC resistant subjects carrying Gln506-Factor V are reportedly defective in this APC cofactor activity.42,43,44 and 45 APC resistance caused by rare factor V mutations that replace Arg306 by Thr46 or Gly47 have been reported, although the relationship to relative risks of thrombosis have not been established.48 A factor V haplotype, designated R2, has also been associated with mild APC resistance.49 The molecular mechanisms and thrombotic risks associated with the R2 factor V haplotype which contains normal Arg506 remain to be defined, although it appears that the R2 haplotype is only a risk factor when present along with the Gln506-factor V allele.50
APC is a normal component of circulating blood that contributes to antithrombotic surveillance mechanisms and prevents thrombosis (see Chap 113).51 Normal subjects have a mean APC concentration of 2.3 ng/ml (38 pM) in the circulation,52 and the in vivo half-life of APC in normal adult human subjects as well as in freshly drawn whole blood is approximately 22 min.53,54 Thus, there is continuous activation of the protein C pathway in vivo. In normal subjects, there is an inverse relationship between levels of circulating APC and of thrombin.55 APC levels are increased when thrombin is acutely generated such as during DIC, ischemia, or surgical procedures.
Because circulating APC has such a long half-life, it provides systemic anticoagulation to down-regulate thrombin generation and to limit extension of hemostatic plugs. Hence, genetic or acquired defects that impair the response to APC are understandably prothrombotic. Elevated plasma levels of prothrombin fragment F1+2 and thrombin-antithrombin complexes are found in many subjects heterozygous or homozygous for Gln506-factor V,56,57,58 and 59 presumably reflecting the impairment of the expression of APC’s anticoagulant activity.
APC resistance has been associated with less intrapartum blood loss, suggesting an evolutionary advantage.60 However, pregnancy loss is increased in some women due to thrombosis in the placental vasculature (see below). Hemophilia A patients who also inherit Gln506-factor V have been reported to have less severe hemorrhagic symptoms.61
CLINICAL FEATURES
The factor V Leiden mutation is present in 3 to 12 percent of Caucasians and is rare in other ethnic groups.62,63 and 64 Deep and superficial venous thromboses are the most common manifestations of this disorder, whereas pulmonary embolism and thromboses in unusual locations appear to be relatively less frequent than in subjects with deficiencies in antithrombin, protein C, or protein S.65,66,67,68 and 69 In patients with venous insufficiency leading to leg ulcers, approximately 25 percent were found to have APC resistance70 or factor V Leiden.71 Cerebral, hepatic, and other thromboses have been reported in patients with factor V Leiden.72,73 and 74 About half the patients will have idiopathic (unprovoked) venous thromboembolism, with 20 percent occurring after surgery, and 30 percent in women who are pregnant or taking birth control pills.75 Pregnancy loss and other obstetric complications occur at an increased rate in women with factor V Leiden (see below).
The risk of thrombosis in subjects with factor V Leiden appears to be somewhat lower than in patients from families with deficiencies of antithrombin or protein C76,77; nonetheless, since factor V Leiden is so common, it accounts for the largest proportion of patients presenting with a first thromboembolic event (20 to 25 percent).78 The relative risk of venous thrombosis in patients heterozygous for factor V Leiden is increased by four to eightfold in studies from Europe and North America.20,67,79,80 and 81 The risk of idiopathic venous thromboembolism for men increases with age, from a relative risk of 1.23 at age 40 to 50 years to 5.97 for those aged 70 years and older.79 First-degree relatives of symptomatic carriers of the factor V mutation develop thromboses at a rate of 0.45 percent per year (0.25 percent per year in the 15- to 30-year age group, and 1.1 percent per year in those over 60).75,77 In several studies, recurrent thrombotic events are rather frequent, occurring at a rate of 5 to 10 percent per year following a first thrombosis.82,83 and 84 However, other investigators found no increased rates of recurrence when compared to thrombosis patients without factor V Leiden.85,86 and 87 Homozygous carriers of factor V Leiden have an odds ratio for venous thrombosis of 50 to 100, and it is estimated that approximately half of such individuals will experience a clinically significant episode during their lives.88 Alth ough thromboses in homozygotes are substantially more common than in heterozygotes, the disorder is far less severe than in subjects with homozygous deficiency of protein C, or protein S.56,88,89 Despite the increased thrombotic risk, the presence of factor V Leiden does not increase overall mortality.90,91,92 and 93
Coronary artery thrombosis has been notably associated with the factor V Leiden mutation in young women17 and men94 also displaying other vascular risk factors. The relative risk of myocardial infarction in carriers of V Leiden from the Netherlands is 1.4, which increased to three- to sixfold if other risk factors such as obesity, smoking, hypertension, or diabetes were present.95 Similar findings have been reported for young women from Washington State, with odds ratios of up to 32 for myocardial infarction in V Leiden carriers who were also smokers.17 However, other studies have failed to find a relationship between APC resistance and myocardial infarction or stroke in older individuals.80,96,97 and 98 Factor V Leiden seems to be relatively common in children who develop cerebral infarction or venous thrombosis.99,100 and 101
PREDISPOSING FACTORS FOR THROMBOSIS IN SUBJECTS WITH FACTOR V LEIDEN
Although isolated factor V Leiden is associated with a relatively mild hypercoagulable state, the risk of thrombosis is greatly magnified when other prothrombotic disorders are also present (Fig. 127-1). These additional risk factors may be hereditary (e.g., protein C deficiency or the prothrombin gene mutation), acquired (antiphospholipid antibodies, hyperhomocysteinemia), physical (inactivity or surgery), due to other diseases (malignancy or inflammation), or hormonal (oral contraceptives or pregnancy).78
Multiple hereditary thrombophilic defects (i.e., gene-gene interaction) are quite common, and are found in up to 15 percent of patients presenting with venous thromboembolism.102 Factor V Leiden has been reported in combination with protein C deficiency,9,103,104 and 105 protein S deficiency,106,107 and 108 antithrombin deficiency,109 the prothrombin gene mutation,110,111 and hyperhomocysteinemia.112,113 and 114 In families with combined defects, thromboses occurred more frequently and at an earlier age in the subjects with two separate defects.
Pregnancy and estrogen-containing oral contraceptives substantially enhance the risk of thrombosis in women with factor V Leiden. Of women who develop venous thromboembolism during pregnancy, 28 to 46 percent will carry the factor V mutation.115,116 and 117 The relative risk of developing thrombosis for heterozygotes during or after pregnancy is increased over threefold, with a higher probability of recurrence (relative risk of 3.86).117 Several studies have examined the risks of thromboembolism in women with factor V Leiden using third-generation oral contraceptives. There is a highly significant increase of 30- to 80-fold in the odds ratio for thrombosis, with an absolute increase in risk from 0.8 to 28.5 per10,000 women per year.118,119 Even higher risks are seen in women who are homozygous for the mutation.120 Data are not yet available on the risk of thrombosis in women with factor V Leiden (with or without a history of thrombosis) who receive hormone replacement therapy. A major side effect of selective estrogen receptor modulators such as tamoxifen or raloxifene may also exert an increased risk of venous thrombosis. Three cases of tamoxifen-associated venous thrombosis were associated with factor V Leiden.121
LABORATORY ASSAYS
Coagulation assays and DNA-based assays are available for the identification of patients with APC resistance. Plasma-based coagulation tests depend on the relative prolongation of the activated partial thromboplastin time (aPTT) or other coagulation screening tests caused by the addition of purified APC. Individuals with resistance to APC have less prolongation of the aPTT than normal. Although an aPTT assay was originally used, current assays employ factor V deficient plasma,35 which makes the test informative in most patients with lupus inhibitors, in pregnant patients, in patients with inflammatory states, and in patients on anticoagulants. The test is sensitive and specific when compared with the genetic test for factor V Leiden.122,123,124,125 and 126 Studies using the first-generation assay (see above) have suggested that an abnormally low APC ratio is associated with venous thrombosis, in both the presence and absence of the factor V Leiden mutation28,29 and with ischemic stroke.30,31 Thus, there is clinically relevant information in the classic aPTT-based APC resistance test that is not obtained using factor V deficient substrate plasma. Tissue-factor-based APC resistance assays can provide additional information about plasma components that differentially modulate the protein C pathway,127,128 and 129 such as “anticoagulant” high-density lipoprotein or as yet unidentified factors that are altered by oral contraceptive usage. The presence of platelets or platelet microparticles in plasma tested for APC resistance using aPTT assays,130,131 and 132 as well as autoantibodies against APC,33 can reduce the anticoagulant response to APC, indicating the need to carefully prepare plasma prior to testing.
Many DNA-based assays for the factor V Leiden polymorphism are available. Genomic DNA is isolated, amplified by polymerase chain reaction (PCR), subjected to restriction fragment length polymorphism analysis, and analyzed for G or A at nucleotide 1691.9 Plasma coagulation tests are often used for screening patients, followed by confirmation of positive results with the DNA assay. Only DNA tests clearly distinguish factor V Leiden heterozygosity from homozygosity. “Pseudo-homozygotes” heterozygous for factor V Ledien and for a dysfunctional factor V allele will have very low APC-resistance ratios in the plasma test but will be heterozygous by the DNA assay for factor V Leiden.133,134 and 135
PROTHROMBIN G20210A GENE POLYMORPHISM
DEFINITION AND HISTORY
In 1996, Poort and colleagues reported that a polymorphism in the 3′-untranslated region of the prothrombin gene, namely nt G20210A, was associated with increased risk of venous thrombosis and with elevated levels of plasma prothrombin.14 This polymorphism likely arose as a single mutation in a Caucasian founder,136 and the polymorphism is currently found in 1 to 5 percent of Caucasians.137
ETIOLOGY AND PATHOGENESIS
Replacement of G by A at nt 20210 in the 3′-untranslated region of the prothrombin gene does not alter transcription of the gene but may increase translation, thus resulting in elevated synthesis and secretion of prothrombin by the liver. The elevated level of plasma prothrombin likely contributes directly to increased thrombotic risk by causing increased thrombin generation.
CLINICAL FEATURES
The prothrombin gene mutation is found largely in Caucasian populations.136 In contrast to factor V Leiden, the frequency of the mutation seems to increase from northern Europe to southern Europe, i.e., only 1.7 percent of the population in northern Europe had the abnormality compared with 3 to 5 percent in the south of Europe and the Middle East.137,138 The prothrombin gene mutation is associated with venous thrombosis in all age groups.139 When sequential patients presenting with a first venous thromboembolism are analyzed, 4 to 8 percent of them will have the mutation, and the odds ratio for thrombosis in subjects with prothrombin 20210A is increased approximately 2- to 5.5-fold.14,102,140,141,142,143,144 and 145 In patients with recurrent thromboembolism or a family history of thrombosis, as many as 15 to 18 percent will have the defect compared with 1 to 3 percent of controls in various populations.14,146 As mentioned earlier, the prothrombin variant is associated with elevated plasma levels of prothrombin (e.g., a mean of 132 percent).14 Increased prothrombin activity or antigen is also associated with an increased risk of thrombosis even in the absence of the mutation.147
As in other forms of hereditary thrombophilia, the prothrombin gene mutation has been found in patients with thrombosis in unusual sites, particularly cerebral sinus vein thrombosis.148,149,150,151,152 and 153 For example, in a study of 40 patients with cerebral vein thrombosis, 20 percent had the gene defect (OR 10.2). Many of these thromboses were in young women taking oral contraceptives, which raises the likelihood of thrombosis even higher (i.e., an OR of 150).150
Individuals who are homozygous for the prothrombin gene mutation appear more likely than heterozygotes to develop thrombosis.154,155 and 156 The mutation also occurs in concert with other hereditary thrombophilic states (8 percent in one study).111,140,142,146,157 When the prothrombin variant was associated with factor V Leiden in young symptomatic patients, overall thrombosis rates were increased as well as spontaneous events and thromboses in unusual locations.111
The prothrombin gene mutation appears generally not to be overrepresented in unselected patients with cerebral vascular or coronary artery disease.146,158,159,160 and 161 However, certain selected groups of patients with arterial thrombosis have an increased likelihood of carrying the mutation.95,141,155,162,163 and 164 In young (younger than 50 years) patients with documented ischemic stroke but without other risk factors such as diabetes, hypertension, or hyperlipidemia, 15 percent had the prothrombin gene mutation (giving an odds ratio for ischemic stroke of 5.1).155 The mutation also appears to be associated with an increased risk of myocardial infarction, especially in those with other major risk factors for coronary heart disease such as smoking.95,162 Finally, a large proportion of a group of young women with acute unexplained spinal cord infarction were found to have the mutation.165 All were taking oral contraceptives and most were smokers.
LABORATORY DIAGNOSIS
Identification of the mutation in the 3′-untranslated region of the prothrombin gene requires DNA analysis following PCR amplification of the pertinent region.14 Although prothrombin levels are elevated, assay of prothrombin activity or prothrombin antigen is usually not sufficiently sensitive or specific to screen for the presence of the mutation or as a more effective predictor of thrombosis.146,147,157
HYPERHOMOCYSTEINEMIA
DEFINITION AND HISTORY
A plasma homocysteine level above the normal range defines hyperhomocysteinemia. Severe hyperhomocysteinemia, also identifiable as homocystinuria, is rare and is an autosomal recessive trait associated with severe defects in cystathionine b-synthase, 5,10-methylenetetrahydrofolate reductase (MTHFR), or possibly other enzymes that affect homocysteine metabolism.166,167 and 168 Such severe abnormalities are associated with neurologic abnormalities, premature cardiovascular disease, stroke, and vascular thrombosis. Mild to moderate hyperhomocysteinemia is an independent risk factor for arteriosclerosis and arterial thrombosis.13,167,169 A meta-analysis of 10 case-control studies concluded that mild hyperhomocysteinemia conveys a significant, though mild, increased risk of venous thrombosis.170
ETIOLOGY AND PATHOGENESIS
Homocysteine is an intermediate in the metabolism of the sulfur-containing amino acids, methionine and cysteine, and homocysteine participates in several metabolic pathways. Remethylation of homocysteine to generate methionine requires the vitamin B12-dependent enzyme, methionine synthase, and 5-methyltetrahydrofolate, which are part of a metabolic pathway that recycles tetrahydrofolate, and 5-methyltetrahydrofolate and involves the enzyme methylenetetrahydrofolate reductase. For the synthesis of cysteine from homocysteine, a transulfuration pathway first involves condensation of homocysteine with serine to generate cystathionine by the vitamin B6-dependent enzyme, cystathionine b-synthase; then deamination and cleavage of cystathionine to yield cysteine and a-ketobutyrate is accomplished by the vitamin B6-dependent enzyme, cystathioninase. The most common known genetic cause of mild hyperhomocysteinemia involves an MTHFR gene polymorphism, nt C677T, that causes a conservative replacement of Ala222 by Val which results in a variant enzyme with reduced specific activity and increased thermolability.113 Homozygosity for this so-called thermolabile form of MTHFR, i.e., homozygosity for TT at nt677, is associated with mild hyperhomocysteinemia.
The most common cause of rare, severe hyperhomocysteinemia is defective cystathionine b-synthase. Suboptimal levels of folate or vitamins B6 or B12 can also contribute to acquired mild to moderate hyperhomocysteinemia by providing inadequate cofactor levels to support the enzymes that regulate homocysteine metabolism. Conversely, administration of folate with vitamins B6 and B12 can reduce homocysteine levels.171,172 To date, no controlled studies of vitamin therapy to reduce homocysteine levels in venous thrombosis patients have been reported nor has it been proved that this vitamin strategy reduces arterial or venous thrombotic risk.
The exact mechanisms by which hyperhomocysteinemia causes increased risk of thrombosis have not been defined, although there is strong evidence that elevated homocysteine levels cause deleterious prothrombotic alterations in a number of normal vascular functions based on animal model studies, tissue culture experiments, and clinical research (see Chap. 114).173,174 and 175
CLINICAL FEATURES
Hyperhomocysteinemia is commonly associated with venous thromboembolism as well as arterial disease. From 10 to 25 percent of patients with primary or recurrent venous thrombosis have plasma homocysteine concentrations that are greater than the 95th percentile of the distribution in normal individuals (i.e., greater than 17 to 22 µmol/liter).167,169,176,177,178,179 and 180 Meta-analysis of multiple studies suggest that the odds ratio for venous thrombosis is 2.5 to 3.0 if homocysteine concentrations are elevated.170 Coagulation activation markers such as F1.2 or plasma levels of activated protein C are increased in patients with hyperhomocysteinemia and thrombosis, suggesting the presence of hypercoagulability.181,182
The association of hyperhomocysteinema and venous thrombosis is stronger among women (e.g., OR 7), and it also increases with age, rising to an odds ratio of 5.5 for individuals who are older than 50 years.176 In most but not all studies,183 the combination of hyperhomocysteinemia in concert with other hypercoagulable states substantially increases the risk of thromboembolism.112,114,169 For example, in the Physicians Health Study, the odds ratio for idiopathic venous thromboembolism in subjects with hyperhomocysteinemia was 3.4, for those with factor V Leiden it was 3.6, but for subjects with both disorders, the odds ratio was greatly increased to over 20.114 Hyperhomocysteinemia is also a strong predictor of recurrent thrombosis (OR 2–3), with reported recurrence rates of up to 10 percentper year for the first 2 years after cessation of oral anticoagulants.177,180
The thermolabile form of MTHFR has been associated with hyperhomocysteinemia, particularly during periods of folate deficiency.184,185 Although controversial, it appears that homozygosity alone for this enzyme defect (which occurs in 10 to 20 percent of normal individuals) is associated with either an absent or a mild increased risk of venous thromboembolism in the absence of associated thrombotic risk factors.102,113,186,187,188 and 189 However, subjects who are homozygous for the MTHFR variant who also have factor V Leiden or the prothrombin gene mutation may be at a mildly to moderately increased risk of thrombosis.102,113,186,188
LABORATORY DIAGNOSIS
Plasma homocysteine concentrations can be measured by HPLC or an immunoassay or by performing a methionine loading test. Both fasting levels and levels after methionine loading have been used to assess hyperhomocysteinemia.190,191,192 and 193 Although the methionine loading test may detect additional subjects with hyperhomocysteinemia, it is not clear that the predictive value for thrombosis is sufficiently increased to warrant the additional effort and cost of this procedure.169,177,193,194 Blood samples for homocysteine levels should be obtained in the fasting state, kept cold, and centrifuged immediately.190,192 Individual measurements reflect average homocysteine concentrations over time (e.g., 4 weeks) reasonably well.190 Serum homocysteine levels are higher than plasma levels, and male values are higher than female values.195 The thermolabile nt 677T variant of MTHFR and mutations in the cystathionine beta-synthase gene can be assessed using DNA-based molecular techniques.196
PROTEIN C DEFICIENCY
DEFINITION AND HISTORY
The first cases of familial heterozygous protein C deficiency (about 50 percent of normal plasma level) associated with venous thrombosis in young adulthood3 and of severe protein C deficiency (less than 1 percent protein C activity) associated with neonatal purpura fulminans197 were reported in 1981. Most typically, hereditary deficiency of protein C results from an autosomal trait in which affected individuals have approximately 50 percent of the normal level of functional plasma protein C. Over 150 different mutations in the protein C gene which are associated with thrombosis have now been reported.198 Heterozygous protein C deficiency conveys a mildly increased risk of venous thrombosis. Fewer than two dozen cases of severe protein C deficiency due to homozygosity or compound heterozygosity have been reported in neonates with purpura fulminans or massive thrombosis. Type I protein C deficiency is defined as a disorder with parallel reductions in both plasma antigen and anticoagulant activity levels, whereas type II deficiency, associated with circulating dysfunctional molecules, involves normal plasma levels of antigen but low levels of anticoagulant activity.
ETIOLOGY AND PATHOGENESIS
Protein C is synthesized in the liver and circulates in plasma as a serine protease zymogen; it is activated by limited proteolysis by thrombin bound to thrombomodulin, possibly with additional acceleration by an endothelial protein C receptor (see Chap. 113). APC is a potent anticoagulant enzyme that down-regulates the blood coagulation pathways by proteolytic and irreversible inactivation of factors Va and VIIIa. Thus, decreased levels of protein C zymogen may impair the inhibition of thrombin generation and contribute to hypercoagulability.
CLINICAL FEATURES
Protein C deficiency occurs in 0.2 to 0.4 percent of normal individuals199,200 and is found in approximately 4 to 5 percent of consecutive outpatients with objectively confirmed deep venous thrombosis.201 Deficiency of protein C is linked to thrombosis (OR 6.5–8),76,201 and many families with hereditary protein C deficiency and thromboembolism have been reported.3,202,203 and 204 The mean age of first thrombosis has been reported to be similar (approximately 45 years) in patients with factor V Leiden and protein C deficiency suggesting similar thrombotic tendencies in the two types of thrombophilia.205 When mortality rates are compared, individuals with protein C deficiency have a normal life-span.206
Variability in clinical expression is a hallmark of the disorder. Subjects identified by screening large numbers of normal individuals (e.g., blood donors) in most instances have neither a personal nor a family history of thromboembolism.199,200 The discrepancy in thrombosis rates between these surveys and studies of families who have striking thrombotic symptoms can be explained in part by the coinheritance of factor V Leiden or another thrombophilic state (Fig. 127-1).9,103,104 and 105,207,208 Polymorphisms in the promotor region of the protein C gene resulting in lower levels of protein C in some of the families could also be involved.209 Recurrent thrombosis in affected families with protein C deficiency is quite common and is unprovoked in about 60 percent of instances.210,211
Deep and superficial venous thrombosis is the most common clinical presentation of protein C deficiency.210,211,212 and 213 By the age of 45, up to 50 percent of heterozygous subjects in clinically affected families will have venous thromboembolism, and half of the episodes will be spontaneous.203 Protein C deficiency has been linked to unusual sites of venous thrombosis including the cerebral and mesenteric veins.210,214 Arterial thrombosis seems to be uncommon, although ischemic stroke and other arterial occlusive events have been reported.76,215
Homozygous protein C deficiency with protein C levels of less than 1 percent produces a fulminating thrombotic diathesis including the dramatic syndrome of neonatal purpura fulminans in affected infants.197,216,217,218 and 219 In a similar scenario, “warfarin skin necrosis,” large areas of thrombotic skin necrosis, appear over central areas of the body (breast, abdomen, genitalia) in subjects with heterozygous protein C deficiency given warfarin.220 In this syndrome in protein C deficient patients, the vitamin K antagonist induces a fall in protein C activity from approximately 50 percent to very low levels because of the short half-life of protein C in vivo (4 to 8 h).221 Because the half-lives of prothrombin, factor IX, and factor X are much longer, a transient hypercoagulable state may arise at the outset of vitamin K-antagonist therapy. Heparin or low-molecular-weight heparin should be used when initiating warfarin treatment in subjects known to be protein C deficient.202
LABORATORY DIAGNOSIS
Most laboratories screen for protein C deficiency with a protein C activity assay that employs a highly specific snake venom protease to activate protein C.222,223 Protein C activity is best assessed with an assay that employs a coagulation rather than a chromogenic end point to identify the greatest number of patients with protein C deficiency.224 Immunoassays are used to distinguish type I defects (reduced antigen and activity) from type II disorders (normal antigen, reduced activity).225 Normal ranges for protein C increase with age (4 percent per decade) so that results need to be interpreted against these age-specific norms.222 Protein C gene promotor polymorphisms also influence plasma concentrations of the protein which can vary from 94 to 106 percent,209,226 and liver disease or oral contraceptives can lower or raise protein C levels respectively.227 Consequently, protein C levels of less than 55 percent (in the absence of oral anticoagulants or overt liver disease) suggest protein C deficiency, but levels from 55 to 70 percent must be considered borderline, and repeated testing or family studies should be undertaken.224 The use of DNA-based assays to identify patients with hereditary protein C deficiency is not practical because more than 150 different mutations have been described.198
The diagnosis of hereditary protein C deficiency in patients who are receiving warfarin is particularly difficult. Protein C antigen levels can be compared with antigen levels for other vitamin K-dependent clotting factors such as factor VII or X, but only if careful control ranges are established for the ratios of protein C to two other vitamin K-dependent factors are established.3,202 In most situations, it is necessary to wait until at least 2 weeks after the end of anticoagulant therapy for a reliable diagnosis. Warfarin should not be restarted before laboratory results have been returned to reduce the possibility of warfarin-induced skin necrosis in patients who are later found to have protein C deficiency.
PROTEIN S DEFICIENCY
DEFINITION AND HISTORY
Familial heterozygous protein S deficiency associated with venous thrombosis was first reported in 1984.4,5 and 6 Since then, many other families with the disorder have been identified.76,228,229,230,231 and 232 Many different mutations in the protein S gene (over 100) associated with thrombosis have been identified.233 Severe deficiency (less than 1 percent of normal protein S levels) due to homozygous or compound heterozygous defects has been reported in only a few infants who presented with neonatal purpura fulminans.234,235 and 236 Protein S enhances the anticoagulant activity of APC, and hence currently available functional assays of protein S measure APC-cofactor activity using protein S–depleted plasma as substrate. Type I protein S deficiency is defined as parallel reductions in both antigen and anticoagulant activity levels in plasma whereas type II deficiency, associated with circulating dysfunctional molecules, involves normal plasma levels of antigen but low levels of anticoagulant activity. Protein S reversibly associates with the plasma complement factor, C4b-binding protein (C4BP), previously known as proline-rich lipoprotein. In normal plasma, approximately 60 percent of protein S is bound to C4BP, and 40 percent is free; importantly, only the free form of protein S functions as a cofactor for APC. This gives rise to another type of protein S deficiency, designated type III deficiency, in which free protein S is low while total protein S antigen is usually in the low normal range.
ETIOLOGY AND PATHOGENESIS
Protein S is principally synthesized in the liver, but other organs may be important sites for its synthesis, including the endothelium, kidney, testes, and brain (see Chap. 113). Because protein S is a cofactor for APC (see Chap. 113), decreased levels of free protein S may impair the down-regulation of thrombin generation and contribute to hypercoagulability. Protein S also exhibits anticoagulant activity that is independent of APC by directly binding to and inhibiting factors Va, VIIIa, and Xa,237,238,239,240,241 and 242 suggesting that deficiency of protein S could also contribute to hypercoagulability by failing to impair factors Va or Xa in the absence of APC. At present, only the APC-cofactor activity of protein S is routinely assayed because there is no generally available standardized assay for the anticoagulant activity of protein S that is independent of APC.
CLINICAL FEATURES
In several studies, approximately 3 percent of unselected outpatients presenting with venous thromboembolism have low levels of protein S201,243,244; higher prevalences are reported for patients under 50 years of age and for patients with a personal or family history of venous thrombosis. The odds ratio for thrombosis in patients with free protein S deficiency has been variably reported to be 1.6,201 2.4,243 8.5,76 and 11.5230 (see below). After an initial venous thrombosis, recurrence rates in protein S–deficient patients average 3.5 percent per year.212,213,245
Deep venous thrombosis and pulmonary embolism are the most common forms of thrombosis associated with protein S deficiency, although superficial vein thrombophlebitis and thrombosis in unusual sites also occur.210,211 and 212 As in other forms of thrombophilia, about 50 percent of thromboses are unprovoked.211 Arterial thrombosis has been reported in a significant number of protein S–deficient patients, particularly in those who smoke or have other thrombotic risk factors.18,106,246,247 Neonatal purpura fulminans has been reported in rare infants with homozygous or compound heterozygous protein S deficiency and very low levels of protein S.234,235 and 236 Warfarin-induced skin necrosis has also been reported in association with protein S deficiency.248
Acquired forms of protein S deficiency are rather common. Oral contraceptive usage decreases plasma protein S levels. Reduced levels of free protein S are regularly found in pregnancy (e.g., as low as 20 to 30 percent of normal),249,250 in patients who are taking oral anticoagulants, and in disseminated intravascular coagulation, liver disease, nephrotic syndrome, inflammatory conditions, and acute thromboembolism.251,252,253 and 254 Protein S deficiency can also occur in concert with the lupus anticoagulant255,256 and as a result of autoantibodies to protein S following varicella or other infections in children.257,258,259,260 and 261
The likelihood of thrombosis varies widely in patients with protein S deficiency. In general, population-based case control studies yield low odds ratios for thrombosis,201 whereas family studies show a high rate of venous thromboembolism in protein S–deficient relatives compared with nonaffected family members.230 Some of the patients identified in the case control studies may have had an acquired deficiency of protein S which was temporary.224 Even more important, several of the families with protein S deficiency have been found to have a second thrombophilic defect, most commonly either factor V Leiden,106,108 or the prothrombin nt 20210A gene mutation.157 Other risk factors, particularly smoking and obesity, also increase the risk of thrombosis in protein S–deficient family members.18,230,231
LABORATORY EVALUATION
Laboratory assays of plasma protein S must be chosen and interpreted with care because the protein circulates both free and bound to C4BP. Moreover, normal ranges differ for males compared with females and depend on age. Free protein S antigen or APC-cofactor anticoagulant activity are better than total protein S antigen in screening for hereditary protein S deficiency.201,262 Free protein S antigen can be assayed using monoclonal antibodies specific for free protein S.263,264 Protein S activity assays may be affected by coexisting APC resistance, although the second-generation assays in which factor V–deficient plasma is used as substrate have improved specificity.265,266 and 267 Assessment of total and free protein S plus protein S activity should allow the classification of patients with protein S defects into types I, II, or III. Type I and type III deficiencies may actually be phenotypic variants of the same disease, because within families, different individuals carrying the same DNA mutation in the protein S gene can present with laboratory findings indicating either type I or type III deficiency.262 Type II deficiency, i.e., normal free protein S antigen with reduced protein S activity, is quite uncommon224 so that screening patients with free protein S antigen levels is clinically reasonable. In normal patients, there is an excellent correlation between free protein S antigen and anticoagulant activity. The lower limit of the normal range for free protein S is lower in females than in males (55 percent versus 65 percent)268; protein S is remarkably sensitive to hormonal status in females.
A coagulation assay for protein S anticoagulant activity independent of APC has been described in which the APTT is determined in the absence and presence of anti–protein S neutralizing polyclonal antibodies added to the test plasma. The APTT is shorter in the presence of antibodies, and the ratio of clotting times is indicative of protein S anticoagulant activity.269 At present, the clinical utility of this interesting assay has not been demonstrated, and the assay is not accessible for routine laboratories.
The high frequency of acquired protein S deficiency makes identification of hereditary defects more difficult. Common acquired conditions leading to low protein S levels should be excluded and tests repeated before making a diagnosis of hereditary thrombophilia. Family studies may also be useful. Oral anticoagulant therapy markedly reduces protein S antigen and activity levels. Assays are not often useful during pregnancy, because the low concentrations of protein S normally seen at that time cause diagnostic confusion.249 Diagnosis of hereditary protein S deficiency using DNA techniques is not favored unless the defect has previously been established in the family because there are numerous different mutations in the protein S gene causing protein S deficiency.
ANTITHROMBIN DEFICIENCY
DEFINITION AND HISTORY
Antithrombin, also known as antithrombin III, is a plasma protease inhibitor that neutralizes thrombin by irreversibly forming a 1:1 complex. The rate of inhibition of thrombin or other serine trypsinlike proteases by antithrombin is catalyzed by heparin. The first family with hereditary antithrombin deficiency and thrombosis was reported by Egeberg in 1965.1 Since then, many more families have been described.270,271 and 272 A database of over 250 mutations in the antithrombin gene is available273 and can be accessed via the internet (http://www.med.ic.ac.uk/dd/ddhc). Type I antithrombin deficiency is defined by low levels of antigen and activity in the absence or presence of heparin. Type II deficiency involves the presence of dysfunctional molecules in the plasma and is defined by normal levels of antigen with defects that affect either the inhibitor’s active center, which complexes with the target enzyme’s active site, or the inhibitor’s heparin binding site which mediates heparin-dependent acceleration of antithrombin’s action. Severe deficiency of antithrombin (less than 5 percent) is very rare, involves defects in heparin-dependent enhancement of antithrombin, and is associated with severe venous and arterial thrombosis.274,275,276 and 277 Type I antithrombin deficiency is found in 0.023 percent of normal individuals in Scotland, whereas type II defects, mostly in asymptomatic individuals and families, is much more common and found in 0.16 percent of people screened.278
ETIOLOGY AND PATHOGENESIS
Antithrombin is a major protease inhibitor that neutralizes factors Xa, IXa, XIa, and thrombin in reactions accelerated in the presence of heparin or by heparan sulfate on endothelial surfaces (see Chap. 113). Therefore, defects in antithrombin compromise the normal inhibition of the coagulation pathways and cause a hypercoagulable state. Molecular antithrombin defects can involve either the reactive center that combines with the active site of the coagulation proteases or the heparin binding region that mediates heparin-dependent acceleration of antithrombin-protease reactions (see Chap. 113).
CLINICAL FEATURES
Antithrombin deficiency is found in approximately 1 percent of consecutive outpatients under 70 years old with a first objectively documented venous thrombosis (see references76,77 and 78), and the odds ratio for thrombosis in patients with antithrombin deficiency is approximately 10 to 20 and is notably greater than in subjects with factor V Leiden.76,78,201,279 Recurrence rates have been reported to be quite high in the first year after a thrombosis (12 to 17 percent) in selected patients with type I antithrombin deficiency,213,245 but lower rates have been reported in other studies (about 4 percent per year).211,280 There is no evidence that there are differences in clinical severity between patients with heterozygous type I defects and those with type II mutations involving the thrombin binding site. Mortality rates are not increased in these patients.281,282 Patients with type II mutations of the heparin-binding site have few if any thrombotic episodes, although homozygous mutations affecting heparin binding are associated with thromboembolism.280
Venous thrombosis of the lower extremities, which occurs at an early age and peaks in the second decade of life, is the most common symptom in antithrombin deficiency.280 Superficial venous thrombosis appears to be somewhat less common than in protein C or protein S deficiency, or in APC resistance.76,210,211 Thrombosis in unusual sites such as the mesenteric or cerebral veins has been reported.210,211 Arterial thrombosis occurs infrequently (about 1 percent of affected patients).283 As previously indicated for patients with other forms of hereditary thrombophilia, gene-gene and gene-environment interactions markedly increase the risk of thrombosis in subjects with antithrombin deficiency by 5-fold and 20-fold respectively.279 Patients with severe antithrombin deficiency, i.e., activity levels less than 5 percent, are exceedingly rare, most likely because the profound deficiency state causes fetal loss in utero. A few infants with homozygous defects involving the heparin-binding region of the molecule have survived, but most have suffered severe venous and arterial thrombosis.274,275,276 and 277 No patients homozygous for reactive center defects have been identified, leading to the speculation that complete deficiency of antithrombin is incompatible with life.
Resistance to the anticoagulant effects of heparin has been observed in some patients with antithrombin deficiency. However, heparin resistance is quite common in general patients with thrombosis. Up to 40 percent of patients without antithrombin deficiency will require more than 40,000 units of heparin daily to prolong the APTT into the therapeutic range.284 Both acute thrombosis and several days of heparin therapy can decrease antithrombin levels, occasionally to as low as 50 percent of normal, which may lead to an erroneous diagnosis of hereditary antithrombin deficiency.285,286 Acquired conditions leading to lowered levels of antithrombin are common and include liver disease, DIC, nephrotic syndrome, chemotherapy with asparaginase, and preeclampsia.287,288,289,290 and 291
LABORATORY DIAGNOSIS
Antithrombin deficiency screening assays should first be performed in the presence of heparin because defects may involve either the reactive center of the inhibitor or the heparin-binding site. If initial results are abnormal, then assays that measure the ability of the inhibitor to neutralize thrombin in the absence of heparin (progressive antithrombin activity) should be done to characterize the abnormality. Antithrombin activity assays that utilize a chromogenic substrate are widely available.292 Most laboratories now use factor Xa or bovine thrombin in their antithrombin assays to avoid the inhibitory effects of heparin cofactor II on human thrombin.293,294 The normal range for antithrombin levels in normal plasma is quite narrow (i.e., 84 to 116 percent).293 Antithrombin antigen measurements are used to help distinguish type I from type II defects. Crossed immunoelectrophoresis using an antithrombin antibody in the presence and absence of heparin can help identify defects in the heparin-binding portion of the molecule.295
In general, patients with type I deficiency and many of those with type II disorders involving the thrombin binding site will have antithrombin activity levels of 40 to 60 percent. Levels of 60 to 84 percent can be due to other type II defects but frequently are a result of acquired antithrombin deficiency such as occurs with mild liver disease, acute thrombosis, or heparin therapy. If these confounding conditions are present, measurement of levels should be repeated and family studies performed if possible.
HIGH LEVELS OF FACTOR VIII AND OTHER COAGULATION FACTORS
DEFINITION AND HISTORY
Based on analysis of the frequency of factor VIII levels that exceed 150 percent of normal values, an elevated factor VIII level has been defined as a significant independent risk factor for venous thrombosis.296,297 Both factor VIII activity and antigen levels are increased.297 The increased risk was similar to that of heterozygosities for factor V Leiden or prothrombin G20210A. Preliminary reports of other epidemiologic studies indicate that elevated levels (higher than 150 percent of normal) of factors XI, IX, X, and V are also risk factors for venous thrombosis.
ETIOLOGY AND PATHOGENESIS
Although factor VIII is an acute phase reactant and elevations can be caused by inflammation, it appears that factor VIII elevations in venous thrombosis patients are not commonly caused by systemic inflammation.297,298 Therefore, elevated factor VIII levels are likely to be directly pathogenic by increasing coagulability of blood via the blood coagulation pathways. Studies of factor VIII levels in different families indicate a significant genetic influence in addition to the known influences from levels of von Willebrand factor, blood group antigens, and the presence of inflammation. Elevations of factor VIII and other coagulation factors of the intrinsic coagulation pathway, e.g., factors XI, IX, X, or V, may contribute to hypercoagulability by increasing thrombin generation.
CLINICAL FEATURES
The clinical presentation of patients with elevated factor VIII levels is not known to differ from that of patients with the other genetic risk factors described in this chapter.
LABORATORY DIAGNOSIS
Factor VIII procoagulant activity is measured with routine coagulation assays commonly used to screen for hemophilia (see Chap. 112 and Chap. 123). In venous thrombosis patients, factor VIII antigen levels correlate with activity measurements.297
HEREDITARY THROMBOTIC DYSFIBRINOGENEMIA
DEFINITION AND HISTORY
Dysfibrinogenemia is defined as a qualitative defect in the molecule due to a mutation in the gene for one of fibrinogen’s polypeptide chains. The hereditary dysfibrinogenemias represent a heterogeneous group of abnormalities that can be asymptomatic or cause either thrombosis or bleeding. Initial reports of dysfibrinogenemias associated with thrombophilia appeared in the 1960s from several laboratories. For a detailed treatment of dysfibrinogenemia, see Chap 124.
ETIOLOGY AND PATHOGENESIS
For normal hemostasis, fibrin is formed after release of fibrinopeptides from fibrinogen due to proteolysis by thrombin and subsequent polymerization of fibrin monomers. Fibrin is then stabilized by covalent cross-links introduced by factor XIIIa. Plasmin-dependent proteolysis of fibrin either to limit formation or growth of a thrombus or to clear fibrin in a timely and normal fashion during healing is essential. Defects in fibrinogen that cause abnormal fibrinolysis cause thrombosis, either because fibrin is not cleared in a normal fashion or because the growth of a normal hemostatic plug is not limited. Specific defects causing hypofibrinolysis can involve alterations of plasmin cleavage sites in fibrin or of sites that promote assembly of components of the fibrinolytic system, e.g., binding sites for plasminogen or plasminogen activators (see Chap. 124).
CLINICAL FEATURES
Patients with hereditary thrombotic dysfibrinogenemias usually present with venous thrombosis at a young age (e.g., 27 to 32 years old).299 An occasional patient will have both thrombosis and bleeding (usually postpartum hemorrhage).299 An increased rate of spontaneous abortion and stillbirth is also observed.299 Approximately 20 percent of reported cases of hereditary dysfibrinogenemia have been associated with thrombosis, whereas about 30 percent manifested bleeding, and the remainder were clinically silent.299 As summarized in detail in Chap. 124, several dozens of reports of abnormal fibrinogens associated with thrombosis have appeared.300 When a large number of patients presenting with thromboembolism were screened, the prevalence rate of dysfibrinogenemia was found to be 0.8 percent.299
LABORATORY DIAGNOSIS
Prolongation of a dilute thrombin time and/or reptilase time due to delayed fibrin polymerization is common with dysfibrinogenemia, as is a disparity between measurement of immunoreactive and clottable fibrinogen. More sophisticated testing often demonstrates abnormal fibrinogen structure or resistance of the fibrin to fibrinolysis. Unfortunately there are no assays readily available that measure the key properties of fibrinogen that are likely to cause thrombosis in patients with dysfibrinogenemia, and therefore this defect may be underdiagnosed.
Therapy relies on anticoagulants. In some instances, the administration of cryoprecipitate as a source of normal fibrinogen should be considered for surgical procedures, both to raise low concentrations of fibrinogen into a hemostatic range and possibly to reduce the risk of thrombosis by dilution of the abnormal prothrombotic fibrinogen.
OTHER POTENTIAL THROMBOPHILIC DISORDERS
Hereditary defects in the fibrinolytic system (see Chap. 116) and in thrombomodulin are potential thrombophilic risk factors. Japanese families with several hereditary dysplasminogenemias301,302 and 303 and hypoplasminogenemias304 have been identified, but these abnormalities were not associated with thrombosis in subjects other than the propositus. Heterozygous plasminogen deficiency is found more often in Asian than in Caucasian populations. One kindred with elevated levels of plasminogen activator inhibitor 1 (PAI-1)and thrombosis was subsequently shown to have protein S deficiency.231 As yet, an association between defects in the fibrinolytic system and thrombosis has not been firmly established.305 Several mutations in the thrombomodulin gene have been discovered in families with thrombosis.306,307 and 308 The genetic defects are scattered throughout the thrombomodulin gene and are associated with variable levels of soluble thrombomodulin in the plasma.307 In aggregate, thrombomodulin defects appear to be relatively common, being found in approximately 5 percent of a group of 200 patients with thromboembolic disease.307 It is not yet clear whether one or more of these mutations constitute a major risk factor, whether they may contribute to the risk of thrombosis in patients with other forms of thrombophilia such as protein S deficiency, or whether they are neutral polymorphisms. At this junc ture, routine screening of thrombosis patients for fibrinolytic or thrombomodulin defects is probably not indicated.
DIAGNOSIS OF THROMBOPHILIA
Plasma or molecular assays are now widely available for each of the common hereditary hypercoagulable states (Table 127-2). Up to 50 percent of patients presenting with a first deep vein thrombosis (DVT) will be found to have an abnormal laboratory test suggesting a thrombophilic defect. Those with recurrent venous thromboembolism or a strong family history of thrombosis are even more likely to have evidence of thrombophilia. Multiple disorders in the same patient are common; e.g., in one series of patients, 16 percent had more than one type of thrombophilia.102

TABLE 127-2 LABORATORY EVALUATION FOR HEREDITARY THROMBOPHILIA

Comprehensive testing for patients with venous thromboembolism should include: an APC resistance test (followed by a factor V Leiden mutation if needed), prothrombin gene mutation analysis, plasma homocysteine concentration, protein C activity (by a clotting assay), protein S activity assay (or free protein S antigen), antithrombin activity, factor VIII activity assay, and fibrinogen concentration (clottable) with a dilute thrombin time (+/– reptilase time) (see Table 127-2). Additional tests for antiphospholipid antibodies should also be considered in patients suspected of having acquired thrombophilia (see Chap. 128). The most appropriate tests for patients with arterial thrombosis are less clear. However, plasma homocysteine, antiphospholipid antibody studies, lipoprotein(a) concentration, and colony assays to search for covert myeloproliferative disorders should be considered. Factor V Leiden, the prothrombin gene mutation, protein S, and other tests for “venous” thrombophilia may prove useful in some patients with premature coronary heart disease or stroke, particularly if other risk factors are present such as smoking, hypertension, diabetes, or obesity. If test results for the more common disorders are normal but the likelihood of a familial hypercoagulable state is high, tests for other causes of thrombosis might be helpful. Experimental tests to consider are assays for elevated levels of factors XI, V, and IX, the 4G4G promoter polymorphism in the PAI-1 gene, a plasminogen activity assay, or perhaps molecular assays for defects in thrombomodulin.
A laboratory evaluation for thrombophilia should be considered if the results of testing could make a difference in the clinical care of the patient or family members. Examples of a potential clinical impact include:

Changes in the duration or intensity of oral anticoagulant therapy

Administration of specific therapy (e.g., antithrombin concentrates, vitamins for homocysteinemia)

More intense prophylaxis for high-risk situations (e.g., surgery, acute illness, immobility)

Better accuracy in estimates of the future risk of thrombosis in clinical settings (e.g., surgery or pregnancy)

Counseling of women as to the risks of oral contraceptives, pregnancy, or hormone replacement therapy

Study of family members at risk of thrombosis
Routine testing of all female family members of a patient found to have factor V Leiden prior to starting oral contraceptives is probably not indicated.75,309 However, if the family has a strong family history of venous thromboembolism in women who were pregnant or taking birth control pills or if the patient has had an episode of venous thrombosis, then test results could help in deciding whether to recommend oral contraceptives or alternative methods of birth control.309
Children with venous or arterial thrombosis (particularly stroke) are also likely to have an underlying thrombophilic disorder. Of these defects, factor V Leiden is the most common, but other disorders including the prothrombin 20210A gene mutation, protein C deficiency, elevations in lipoprotein(a), and antiphospholipid antibodies have been reported.99,100,310,311,312 and 313
Laboratory testing is best performed several weeks after completion of a course of oral anticoagulants in patients with thrombosis, to avoid confounding effects of acute thrombosis or heparin or warfarin therapy on the assay results. However, stopping anticoagulants in some patients with a high risk of recurrent thromboembolism may not be advisable. With the exception of assays for protein C and protein S, all other thrombophilic factors can be assayed in patients taking oral anticoagulants. Options for assessment of protein C or protein S levels in patients requiring warfarin include comparing their relative antigen levels with other “benchmark” vitamin K-dependent clotting factors3,202 or obtaining assays on family members. Alternatively, heparin or low-molecular-weight (LMW) heparin can be substituted for warfarin for a period of time (approximately 2 weeks) prior to drawing blood for analysis.
THERAPY OF THROMBOPHILIA
Thrombophilia patients who develop a DVT or PE are initially given standard venous thromboembolism treatment with heparin or LMW heparin for acute therapy and warfarin for longer-term protection. Warfarin has been shown to effectively prevent recurrent thromboembolism at an INR range of 2 to 3.314,315 Higher intensities of warfarin are unnecessary and will increase the risk of bleeding. The absolute risk of major hemorrhage with oral anticoagulants in patients with venous thromboembolism averages 1 to 3 percent per year, with a fatality rate of 0.2 to 0.4 percent per year.314,316
The optimal duration of anticoagulant treatment in patients with thrombosis and a history of thombophilia is an important clinical question.314,317 Warfarin therapy is usually given for 6 months315 following a thrombotic event (see Chap. 132). However, longer treatment may be indicated in patients with hereditary thrombophilia if the risk of additional thromboemboli substantially outweighs the risk of bleeding due to oral anticoagulants. The lack of reliable data on the absolute risks of recurrent thrombosis in patients with one or more thrombophilic states makes clinical decision making more difficult.314,317 However, several factors make recurrent thrombosis more likely, and, if present, longer-term treatment is more appropriate:

A spontaneous rather than a provoked thrombosis or pulmonary embolism316

A high odds ratio for thrombosis; e.g., antithrombin deficiency (OR = 8) versus factor V Leiden (OR = 2.5)

A strong family history of thrombosis (suggesting the presence of multiple hereditary defects)

A history of recurrent thromboses

Multiple inherited or acquired risk factors (e.g., factor V Leiden + prothrombin G20210A gene mutation; or factor V Leiden + antiphospholipid antibodies)

Permanent rather than temporary major risk factors

Unusual or life-threatening thromboses (e.g., cerebral vein or mesenteric thrombosis; ileofemoral deep venous thrombosis with multiple pulmonary emboli).
Decisions as to long-term anticoagulant therapy are best tailored to individual patients. A thorough assessment should include: (1) an estimate of the future risk of thrombosis; (2) an estimate of the future risk of major or fatal hemorrhage; and (3) patient preferences (e.g., impact of the decision on occupational or social situations).
Prophylactic oral anticoagulation therapy is usually not warranted in subjects who have not yet suffered a thrombotic event but who are discovered to have hereditary thrombophilia because of family testing or some other reason. In this instance, the risks of hemorrhage due to warfarin (about 1 to 3 percent per year) clearly outweigh the risk of thrombosis (e.g., 0.4 percent per year in asymptomatic individuals with APC resistance). In contrast, most authorities would recommend long-term antithrombotic treatment for patients who have suffered recurrent thromboses and who have more than one hereditary or acquired hypercoagulable state. More clinical trial data are needed before persuasive clinical guidelines can be recommended for patients with narrower risk/benefit ratios; e.g., a young patient with a first spontaneous but extensive DVT and a single prothrombotic disorder such as the factor V Leiden mutation.
If oral anticoagulant therapy is not used, an alternative approach includes intensive antithrombotic prophylaxis (e.g., LMW heparin) for events with a high risk of thrombosis such as surgery, infectious (e.g., pneumonia) or inflammatory diseases (e.g., inflammatory bowel disease), or prolonged periods of inactivity. Effective prophylaxis should reduce the risk of thromboembolism by about half, since approximately 50 percent of thromboses in patients with hereditary hypercoagulable states can be attributed to a known provoking factor. Other recommendations include patient education as to the signs and symptoms of acute DVT or PE, facilitation of diagnostic testing should symptoms occur, and continued follow-up in case new laboratory tests or clinical recommendations appear in the future.
Specific therapies are available for some thrombophilic disorders. Antithrombin concentrates are now widely available and can be administered for surgery, major trauma, and at the time of delivery in patients with antithrombin deficiency.318,319 and 320 Protein C and activated protein C concentrates are under development and, when available, may be useful in infants or children with homozygous protein C deficiency, or in heterozygous subjects during surgery or other major stresses.321,322 and 323 Cryoprecipitate is a source of normal fibrinogen, which can be useful for replacement of normal fibrinogen in patients with hereditary dysfibrinogenemia. Finally, although not yet proved to prevent thrombosis in patients with hyperhomocysteinemia, B vitamins (folic acid, pyridoxine, B12) effectively lower homocysteine concentrations into the normal range.171,172
THROMBOPHILIA AND PREGNANCY, ORAL CONTRACEPTIVES, AND HORMONE REPLACEMENT THERAPY
MANAGEMENT OF PREGNANCY IN THROMBOPHILIC WOMEN
Pregnancy substantially increases the risk of thrombosis in women with thrombophilia.324 To date, treatment guidelines supported by clinical trial data for the treatment of these women are not available. Screening all women for a thrombophilic state prior to pregnancy does not seem warranted based on an extremely high cost:benefit ratio.325,326 Similarly, routine heparin prophylaxis for previously asymptomatic individuals known to carry the V Leiden gene mutation or other mild thrombophilic defects is not indicated. Prophylactic heparin (or LMW heparin if proved safe and effective in pregnancy) should be considered for women with a history of venous thromboembolism, particularly if the prior thrombosis was related to pregnancy or oral contraceptives.324,327,328 The potential antithrombotic benefit should offset the risks of heparin-induced osteopenia, bleeding, or heparin-induced thrombocytopenia. Venous thromboembolism that occurs during pregnancy requires therapeutic doses of heparin for the remainder of the pregnancy, followed by postpartum anticoagulants for at least 4 to 6 weeks.324 Antithrombin concentrates (along with low-dose heparin) should be considered for women with hereditary antithrombin deficiency during the peripartum period or during complications of pregnancy.
PREGNANCY LOSS DUE TO THROMBOPHILIA
Thrombophilia is a cause of fetal loss and other complications of pregnancy, most likely due to thrombosis of the placental vasculature.326,329 Fetal loss is often manifested by stillbirth (second or third trimester) rather than first trimester miscarriage.329 In addition, severe preeclampsia, fetal growth retardation, and placental infarction have all been linked to maternal or fetal hypercoagulable states.330,331,332,333 and 334 Most of the hereditary thrombophilic states have been implicated. Factor V Leiden has been linked to fetal loss (OR 2–3),329,335,336,337 and 338 as have protein C deficiency (OR 2.3), protein S deficiency (OR 3.3), and antithrombin deficiency (OR 5.2).329 If combined defects are present, the odds ratio for fetal loss increases to 14.3.329 Hyperhomocysteinemia has been associated with placental abruption, placental infarction, and stillbirth; moreover, homozygosity for the thermolabile MTHFR defect has also been linked to pregnancy complications.330,331,334,339 Although quantitative data are not available, pregnancy loss appears to be increased in women with hereditary thrombotic dysfibrinogenemia.299
When adverse outcomes of pregnancy were combined, including stillbirth, preeclampsia, abruptio placentae, and fetal growth retardation, 52 percent of affected women were found to have thrombophilia compared to a rate of 17 percent in women with normal pregnancies.330 Diagnostic studies for thrombophilia should be considered for women with recurrent midtrimester pregnancy loss or other adverse pregnancy outcomes, particularly if future studies suggest that antithrombotic treatment (e.g., low-dose heparin, LMW heparin, or aspirin) is effective.340
ESTROGENS: ORAL CONTRACEPTIVES AND HORMONE REPLACEMENT THERAPY
Oral contraceptives increase the risk of thrombosis in women with hereditary thrombophilia.118,119,341,342 and 343 For example, the odds ratio for thrombosis in women with factor V Leiden who use third-generation oral contraceptives is increased 30- to 50-fold.118,119 In absolute numbers this represents an increase in risk from 1/12,500 women per year without V Leiden to 1/400 women per year in women with factor V Leiden.119 The thrombotic risk associated with birth control pills is greater in women who are homozygous for factor V Leiden.120 Screening for the factor V mutation prior to the administration of oral contraceptives is probably not cost-effective. By one estimate, it would be necessary to screen 2.25 million women to detect 90,000 women with V Leiden, in order to prevent one death from venous thromboembolism by withholding the oral contraceptives.344 However, if a woman with factor V Leiden has a history of thrombosis or is homozygous for the mutation, then avoidance of oral contraceptives would certainly be prudent.119,120,345 Oral contraceptives probably should not be recommended for women known to be deficient in antithrombin, protein C, and possibly protein S.341
Whether to recommend hormone replacement therapy in women with hereditary thrombophilia is a particularly difficult question. The relative risk of venous thrombosis with replacement estrogens is significantly increased by a factor of 2 to 4 when large groups of women are studied, but the absolute risk of thrombosis is quite low; i.e., 1 excess thrombosis per 5000 women per year.346,347,348,349 and 350 However, studies are not yet available to estimate any potential increase in the relative or absolute risk of thrombosis in women who use hormone replacement therapy and carry the factor V Leiden mutation or other genetic thrombophilic risk factors.
Use of selective estrogen receptor modulators such as tamoxifen or raloxifene increases the risk of venous thrombosis. Three cases of tamoxifen-associated venous thrombosis associated with factor V Leiden have been reported.121 Given the increasing use of selective estrogen receptor modulators for treating or preventing breast cancer and osteoporosis, it will be important to determine whether thrombophilic genetic risk factors (e.g., those listed in Table 127-1) will increase the liklihood of selective estrogen receptor modulator–associated venous thrombosis.
CHAPTER REFERENCES

1.
Egeberg O: Inherited antithrombin deficiency causing thrombophilia. Thromb Diath Haemorrh 13:516, 1963.

2.
Stenflo J, Fernlund P, Egan W, Roepstorff P: Vitamin K-dependent modifications of glutamic acid residues in prothrombin. Proc Natl Acad Sci 71:2730, 1974.

3.
Griffin JH, Evatt B, Zimmerman TS, Kleiss AJ: Deficiency of protein C in congenital thrombotic disease. J Clin Invest 68:1370, 1981.

4.
Schwarz HP, Fischer M, Hopmeier P, Batard MA, Griffin JH: Plasma protein S deficiency in familial thrombotic disease. Blood 64:1297, 1984.

5.
Comp PC, Nixon RR, Cooper MR, Esmon CT: Familial protein S deficiency is associated with recurrent thrombosis. J Clin Invest 74:2082, 1984.

6.
Comp PC, Esmon CT: Recurrent venous thromboembolism in patients with a partial deficiency of protein S. N Engl J Med 311:1525, 1984.

7.
Dahlback B, Carlsson M, Svensson PJ: Familial thrombophilia due to a previously unrecognized mechanism characterized by poor anticoagulant response to activated protein C: prediction of a cofactor to activated protein C. Proc Natl Acad Sci 90:1004, 1993.

8.
Svensson PJ, Dahlbäck B: Resistance to activated protein C as a basis for venous thrombosis. N Engl J Med 330:517, 1994.

9.
Bertina RM, Koeleman BPC, Koster T, et al: Mutation in blood coagulation factor V associated with resistance to activated protein C. Nature 369:64, 1994.

10.
Greengard JS, Sun X, Xu X, et al: Activated protein C resistance caused by Arg506Gln mutation in factor Va. Lancet 343:1361, 1994.

11.
Voorberg J, Roelse J, Koopman R, et al: Association of idiopathic venous thromboembolism with single point-mutation at Arg506 of factor V. Lancet 343:1535, 1994.

12.
Bienvenu T, Ankri A, Chadefaux B, Montalescot G, Kamoun P: Elevated total plasma homocysteine, a risk factor for thrombosis. Relation to coagulation and fibrinolytic parameters. Thromb Res 70:123, 1993.

13.
McCully KS: Vascular pathology of homocysteinemia: implications for the pathogenesis of arteriosclerosis. Am J Pathol 56:111, 1969.

14.
Poort SR, Rosendaal FR, Reitsma PH, Bertina RM: A common genetic variation in the 3′-untranslated region of the prothrombin gene is associated with elevated plasma prothrombin levels and an increase in venous thrombosis. Blood 88:3698, 1996.

15.
Bertina RM: Molecular risk factors for thrombosis. Thromb Haemost 82:601, 1999.

16.
Rosendaal FR: Risk factors for venous thrombotic disease. Thromb Haemost 82:610, 1999.

17.
Rosendaal FR, Siscovick DS, Schwartz SM, et al: Factor V Leiden (resistance to activated protein C) increases the risk of myocardial infarction in young women. Blood 89:2817, 1997.

18.
Zoller B, Garcia dF, Dahlback B: A common 4G allele in the promoter of the plasminogen activator inhibitor-1 (PAI-1) gene as a risk factor for pulmonary embolism and arterial thrombosis in hereditary protein S deficiency. Thromb Haemost 79:802, 1998.

19.
Silverstein MD, Heit JA, Mohr DN, et al: Trends in the incidence of deep vein thrombosis and pulmonary embolism: a 25-year population-based study. Arch Intern Med 158:585, 1998.

20.
Koster T, Rosendaal FR, Deronde H, et al: Venous thrombosis due to poor anticogaulant response to activated protein-C-Leiden Thrombophilia Study. Lancet 342:1503, 1993.

21.
Griffin JH, Evatt B, Wideman C, Fernandez JA: Anticoagulant protein C pathway defective in majority of thrombophilic patients. Blood 82:1989, 1993.

22.
Marciniak E, Romond EH: Impaired catalytic function of activated protein C: a new in vitro manifestation of lupus anticoagulant. Blood 74:2426, 1989.

23.
Malia RG, Kitchen S, Greaves M, Preston FE: Inhibition of activated protein C and its cofactor protein S by antiphospholipid antibodies. Br J Haematol 76:101, 1990.

24.
Amer L, Kisiel W, Searles RP, Williams RC Jr: Impairment of the protein C anticoagulant pathway in a patient with systemic lupus erythematosus, anticardiolipin antibodies and thrombosis. Thromb Res 57:247, 1990.

25.
Dahlback B, Carlsson M: Factor VIII defect associated with familial thrombophilia. Thromb Haemost 65:658, 1991.

26.
Zoller B, Dahlback B: Linkage between inherited resistance to activated protein C and factor V gene mutation in venous thrombosis. Lancet 343:1536, 1994.

27.
Zoller B, Svensson PJ, He X, Dahlback B: Identification of the same factor V gene mutation in 47 out of 50 thrombosis-prone families with inherited resistance to activated protein C. J Clin Invest 94:2521, 1994.

28.
De Visser MCH, Rosendaal FR, Bertina RM: A reduced sensitivity for activated protein C in the absence of factor V Leiden increases the risk of venous thrombosis. Blood 93:1271, 1999.

29.
Rodeghiero F, Tosetto A: Activated protein C resistance and factor V Leiden mutation are independent risk factors for venous thromboembolism. Ann Intern Med 130:643, 1999.

30.
Fisher M, Fernandez JA, Ameriso SF, et al: Activated protein C resistance in ischemic stroke not due to factor V arginine506®glutamine mutation. Stroke 27:1163, 1996.

31.
Van der Bom JG, Bots ML, Haverkate F, et al: Reduced response to activated protein C is associated with increased risk for cerebrovascular disease. Ann Intern Med 125:265, 1996.

32.
Oosting JD, Derksen RHWM, Bobbink IWG, et al: Antiphospholipid antibodies directed against a combination of phospholipids with prothrombin, protein C, or protein S: an explanation for their pathogenic mechanism? Blood 81:2618, 1993.

33.
Zivelin A, Gitel S, Griffin JH, et al: Extensive venous and arterial thrombosis associated with an inhibitor to activated protein C. Blood 94:895, 1999.

34.
Zivelin A, Griffin JH, Xu X, et al: A single genetic origin for a common caucasian risk factor for venous thrombosis. Blood 89:397, 1997.

35.
Sun X, Evatt B, Griffin JH: Blood coagulation factor Va abnormality associated with resistance to activated protein C in venous thrombophilia. Blood 83:3120, 1994.

36.
Heeb MJ, Kojima Y, Greengard JS, Griffin JH: Activated protein C resistance: molecular mechanisms based on studies using purified Gln506-factor V. Blood 85:3405, 1995.

37.
Kalafatis M, Bertina RM, Rand MD, Mann KG: Characterization of the molecular defect in factor VR506Q. J Biol Chem 270:4053, 1995.

38.
Nicolaes GAF, Tans G, Thomassen MCLGD, et al: Peptide bond cleavages and loss of functional activity during inactivation of factor Va and factor VaR506Q by activated protein C. J Biol Chem 270:21158, 1995.

39.
Rosing J, Hoekema L, Nicolaes GA, et al: Effects of protein S and factor Xa on peptide bond cleavages during inactivation of factor Va and factor VaR506Q by activated protein C. J Biol Chem 270:27852, 1995.

40.
Kalafatis M, Rand MD, Mann KG: The mechanism of inactivation of human factor V and human factor Va by activated protein C. J Biol Chem 269:31869, 1994.

41.
Shen L, Dahlback B: Factor V and protein S as synergistic cofactors to activated protein C in degradation of factor VIIIa. J Biol Chem 269:18735, 1994.

42.
Dahlback B, Hildebrand B: Inherited resistance to activated protein C is corrected by anticoagulant cofactor activity found to be a property of factor V. Proc Natl Acad Sci 91:1396, 1994.

43.
Thorelli E, Kaufman RJ, Dahlback B: Cleavage of factor V at Arg 506 by activated protein C and the expression of anticoagulant activity of factor V. Blood 93:2552, 1999.

44.
Dahlback B: Procoagulant and anticoagulant properties of coagulation factor V: factor V Leiden (APC resistance) causes hypercoagulability by dual mechanisms. J Lab Clin Med 133:415, 1999.

45.
Varadi K, Rosing J, Tans G, et al: Factor V enhances the cofactor function of protein S in the APC-mediated inactivation of factor VIII: influence of the factor VR506Q mutation. Thromb Haemost 76:208, 1996.

46.
Williamson D, Brown K, Luddington R, Baglin C, Baglin T: Factor V Cambridge: a new mutation (Arg306®Thr) associated with resistance to activated protein C. Blood 91:1140, 1998.

47.
Chan WP, Lee CK, Kwong YL, Lam CK, Liang R: A novel mutation of Arg306 of factor V gene in Hong Kong Chinese. Blood 91:1135, 1998.

48.
Franco RF, Maffei FH, Lourenco D, et al: Factor VArg306®Thr (factor V Cambridge) and factor V Arg306®Gly mutations in venous thrombotic disease. Br J Haematol 103:888, 1998.

49.
Bernardi F, Faioni EM, Castoldi E, et al: A factor V genetic component differing from factor V R506Q contributes to the activated protein C resistance phenotype. Blood 90:1552, 1997.

50.
Alhenc-Gelas M, Nicaud V, Gandrille S, et al: The factor V gene A4070G mutation and the risk of venous thrombosis. Thromb Haemost 81:193, 1999.

51.
Griffin JH: Blood coagulation. The thrombin paradox [news; comment]. Nature 378:337, 1995.

52.
Gruber A, Griffin JH. Direct detection of activated protein C in blood from human subjects. Blood 79:2340, 1992.

53.
Okajima K, Koga S, Kaji M, et al: Effect of protein C and activated protein C on coagulation and fibrinolysis in normal human subjects. Thromb Haemost 63:48, 1990.

54.
Heeb MJ, Gruber A, Griffin JH: Identification of divalent metal ion-dependent inhibition of activated protein C by alpha 2-macroglobulin and alpha 2-antiplasmin in blood and comparisons to inhibition of factor Xa, thrombin, and plasmin. J Biol Chem 266:17606, 1991.

55.
Fernandez JA, Petaja J, Gruber A, Griffin JH: Activated protein C correlates inversely with thrombin levels in resting healthy individuals. Am J Hematol 56:29, 1997.

56.
Greengard JS, Eichinger S, Griffin JH, Bauer KA: Variability of thrombosis among homozygous siblings with resistance to activated protein C due to an Arg®Gln mutation in the gene for factor V. N Engl J Med 331:1559, 1994.

57.
Martinelli I, Bottasso B, Duca F, Faioni E, Mannucci PM: Heightened thrombin generation in individuals with resistance to activated protein C. Thromb Haemost 75:703, 1996.

58.
Simioni P, Scarano L, Gavasso S, et al: Prothrombin fragment 1+2 and thrombin-antithrombin complex levels in patients with inherited APC resistance due to factor V Leiden mutation. Br J Haematol 92:435, 1996.

59.
Zoller B, Holm J, Svensson P, Dahlback B: Elevated levels of prothrombin activation fragment 1+2 in plasma from patients with heterozygous Arg506 to Gln mutation in the factor V gene (APC-resistance) and/or inherited protein S deficiency. Thromb Haemost 75:270, 1996.

60.
Lindqvist PG, Svensson PJ, Dahlback B, Marsal K: Factor V Q506 mutation (activated protein C resistance) associated with reduced intrapartum blood loss—a possible evolutionary selection mechanism. Thromb Haemost 79:69, 1998.

61.
Nichols WC, Amano K, Cacheris PM, et al: Moderation of hemophilia A phenotype by the factor V R506Q mutation. Blood 88:1183, 1996.

62.
Rees DC, Cox M, Clegg JB: World distribution of factor V Leiden. Lancet 346:1133, 1995.

63.
Ridker PM, Miletich JP, Hennekens CH, Buring JE: Ethnic distribution of factor V Leiden in 4047 men and women—implications for venous thromboembolism screening. JAMA 277:1305, 1997.

64.
Gregg JP, Yamane AJ, Grody WW. Prevalence of the factor V-Leiden mutation in four distinct American ethnic populations. Am J Med Genet 73:334, 1997;

65.
Dahlback B: Resistance to activated protein C as risk factor for thrombosis: molecular mechanisms, laboratory investigation, and clinical management. Sem Hematol 34:217, 1997.

66.
Turkstra F, Karemaker R, Kuijer PMM, Prins MH, Büller HR: Is the prevalence of the factor V Leiden mutation in patients with pulmonary embolism and deep vein thrombosis really different? Thromb Haemost 81:345, 1999.

67.
Manten B, Westendorp RGJ, Koster T, Reitsma PH, Rosendaal FR: Risk factor profiles in patients with different clinical manifestations of venous thromboembolism: a focus on the factor V Leiden mutation. Thromb Haemost 76:510, 1996.

68.
Desmarais S, De Moerloose P, Reber G, et al: Resistance to activated protein C in an unselected population of patients with pulmonary embolism. Lancet 347:1374, 1996.

69.
Vandenbroucke JP, Bertina RM, Holmes ZR, et al: Factor V Leiden and fatal pulmonary embolism. Thromb Haemost 79:511, 1998.

70.
Munkvad S, Jorgensen M: Resistance to activated protein C: a common anticoagulant deficiency in patients with venous leg ulceration. Br J Dermatol 134:296, 1996.

71.
Maessen-Visch MB, Hamulyak K, Tazelaar DJ, Crombag NHCM, Neumann HAM: The prevalence of factor V Leiden mutation in patients with leg ulcers and venous insufficiency. Arch Dermatol 135:41, 1999.

72.
Zuber M, Toulon P, Marnet L, Mas JL: Factor V Leiden mutation in cerebral venous thrombosis. Stroke 27:1721, 1996.

73.
Lüdemann P, Nabavi DG, Junker R, et al: Factor V Leiden mutation is a risk factor for cerebral venous thrombosis—a case-control study of 55 patients. Stroke 29:2507, 1998.

74.
Leebeek FWG, Lameris JS, Van Buuren HR, et al: Budd-Chiari syndrome, portal vein and mesenteric vein thrombosis in a patient homozygous for factor V Leiden mutation treated by TIPS and thrombolysis. Br J Haematol 102:929, 1998.

75.
Middeldorp S, Henkens CMA, Koopman MMW, et al: The incidence of venous thromboembolism in family members of patients with factor V Leiden mutation and venous thrombosis. Ann Intern Med 128:15, 1998.

76.
Martinelli I, Mannucci PM, De Stefano V, et al: Different risks of thrombosis in four coagulation defects associated with inherited thrombophilia: a study of 150 families. Blood 92:2353, 1998.

77.
Bucciarelli P, Rosendaal FR, Tripodi A, et al: Risk of venous thromboembolism and clinical manifestations in carriers of antithrombin, protein C, protein S deficiency, or activated protein C resistance—a multicenter collaborative family study. Arterioscler Thromb Vasc Biol 19:1026, 1999.

78.
Rosendaal FR: Venous thrombosis: a multicausal disease. Lancet 353:1167, 1999.

79.
Ridker PM, Glynn RJ, Miletich JP, et al: Age-specific incidence rates of venous thromboembolism among heterozygous carriers of factor V Leiden mutation. Ann Intern Med 126:528, 1997.

80.
Ridker PM, Hennekens CH, Lindpaintner K, et al: Mutation in the gene coding for coagulation facator V and the risk of myocardial infarction, stroke, and venous thrombosis in apparently healthy men. N Engl J Med 332:912, 1995.

81.
Price DT, Ridker PM: Factor V Leiden mutation and the risks for thromboembolic disease: a clinical perspective. Ann Intern Med 127:895, 1997.

82.
Ridker PM, Miletich JP, Stampfer MJ, et al: Factor V Leiden and risks of recurrent idiopathic venous thromboembolism. Circulation 92:2800, 1995.

83.
Simioni P, Prandoni P, Lensing AWA, et al: The risk of recurrent venous thromboembolism in patients with an Arg506®Gln mutation in the gene for factor V (factor Leiden). N Engl J Med 336:399, 1997.

84.
Baglin C, Brown K, Luddington R, Baglin T: Rick of recurrent venous thromboembolism in patients with the factor V Leiden (FVR 506Q) mutation: effect of warfarin and prediction by precipitating factors. Br J Haematol 100:764, 1998.

85.
Eichinger S, Minar E, Hirschl M, et al: The risk of early recurrent venous thromboembolism after oral anticoagulant therapy in patients with the G20210A transition in the prothrombin gene. Thromb Haemost 81:14, 1999.

86.
Rintelen C, Pabinger I, Knöbl P, Lechner K, Mannhalter C: Probability of recurrence of thrombosis in patients with and without factor V Leiden. Thromb Haemost 75:229, 1996.

87.
Lindmarker P, Schulman S, Sten-Linder M, et al: The risk of recurrent venous thromboembolism in carriers and non-carriers of the G1691A allele in the coagulation factor V gene and the G20210A allele in the prothrombin gene. Thromb Haemost 81:684, 1999.

88.
Rosendaal FR, Koster T, Vandenbroucke JP, Reitsma PH: High risk of thrombosis in patients homozygous for factor V Leiden (activated protein C resistance). Blood 85:1504, 1995.

89.
Emmerich J, Alhenc-Gelas M, Aillaud MF, et al: Clinical features in 36 patients homozygous for the ARG 506®GLN factor V mutation. Thromb Haemost 77:620, 1997.

90.
Mari D, Mannucci PM, Duca F, Bertolini S, Franceschi C: Mutant factor V (Arg506Gln) in health centenarians. Lancet 347:1044, 1996.

91.
Heijmans BT, Westendorp RGJ, Knook DL, Kluft C, Slagboom PE: The risk of mortality and the factor V Leiden mutation in a population-based cohort. Thromb Haemost 80:607, 1998.

92.
Hille ETM, Westendorp RGJ, Vandenbroucke JP, Rosendaal FR: Mortality and causes of death in families with the factor V Leiden mutation (resistance to activated protein C). Blood 89:1963, 1997.

93.
Rees DC, Liu YT, Cox MJ, Elliott P, Wainscoat JS: Factor V Leiden and thermolabile methylenetetrahydrofolate reductase in extreme old age. Thromb Haemost 78:1357, 1997.

94.
Inbal A, Freimark D, Modan B, et al: Synergistic effects of prothrombotic polymorphisms and atherogenic factors on the risk of myocardial infarction in young males. Blood 93:2186, 1999.

95.
Doggen CJM, Cats VM, Bertina RM, Rosendaal FR: Interaction of coagulation defects and cardiovascular risk factors—increased risk of myocardial infarction associated with factor V Leiden or prothrombin 20210A. Circulation 97:1037, 1998.

96.
Press RD, Liu XY, Beamer N, Coull BM: Ischemic stroke in the elderly—role of the common factor V mutation causing resistance to activated protein C. Stroke 27:44, 1996.

97.
Van Bockxmeer FM, Baker RI, Taylor RR: Premature ischaemic heart disease and the gene for coagulation factor V. Nature Med 1:185, 1995.

98.
Cushman M, Rosendaal FR, Psaty BM, et al: Factor V Leiden is not a risk factor for arterial vascular disease in the elderly: results from the Cardiovascular Health Study. Thromb Haemost 79:912, 1998.

99.
Becker S, Heller CH, Gropp F, Scharrer I, Kreuz W: Thrombophilic disorders in children with cerebral infarction. Lancet 352:1756, 1998.

100.
Nowak-Gottl U, Koch HG, Aschka I, et al: Resistance to activated protein C (APCR) in children with venous or arterial thromboembolism. Br J Haematol 92:992, 1996.

101.
Sifontes MT, Nuss R, Hunger SP, et al: Activated protein C resistance and the factor V Leiden mutation in children with thrombosis. Am J Hematol 57:29, 1998.

102.
Salomon O, Steinberg DM, Zivelin A, et al: Single and combined prothrombotic factors in patients with idiopathic venous thromboembolism—prevalence and risk assessment. Arterioscler Thromb Vasc Biol 19:511, 1999.

103.
Gandrille S, Greengard JS, Alhenc-Gelas M, et al: Incidence of activated protein C resistance caused by the ARG 506 GLN mutation in factor V in 113 unrelated symptomatic protein C-deficient patients. The French Network on the behalf of INSERM. Blood 86:219, 1995.

104.
Brenner B, Zivelin A, Lanir N, et al: Venous thromboembolism associated with double heterozygosity for R506Q mutation of factor V and for T298M mutation of protein C in a large family of a previously described homozygous protein C deficient newborn with massive thrombosis. Blood 88:877, 1996.

105.
Koeleman BPC, Reitsma PH, Allaart CF, Bertina RM: Activated protein C resistance as an additional risk factor for thrombosis in protein C-deficient families. Blood 84:1031, 1994.

106.
Zoller B, Berntsdotter A, Garcia de Frutos P, Dahlback B: Resistance to activated protein C as an additional genetic risk factor in hereditary deficiency of protein S. Blood 85:3518, 1995.

107.
Zoller B, He X, Dahlback B: Homozygous APC-resistance combined with inherited type I protein S deficiency in a young boy with severe thrombotic disease. Thromb Haemost 73:743, 1995.

108.
Koeleman BP, van Rumpt D, Hamulyak K, Reitsma PH, Bertina RM: Factor V Leiden: an additional risk factor for thrombosis in protein S deficient families? Thromb Haemost 74:580, 1995.

109.
Van Boven HH, Reitsma PH, Rosendaal FR, et al: Factor V Leiden (FV R506Q) in families with inherited antithrombin deficiency. Thromb Haemost 75:417, 1996.

110.
Tosetto A, Rodeghiero F, Martinelli I, et al: Additional genetic risk factors for venous thromboembolism in carriers of the factor V Leiden mutation. Br J Haematol 103:871, 1998.

111.
Ehrenforth S, Prondsinski MV, Aygören-Pürsün E, Scharrer I, Ganser A: Study of the prothrombin gene 20210 GA variant in FV:Q506 carriers in relationship to the presence or absence of juvenile venous thromboembolism. Arterioscler Thromb Vasc Biol 19:276, 1999.

112.
Mandel H, Brenner B, Berant M, et al: Coexistence of hereditary homocystinuria and Factor V Leiden—effect on thrombosis. N Engl J Med 334:763, 1996.

113.
Cattaneo M, Tsai MY, Bucciarelli P, et al: A common mutation in the methylenetetrahydrofolate reductase gene (C677T) increases the risk for deep-vein thrombosis in patients with mutant factor V (factor V:Q506). Arterioscler Thromb Vasc Biol 17:1662, 1997.

114.
Ridker PM, Hennekens CH, Selhub J, et al: Interrelation of hyperhomocysteinemia, factor V Leiden, and risk of future venous thromboembolism. Circulation 95:1777, 1997.

115.
Dizon-Townson DS, Nelson LM, Jang H, Varner MW, Ward K: The incidence of the factor V Leiden mutation in an obstetric population and its relationship to deep vein thrombosis. Am J Obstet Gynecol 176:883, 1997.

116.
Hallak M, Senderowicz J, Cassel A, et al: Activated protein C resistance (factor V Leiden) associated with thrombosis in pregnancy. Am J Obstet Gynecol 176:889, 1997.

117.
Bokarewa MI, Bremme K, Blomback M: Arg506-Gln mutation in factor V and risk of thrombosis during pregnancy. Br J Haematol 92:473, 1996.

118.
Bloemenkamp KWM, Rosendaal FR, Helmerhorst FM, Büller HR, Vandenbroucke JP: Enhancement by factor V Leiden mutation of risk of deep-vein thrombosis associated with oral contraceptives containing third- generation progestagen. Lancet 346:1593, 1995.

119.
Vandenbroucke JP, Koster T, Brit E, et al: Increased risk of venous thrombosis in oral-contraceptive users who are carriers of factor V Leiden mutation. Lancet 344:1453, 1994.

120.
Rintelen C, Mannhalter C, Ireland H, et al: Oral contraceptives enhance the risk of clinical manifestation of venous thrombosis at a young age in females homozygous for factor V Leiden. Br J Haematol 93:487, 1996.

121.
Weitz IC, Israel VK, Liebman HA: Tamoxifen-associated venous thrombosis and activated protein C resistance due to factor V Leiden. Cancer 79:2024, 1997.

122.
Trossart M, Conard J, Horellou MH, et al: Modified APC resistance assay for patients on oral anticoagulants. Lancet 344:1709, 1994.

123.
Svensson PJ, Zoller B, Dahlback B: Evaluation of original and modified APC-resistance tests in unselected outpatients with clinically suspected thrombosis and in healthy controls. Thromb Haemost 77:332, 1997.

124.
Kapiotis S, Quehenberger P, Jilma B, et al: Improved characteristics of aPC-resistance assay. Coatest aPC resistance by predilution of samples with factor V deficiency plasma. Am J Clin Pathol 106:588, 1996.

125.
Dahlback B: Resistance to activated protein C, the Arg506 to Gln mutation in the factor V gene, and venous thrombosis. Thromb Haemost 73:739, 1995.

126.
Jorquera JI, Montoro JM, Fernandez MA, Aznar JA, Aznar J: Modified test for activated protein C resistance. Lancet 344:1162, 1994.

127.
Le DT, Griffin JH, Greengard JS, Mujumdar V, Rapaport SI: Use of a generally applicable tissue factor-dependent factor V assay to detect activated protein C-resistant factor Va in patients receiving warfarin and in patients with a lupus anticoagulant. Blood 85:1704, 1995.

128.
Griffin JH, Kojima K, Banka CL, Curtiss LK, Fernández JA: High-density lipoprotein enhancement of anticoagulant activities of plasma protein S and activated protein C. J Clin Invest 103:219, 1999.

129.
Curvers J, Thomassen MCLG, Nicolaes GAF, et al: Acquired APC resistance and oral contraceptives: differences between two functional tests. Br J Haematol 105:88, 1999.

130.
Stearns-Kurosawa DJ, Kurosawa S, Mollica JS, Ferrell GL, Esmon CT: The endothelial cell protein C receptor augments protein C activation by the thrombin-thrombomodulin complex. Proc Natl Acad Sci 93:10212, 1996.

131.
Cooper PC, Abuzenadah A, Preston FE: APC resistance test, a new phenomenon—the role of platelets. Br J Haematol 86(suppl):33, 1999.

132.
Shizuka R, Kanda T, Amagai H, Kobayashi I: False-positive activated protein C (APC) sensitivity ratio caused by freezing and by contamination of plasma with platelets. Thromb Res 78:189, 1995.

133.
Simioni P, Scudeller A, Radossi P, et al: “Pseudo homozygous” activated protein C resistance due to double heterozygous factor V defects (factor V Leiden mutation and type I quantitative factor V defect) associated with thrombosis: report of two cases belonging to two unrelated kindreds. Thromb Haemost 75:422, 1996.

134.
Castoldi E, Kalafatis M, Lunghi B, et al: Molecular bases of pseudo-homozygous APC resistance: the compound heterozygosity for FV R506Q and a FV Null mutation results in the exclusive presence of FV Leiden molecules in plasma. Thromb Haemost 80:403, 1998.

135.
Kalafatis M, Bernardi F, Simioni P, et al: Phenotype and genotype expression in pseudohomozygous factor VLEIDEN—the need for phenotype analysis. Arterioscler Thromb Vasc Biol 19:336, 1999.

136.
Zivelin A, Rosenberg N, Faier S, et al: A single genetic origin for the common prothrombotic G20210A polymorphism in the prothrombin gene. Blood 92:1119, 1998.

137.
Rosendaal FR, Doggen CJM, Zivelin A, et al: Geographic distribution of the 20210 G to A prothrombin variant. Thromb Haemost 79:706, 1998.

138.
Souto JC, Coll I, Llobet D, et al: The prothrombin 20210A allele is the most prevalent genetic risk factor for venous thromboembolism in the Spanish population. Thromb Haemost 80:366, 1998.

139.
Rosendaal FR, Vos HL, Poort SL, Bertina RM: Prothrombin 20210A variant and age at thrombosis. Thromb Haemost 79:444, 1998.

140.
Leroyer C, Mercier B, Oger E, et al: Prevalence of 20210 A allele of the prothrombin gene in venous thromboembolism patients. Thromb Haemost 80:49, 1998.

141.
Arruda VR, Annichino-Bizzacchi JM, Gonçalves MS, Costa FF: Prevalence of the prothrombin gene variant (nt20210A) in venous thrombosis and arterial disease. Thromb Haemost 78:1430, 1997.

142.
Margaglione M, Brancaccio V, Giuliani N, et al: Increased risk for venous thrombosis in carriers of the prothrombin G®A20210 gene variant. Ann Intern Med 129:89, 1998.

143.
Hillarp A, Zoller B, Svensson PJ, Dahlback B: The 20210 A allele of the prothrombin gene is a common risk factor among Swedish outpatients with verified deep venous thrombosis. Thromb Haemost 78:990, 1997.

144.
Cumming AM, Keeney S, Salden A, et al: The prothrombin gene G 20210A variant: Prevalence in a U.K. anticoagulant clinic population. Br J Haematol 98:353, 1997.

145.
Brown K, Luddington R, Williamson D, Baker P, Baglin T: The risk of venous thromboembolism associated with a G to A transition at position 20210 in the 3′-untranslated region of the prothrombin gene. Br J Haematol 98:907, 1997.

146.
Ferraresi P, Marchetti G, Legnani C, et al: The heterozygous 20210 G/A prothrombin genotype is associated with early venous thrombosis in inherited thrombophilias and is not increased in frequency in artery disease. Arterioscler Thromb Vasc Biol 17:2418, 1997.

147.
Simioni P, Tormene D, Manfrin D, et al: Prothrombin antigen levels in symptomatic and asymptomatic carriers of the 20210A prothrombin variant. Br J Haematol 103:1045, 1998.

148.
De Stefano V, Chiusolo P, Paciaroni K, et al: Hepatic vein thrombosis in a patient with mutant prothrombin 20210A allele. Thromb Haemost 80:519, 1998.

149.
Darnige L, Jezequel P, Amoura Z, et al: Mesenteric venous thrombosis in two patients heterozygous for the 20210 A allele of the prothrombin gene. Thromb Haemost 80:703, 1998.

150.
Martinelli I, Sacchi E, Landi G, et al: High risk of cerebral-vein thrombosis in carriers of a prothrombin-gene mutation and in users of oral contraceptives. N Engl J Med 338:1793, 1998.

151.
Biousse V, Conard J, Brouzes C, et al: Frequency of the 20210 G®A mutation in the 3′-untranslated region of the prothrombin gene in 35 cases of cerebral venous thrombosis. Stroke 29:1398, 1998.

152.
Chamouard P, Pencreach E, Maloisel F, et al: Frequent factor II G20210A mutation in idiopathic portal vein thrombosis. Gastroenterology 116:144, 1999.

153.
Reuner KH, Ruf A, Grau A, et al: Prothrombin gene G20210®A transition is a risk factor for cerebral venous thrombosis. Stroke 29:1765, 1998.

154.
Zawadzki C, Gaveriaux V, Trillot N, et al: Homozygous G20210A transition in the prothrombin gene associated with severe venous thrombotic disease: two cases in a French family. Thromb Haemost 80:1027, 1998.

155.
De Stefano V, Chiusolo P, Paciaroni K, et al: Prothrombin G20210A mutant genotype is a risk factor for cerebrovascular ischemic disease in young patients. Blood 91:3562, 1998.

156.
Howard TE, Marusa M, Channell C, Duncan A: A patient homozygous for a mutation in the prothrombin gene 3”-untranslated region associated with massive thrombosis. Blood Coagul Fibrinolysis 8:316, 1997.

157.
Makris M, Preston FE, Beauchamp NJ, et al: Co-inheritance of the 20210A allele of the prothrombin gene increases the risk of thrombosis in subjects with familial thrombophilia. Thromb Haemost 78:1426, 1997.

158.
Corral J, Gonzalez-Conejero R, Lozano ML, et al: The venous thrombosis risk factor 20210 A allele of the prothrombin gene is not a major risk factor for arterial thrombotic disease. Br J Haematol 99:304, 1997.

159.
Eikelboom JW, Baker RI, Parsons R, Taylor RR, Van Bockxmeer FM: No association between the 20210 G/A prothrombin gene mutation and premature coronary artery disease. Thromb Haemost 80:878, 1998.

160.
Redondo M, Watzke HH, Stucki B, et al: Coagulation factors II, V, VII, and X, prothrombin gene 20210G®A transition, and factor V Leiden in coronary artery disease—high factor V clotting activity is an independent risk factor for myocardial infarction. Arterioscler Thromb Vasc Biol 19:1020, 1999.

161.
Ridker PM, Hennekens CH, Miletich JP: G20210A mutation in prothrombin gene and risk of myocardial infarction, stroke, and venous thrombosis in a large cohort of US men. Circulation 99:999, 1999.

162.
Rosendaal FR, Siscovick DS, Schwartz SM, et al: A common prothrombin variant (20210 G to A) increases the risk of myocardial infarction in young women. Blood 90:1747, 1997.

163.
Franco RF, Trip MD, Ten Cate H, et al: The 20210 G®A mutation in the 3′-untranslated region of the prothrombin gene and the risk for arterial thrombotic disease. Br J Haematol 104:50, 1999.

164.
Gardemann A, Arsic T, Katz N, et al: The factor II G20210A and factor V G1691A gene transitions and coronary heart disease. Thromb Haemost 81:208, 1999.

165.
Mercier E, Quere I, Campello C, Mares P, Gris JC: The 20210A allele of the prothrombin gene is frequent in young women with unexplained spinal cord infarction. Blood 92:1840, 1998.

166.
Mudd SH, Levy Hl, Skovby F: Disorders of transsulfuration, in Scriver CR, Beaudet AL, Sly WS, Valle D, The metabolic and molecular bases of inherited disease, p 1279. McGraw-Hill, New York, 1995.

167.
Cattaneo M: Hyperhomocysteinemia, atherosclerosis and thrombosis. Thromb Haemost 81:165, 1999.

168.
D’Angelo A, Selhub J: Homocysteine and thrombotic disease. Blood 90:1, 1997.

169.
Fermo I, D’Angelo SV, Paroni R, et al: Prevaleance of moderate hyperhomocysteinemia in patients with early-onset venous and arterial occlusive disease. Ann Intern Med 123:747, 1995.

170.
Den Heijer M, Rosendaal FR, Blom HJ, Gerrits WBJ, Bos GMJ: Hyperhomocysteinemia and venous thrombosis: a meta-analysis. Thromb Haemost 80:874, 1998.

171.
den Heijer M, Brouwer IA, Bos GMJ, et al: Vitamin supplementation reduces blood homocysteine levels—a controlled trial in patients with venous thrombosis and healthy volunteers. Arterioscler Thromb Vasc Biol 18:356, 1998.

172.
Woodside JV, Yarnell JWG, McMaster D, et al: Effect of B-group vitamins and antioxidant vitamins on hyperhomocysteinemia: a double-blind, randomized, factorial-design, controlled trial. Am J Clin Nutr 67:858, 1998.

173.
Harker LA, Ross R, Slichter SJ, Scott CR: Homocystine-induced arteriosclerosis: the role of endothelial cell injury and platelet response in its genesis. J Clin Invest 58:731, 1976.

174.
Lentz SR, Sobey CG, Piegors DJ, et al: Vascular dysfunction in monkeys with diet-induced hyperhomocyst(e)inemia. J Clin Invest 98:24, 1996.

175.
Lentz SR: Mechanisms of thrombosis in hyperhomocysteinemia. Curr Opin Hematol 5:343, 1998.

176.
Den Heijer M, Koster T, Blom HJ, et al: Hyperhomocysteinemia as a risk factor for deep-vein thrombosis. N Engl J Med 334:759, 1996.

177.
Den Heijer M, Blom HJ, Gerrits WBJ, et al: Is hyperhomocysteinaemia a risk factor for recurrent venous thrombosis? Lancet 345:882, 1995.

178.
Simioni P, Prandoni P, Burlina A, et al: Hyperhomocysteinemia and deep-vein thrombosis—a case-control study. Thromb Haemost 76:883, 1996.

179.
Falcon CR, Cattaneo M, Panzeri D, Martinelli I, Mannucci PM: High prevalence of hyperhomocyst(e)inemia in patients with juvenile venous thrombosis. Arterioscler Thromb 14:1080, 1994.

180.
Eichinger S, Stümpflen A, Hirschl M, et al: Hyperhomocysteinemia is a risk factor of recurrent venous thromboembolism. Thromb Haemost 80:566, 1998.

181.
Kyrle PA, Stümpflen A, Hirschl M, et al: Levels of prothrombin fragment F1+2 in patients with hyperhomocysteinemia and a history of venous thromboembolism. Thromb Haemost 78:1327, 1997.

182.
Cattaneo M, Franchi F, Zighetti ML, et al: Plasma levels of activated protein C in healthy subjects and patients with previous venous thromboembolism—relationships with plasma homocysteine levels. Arterioscler Thromb Vasc Biol 18:1371, 1998.

183.
Kluijtmans LAJ, Boers GHJ, Verbruggen B, et al: Homozygous cystathionine b-synthase deficiency, combined with factor V Leiden or thermolabile methylenetetrahydrofolate reductase in the risk of venous thrombosis. Blood 91:2015, 1998.

184.
Kang SS, Zhou J, Wong PW, Kowalisyn J, Strokosch G: Intermediate homocysteinemia: a thermolabile variant of methylenetetrahydrofolate reductase. Am J Hum Genet 43:414, 1988.

185.
De Franchis R, Mancini FP, D’Angelo A, et al: Elevated total plasma homocysteine and 677C®T mutation of the 5,10-methylenetetrahydrofolate reductase gene in thrombotic vascular disease [letter]. Am J Hum Genet 59:262, 1996.

186.
Margaglione M, D’Andrea G, D’Addedda M, et al: The methylenetetrahydrofolate reductase TT677 genotype is associated with venous thrombosis independently of the coexistence of the FV leiden and the prothrombin A20210 mutation. Thromb Haemost 79:907, 1998.

187.
Kluijtmans LAJ, den Heijer M, Reitsma PH, et al: Thermolabile methylenetetrahydrofolate reductase and factor V Leiden in the risk of deep-vein thrombosis. Thromb Haemost 79:254, 1998.

188.
Legnani C, Palareti G, Grauso F, et al: Hyperhomocyst(e)inemia and a common methylenetetrahydrofolate reductase mutation (Ala223Val MTHFR) in patients with inherited thrombophilic coagulation defects. Arterioscler Thromb Vasc Biol 17:2924, 1997.

189.
Tosetto A, Missiaglia E, Frezzato M, Rodeghiero F: The VITA Project: C677T mutation in the methylene-tetrahydrofolate reductase gene and risk of venous thromboembolism. Br J Haematol 97:804, 1997.

190.
Garg UC, Zheng ZJ, Folsom AR, et al: Short-term and long-term variability of plasma homocysteine measurement. Clin Chem 43:141, 1997.

191.
Refsum H, Fiskerstrand T, Guttormsen AB, Ueland PM: Assessment of homocysteine status. J Inherit Metab Dis 20:286, 1997.

192.
Miner SES, Evrovski J, Cole DEC: Clinical chemistry and molecular biology of homocysteine metabolism: an update. Clin Biochem 30:189, 1997.

193.
Van Der Griend R, Haas FJLM, Duran M, et al: Methionine loading test is necessary for detection of hyperhomocysteinemia. J Lab Clin Med 132:67, 1998.

194.
Welch GN, Loscalzo J: Homocysteine and atherothrombosis. N Engl J Med 338:1042, 1998.

195.
Jacobsen DW, Gatautis VJ, Green R, et al: Rapid HPLC determination of total homocysteine and other thiols in serum and plasma: sex differences and correlation with cobalamin and folate concentrations in healthy subjects [see comments]. Clin Chem 40:873, 1994.

196.
Frosst P, Blom HJ, Milos R, et al: A candidate genetic risk factor for vascular disease: a common mutation in methylenetetrahydrofolate reductase [letter]. Nat Genet 10:111, 1995.

197.
Branson HE, Katz J, Marble R, Griffin JH: Inherited protein C deficiency and coumarin-responsive chronic relapsing purpura fulminans in a newborn infant. Lancet 2:1165, 1983.

198.
Reitsma PH, Bernardi F, Doig RG, et al: Protein C deficiency: a database of mutations, 1995 update. On behalf of the Subcommittee on Plasma Coagulation Inhibitors of the Scientific and Standardization Committee of the ISTH. [Review] [112 refs]. Thromb Haemost 73:876, 1995.

199.
Miletich J, Sherman L, Broze G: Absence of thrombosis in subjects with heterozygous protein C deficiency. N Engl J Med 317:991, 1987.

200.
Tait RC, Walker ID, Reitsma PH, et al: Prevalence of protein C deficiency in the healthy population. Thromb Haemost 73:87, 1995.

201.
Koster T, Rosendaal FR, Briët E, et al: Protein C deficiency in a controlled series of unselected outpatients: an infrequent but clear risk factor for venous thrombosis (Leiden thrombophilia study). Blood 85:2756, 1995.

202.
Bertina RM, Broekmans AW, van dL I, Mertens K: Protein C deficiency in a Dutch family with thrombotic disease. Thromb Haemost 48:1, 1982.

203.
Allaart CF, Poort SR, Rosendaal FR, et al: Increased risk of venous thrombosis in carriers of hereditary protein C deficiency defect. Lancet 341:134, 1993.

204.
Bovill EG, Bauer KA, Dickerman JD, Callas P, West B: The clinical spectrum of heterozygous protein C deficiency in a large New England kindred. Blood 73:712, 1989.

205.
Lensen RPM, Rosendaal FR, Koster T, et al: Apparent different thrombotic tendency in patients with factor V Leiden and protein C deficiency due to selection of patients. Blood 88:4205, 1996.

206.
Allaart CF, Rosendaal FR, Noteboom WMP, Vandenbroucke JP, Briet E: Survival in families with hereditary protein C deficiency, 1820 to 1993. Br Med J 311:910, 1995.

207.
Hasstedt SJ, Bovill EG, Callas PW, Long GL: An unknown genetic defect increases venous thrombosis risk, through interaction with protein C deficiency. Am J Hum Genet 63:569, 1998.

208.
Gandrille S, Priollet P, Capron L, et al: Association of inherited dysfibrinogenaemia and protein C deficiency in two unrelated families. Br J Haematol 68:329, 1988.

209.
Spek CA, Koster T, Rosendaal FR, Bertina RM, Reitsma PH: Genotypic variation in the promoter region of the protein C gene is associated with plasma protein C levels and thrombotic risk. Arterioscler Thromb Vasc Biol 15:214, 1995.

210.
Pabinger I, Schneider B: Thrombotic risk in hereditary antithrombin III, protein C, or protein S deficiency—a cooperative, retrospective study. Arterioscler Thromb Vasc Biol 16:742, 1996.

211.
De Stefano V, Leone G, Mastrangelo S, et al: Clinical manifestations and management of inherited thrombophilia: retrospective analysis and follow-up after diagnosis of 238 patients with congenital deficiency of antithrombin III, protein C, protein S. Thromb Haemost 72:352, 1994.

212.
Pabinger I, Kyrle PA, Heistinger M, et al: The risk of thromboembolism in asymptomatic patients with protein C and protein S deficiency: a prospective cohort study. Thromb Haemost 71:441, 1994.

213.
Van den Belt AGM, Sanson BJ, Simioni P, et al: Recurrence of venous thromboembolism in patients with familial thrombophilia. Arch Intern Med 157:2227, 1997.

214.
De Bruijn SFTM, Stam J, Koopman MMW, Vandenbroucke JP: Cerebral venous sinus thrombosis study: case-control study of risk of cerebral sinus thrombosis in oral contraceptive users who are carriers of hereditary prothrombotic conditions. Br Med J 316:589, 1998.

215.
Camerlingo M, Finazzi G, Casto L, et al: Inherited protein C deficiency and nonhemorrhagic arterial stroke in young adults. Neurology 41:1371, 1991.

216.
Seligsohn U, Berger A, Abend M, et al: Homozygous protein C deficiency manifested by massive venous thrombosis in the newborn. N Engl J Med 310:559, 1984.

217.
Sills RH, Marlar RA, Montgomery RR, Deshpande GN, Humbert JR: Severe homozygous protein C deficiency. J Pediatr 105:409, 1984.

218.
Marciniak E, Wilson HD, Marlar RA: Neonatal purpura fulminans: a genetic disorder related to the absence of protein C in blood. Blood 65:15, 1985.

219.
Monagle P, Andrew M, Halton J, et al: Homozygous protein C deficiency: Description of a new mutation and successful treatment with low molecular weight heparin. Thromb Haemost 79:756, 1998.

220.
McGehee WG, Klotz TA, Epstein DJ, Rapaport SI: Coumarin necrosis associated with hereditary protein C deficiency. Ann Int Med 101:59, 1984.

221.
Vigano D’A, Comp PC, Esmon CT, D’Angelo A: Relationship between protein C antigen and anticoagulant activity during oral anticoagulation and in selected disease states. J Clin Invest 77:416, 1986.

222.
Miletich JP: Laboratory diagnosis of protein C deficiency. [Review] [31 refs]. Semin Thromb Hemost 16:169, 1990.

223.
Francis RBJ, Seyfert U: Rapid amidolytic assay of protein C in whole plasma using an activator from the venom of Agkistrodon contortrix. Am J Clin Pathol 87:619, 1987.

224.
Aiach M, Borgel D, Gaussem P, et al: Protein C and protein S deficiencies. Semin Hematol 34:205, 1997.

225.
Berdeaux DH, Abshire TC, Marlar RA: Dysfunctional protein C deficiency (Type II). A report of 11 cases in 3 American families and review of the literature. Am J Clin Pathol 99:677, 1993.

226.
Aiach M, Nicaud V, Alhenc-Gelas M, et al: Complex association of protein C gene promoter polymorphism with circulating protein C levels and thrombotic risk. Arterioscler Thromb Vasc Biol 19:1573, 1999.

227.
Tait RC, Walker ID, Islam SI, et al: Protein C activity in healthy volunteers—influence of age, sex, smoking and oral contraceptives. Thromb Haemost 70:281, 1993.

228.
Engesser L, Broekmans AW, Briet E, Brommer EJP, Bertina RM: Hereditary protein S deficiency: clinical manifestations. Ann Intern Med 106:677, 1987.

229.
Reitsma PH, Ploos van Amstel HK, Bertina RM: Three novel mutations in five unrelated subjects with hereditary protein S deficiency type I. J Clin Invest 93:486, 1994.

230.
Simmonds RE, Ireland H, Lane DA, et al: Clarification of the risk for venous thrombosis associated with hereditary protein S deficiency by investigation of a large kindred with a characterized gene defect. Ann Intern Med 128:8, 1998.

231.
Bolan CD, Krishnamurti C, Tang DB, Carrington LR, Alving BM: Association of protein S deficiency with thrombosis in a kindred with increased levels of plasminogen activator inhibitor-1. Ann Intern Med 119:779, 1993.

232.
Broekmans AW, Bertina RM, Reinalda-Poot J, et al: Hereditary protein S deficiency and venous thrombo-embolism. A study in three Dutch families. Thromb Haemost 53:273, 1985.

233.
Gandrille S, Borgel D, Ireland H, et al: Protein S deficiency: a database of mutations. For the Plasma Coagulation Inhibitors Subcommittee of the Scientific and Standardization Committee of the International Society on Thrombosis and Haemostasis. Thromb Haemost 77:1201, 1997.

234.
Mahasandana C, Suvatte V, Chuansumrit A, et al: Homozygous protein S deficiency in an infant with purpura fulminans. J Pediatr 117:750, 1990.

235.
Pegelow CH, Ledford M, Young JN, Zilleruelo G: Severe protein S deficiency in a newborn. [Review] [26 refs]. Pediatrics 89:674, 1992.

236.
Pung-amritt P, Poor SR, Vos HL, et al: Compound heterozygosity for one novel and one recurrent mutation in a Thai patient with severe protein S deficiency. Thromb Haemost 81:189, 1999.

237.
Heeb MJ, Mesters RM, Tans G, Rosing J, Griffin JH: Binding of protein S to factor Va associated with inhibition of prothrombinase that is independent of activated protein C. J Biol Chem 268:2872, 1993.

238.
Heeb MJ, Rosing J, Bakker HM, et al: Protein S binds to and inhibits factor Xa. Proc Natl Acad Sci USA 91:2728, 1994.

239.
Hackeng TM, Van’t Veer C, Meijers JCM, Bouma BN: Human protein S inhibits prothrombinase complex activity on endothelial cells and platelets via direct interactions with factors Va and Xa. J Biol Chem 269:21051, 1994.

240.
Koppelman SJ, Hackeng TM, Sixma JJ, Bouma BN: Inhibition of the intrinsic factor X activating complex by protein S: evidence for a specific binding of protein S to factor VIII. Blood 86:1062, 1995.

241.
Koppelman SJ, Van’t Veer C, Sixma JJ, Bouma BN: Synergistic inhibition of the intrinsic factor X activation by protein S and C4b-binding protein. Blood 86:2653, 1995.

242.
Van’t Veer C, Hackeng TM, Biesbroeck D, Sixma JJ, Bouma BN: Increased prothrombin activation in protein S-deficient plasma under flow conditions on endothelial cell matrix: an independent anticoagulant function of protein S in plasma. Blood 85:1815, 1995.

243.
Faioni EM, Valsecchi C, Palla A, et al: Free protein S deficiency is a risk factor for venous thrombosis. Thromb Haemost 78:1343, 1997.

244.
Heijboer H, Brandjes DPM, Büller HR, Sturk A, Ten Cate JW: Deficiencies of coagulation-inhibiting and fibrinolytic proteins in outpatients with deep-vein thrombosis. N Engl J Med 323:1512, 1990.

245.
Finazzi G, Barbui T: Different incidence of venous thrombosis in patients with inherited deficiencies of antithrombin III, protein C and protein S. Thromb Haemost 71:15, 1994.

246.
Coller BS, Owen J, Jesty J, et al: Deficiency of plasma protein S, protein C, or antithrombin III and arterial thrombosis. Arteriosclerosis 7:456, 1987.

247.
Allaart CF, Aronson DC, Ruys T, et al: Hereditary protein S deficiency in young adults with arterial occlusive disease. Thromb Haemost 64:206, 1990.

248.
Grimaudo V, Gueissaz F, Hauert J, et al: Necrosis of skin induced by coumarin in a patient deficient in protein S. Br Med J 298:233, 1989.

249.
Comp PC, Thurnau GR, Welsh J, Esmon CT: Functional and immunologic protein S levels are decreased during pregnancy. Blood 68:881, 1986.

250.
Malm J, Laurell M, Dahlback B: Changes in the plasma levels of vitamin K-dependent proteins C and S and of C4b-binding protein during pregnancy and oral contraception. Br J Haematol 68:437, 1988.

251.
Comp PC, Doray D, Patton D, Esmon CT: An abnormal plasma distribution of protein S occurs in functional protein S deficiency. Blood 67:504, 1986.

252.
D’Angelo A, Vigano-D’Angelo S, Esmon CT, Comp PC: Acquired deficiences of protein S. Protein S activity during oral anticoagulation, in liver disease, and in disseminated intravascular coagulation. J Clin Invest 81:1445, 1988.

253.
Vigano-D’Angelo S, D’Angelo A, Kaufman CE, et al: Protein S deficiency occurs in the nephrotic syndrome. Ann Intern Med 107:42, 1987.

254.
Aadland E, Odegaard OR, Roseth A, Try K: Free protein S deficiency in patients with chronic inflammatory bowel disease. Scand J Gastroenterol 27:957, 1992.

255.
Parke AL, Weinstein RE, Bona RD, Maier DB, Walker FJ: The thrombotic diathesis associated with the presence of phospholipid antibodies may be due to low levels of free protein S. Am J Med 93:49, 1992.

256.
Ginsberg JS, Demers C, Brill-Edwards P, et al: Acquired free protein S deficiency is associated with antiphospholipid antibodies and increased thrombin generation in patients with systemic lupus erythematosus. Am J Med 98:379, 1995.

257.
Levin M, Eley BS, Louis J, et al: Postinfectious purpura fulminans caused by an autoantibody directed against protein S. J Pediatr 127:355, 1995.

258.
Blanco A, Bonduel M, Peñalva L, Hepner M, Lazzari M: Deep vein thrombosis in a 13-year-old boy with hereditary protein S deficiency and a review of the pediatric literature. Am J Hematol 45:330, 1994.

259.
D’Angelo A, Della Valle P, Crippa L, et al: Brief report: Autoimmune protein S deficiency in a boy with severe thromboembolic disease. N Engl J Med 328:1753, 1993.

260.
Bergmann F, Hoyer PF, Vigano D’Angelo S, et al: Severe autoimmune protein S deficiency in a boy with idiopathic purpura fulminans. Br J Haematol 89:610, 1995.

261.
Woods CR, Johnson CA: Varicella purpura fulminans associated with heterozygosity for factor V Leiden and transient protein S deficiency. Pediatrics 102:1208, 1998.

262.
Zoller B, Garcia de Frutos P, Dahlback B: Evaluation of the relationship between protein S and C4b-binding protein isoforms in hereditary protein S deficiency demonstrating type I and type III deficiencies to be phenotypic variants of the same genetic disease. Blood 85:3524, 1995.

263.
Wolf M, Boyer-Neumann C, Peynaud-Debayle E, et al: Clinical applications of a direct assay of free protein S antigen using monoclonal antibodies. A study of 59 cases. Blood Coagul Fibrinolysis 5:187, 1994.

264.
Amiral J, Grosley B, Boyer-Neumann C, et al: New direct assay of free protein S antigen using two distinct monoclonal antibodies specific for the free form. Blood Coagul Fibrinolysis 5:179, 1994.

265.
Faioni EM, Boyer-Neumann C, Franchi F, et al: Another protein S functional assay is sensitive to resistance to activated protein C. Thromb Haemost 72:648, 1994.

266.
Brunet D, Barthet MC, Morange PE, et al: Protein S deficiency: different biological phenotypes according to the assays used. Thromb Haemost 79:446, 1998.

267.
Wolf M, Boyer-Neumann C, Leroy-Matheron C, et al: Functional assay of protein S in 70 patients with congenital and acquired disorders. Blood Coagul Fibrinolysis 2:705, 1991.

268.
Gari M, Falkon L, Urrutia T, et al: The influence of low protein S plasma levels in young women, on the definition of normal range. Thromb Res 73:149, 1994.

269.
Van Wijnen M, Van’t Veer C, Meijers JCM, Bertina RM, Bouma BN: A plasma coagulation assay for an activated protein C-independent anticoagulant activity of protein S. Thromb Haemost 80:930, 1998.

270.
Demers C, Ginsberg JS, Hirsh J, Henderson P, Blajchman MA: Thrombosis in antithrombin-III-deficient persons. Report of a large kindred and literature review. Ann Intern Med 116:754, 1992.

271.
Van Boven HH, Lane DA: Antithrombin and its inherited deficiency states. Semin Hematol 34:188, 1997.

272.
Blajchman MA, Austin RC, Fernandez-Rachubinski F, Sheffield WP: Molecular basis of inherited human antithrombin deficiency. Blood 80:2159, 1992.

273.
Lane DA, Bayston T, Olds RJ, et al: Antithrombin mutation database: 2nd (1997) update. For the Plasma Coagulation Inhibitors Subcommittee of the Scientific and Standardization Committee of the International Society on Thrombosis and Haemostasis. Thromb Haemost 77:197, 1997.

274.
Sakuragawa N, Takahashi K, Kondo S, Koide T: Antithrombin III Toyama: a hereditary abnormal antithrombin III of a patient with recurrent thrombophlebitis. Thromb Res 31:305, 1983.

275.
Fischer AM, Cornu P, Sternberg C, et al: Antithrombin III Alger: a new homozygous AT III variant. Thromb Haemost 55:218, 1986.

276.
Okajima K, Ueyama H, Hashimoto Y, et al: Homozygous variant of antithrombin III that lacks affinity for heparin, AT III Kumamoto. Thromb Haemost 61:20, 1989.

277.
Boyer C, Wolf M, Vedrenne J, Meyer D, Larrieu MJ: Homozygous variant of antithrombin III: AT III Fontainebleau. Thromb Haemost 56:18, 1986.

278.
Tait RC, Walker ID, Perry DJ, et al: Prevalence of antithrombin deficiency in the healthy population. Br J Haematol 87:106, 1994.

279.
Van Boven HH, Vandenbroucke JP, Briët E, Rosendaal FR: Gene-gene and gene-environment interactions determine risk of thrombosis in families with inherited antithrombin deficiency. Blood 94:2590, 1999.

280.
Hirsh J, Piovella F, Pini M: Congenital antithrombin III deficiency. Incidence and clinical features. Am J Med 87 [suppl 3B]:34S, 1989.

281.
Rosendaal FR, Heijboer H, Briet E, et al: Mortality in hereditary antithrombin III deficiency—1830 to 1989. Lancet 337:260, 1991.

282.
Van Boven HH, Olds RJ, Thein S-L, et al: Hereditary antithrombin deficiency: heterogeneity of the molecular basis and mortality in Dutch families. Blood 84:4209, 1994.

283.
Coller BS, Owen J, Jesty J, et al: Deficiency of plasma protein S, protein C, or antithrombin III and arterial thrombosis. Arteriosclerosis 7:456, 1987.

284.
Levine MN, Hirsh J, Gent M, et al: A randomized trial comparing activated thromboplastin time with heparin assay in patients with acute venous thromboembolism requiring large daily doses of heparin. Arch Intern Med 154:49, 1994.

285.
De Boer AC, van Riel LA, den Ottolander GJ: Measurement of antithrombin III, alpha 2-macroglobulin and alpha 1-antitrypsin in patients with deep venous thrombosis and pulmonary embolism. Thromb Res 15:17, 1979.

286.
Marciniak E, Gockerman JP: Heparin-induced decrease in circulating antithrombin-III. Lancet 2:581, 1977.

287.
Von Kaulla E, Von Kaulla KN: Antithrombin 3 and diseases. Am J Clin Pathol 48:69, 1967.

288.
Damus PS, Wallace GA: Immunologic measurement of antithrombin III-heparin cofactor and alpha2 macroglobulin in disseminated intravascular coagulation and hepatic failure coagulopathy. Thromb Res 6:27, 1975.

289.
Kauffmann RH, Veltkamp JJ, van Tilburg NH, Van Es LA: Acquired antithrombin III deficiency and thrombosis in the nephrotic syndrome. Am J Med 65:607, 1978.

290.
Buchanan GR, Holtkamp CA: Reduced antithrombin III levels during L-asparaginase therapy. Med Pediatr Oncol 8:7, 1980.

291.
Weenink GH, Treffers PE, Vijn P, Smorenberg-Schoorl ME, Ten Cate JW: Antithrombin III levels in preeclampsia correlate with maternal and fetal morbidity. Am J Obstet Gynecol 148:1092, 1984.

292.
Abildgaard U, Lie M, Odegard OR: Antithrombin (heparin cofactor) assay with “new” chromogenic substrates (S-2238 and Chromozym TH). Thromb Res 11:549, 1977.

293.
Demers C, Henderson P, Blajchman MA, et al: An antithrombin III assay based on factor Xa inhibition provides a more reliable test to identify congenital antithrombin III deficiency than an assay based on thrombin inhibition. Thromb Haemost 69:231, 1993.

294.
Bohner J, Von Pape K-W, Blaurock M: Thrombin-based antithrombin assays show over-estimation of antithrombin III activity in patients on heparin therapy due to heparin cofactor II influence. Thromb Haemost 71:280, 1994.

295.
Sas G, Pepper DS, Cash JD: Plasma and serum antithrombin III: differentiation by crossed immunoelectrophoresis. Thromb Res 6:87, 1975.

296.
Koster T, Blann AD, Briët E, Vandenbroucke JP, Rosendaal FR: Role of clotting factor VIII in effect of von Willebrand factor on occurrence of deep-vein thrombosis. Lancet 345:152, 1995.

297.
O’Donnell J, Tuddenham EG, Manning R, et al: High prevalence of elevated factor VIII levels in patients referred for thrombophilia screening: role of increased synthesis and relationship to the acute phase reaction. Thromb Haemost 77:825, 1997.

298.
Kamphuisen PW, Eikenboom JC, Vos HL, et al: Increased levels of factor VIII and fibrinogen in patients with venous thrombosis are not caused by acute phase reactions. Thromb Haemost 81:680, 1999.

299.
Haverkate F, Samama M: Familial dysfibrinogenemia and thrombophilia. Report on a study of the SSC Subcommittee on Fibirnogen. Thromb Haemost 73:151, 1995.

300.
Martinez J: Congenital dysfibrinogenemia. Curr Opin Hematol 4:357, 1997.

301.
Aoki N, Moroi M, Sakata Y, Yoshida N, Matsuda M: Abnormal plasminogen. A hereditary molecular abnormality found in a patient with recurrent thrombosis. J Clin Invest 61:1186, 1978.

302.
Miyata T, Iwanaga S, Sakata Y, Aoki N: Plasminogen Tochigi: Inactive plasmin resulting from replacement of alanine-600 by threonine in the active site. Proc Natl Acad Sci USA 79:6132, 1982.

303.
Miyata T, Iwanaga S, Sakata Y, et al: Plasminogens Tochigi II and Nagoya: two additional molecular defects with Ala600Val replacement found in plasmin light chain variants. J Biochem (Tokyo) 96:277, 1984.

304.
Shigekiyo T, Uno Y, Tomonari A, et al: Type I congenital plasminogen deficiency is not a risk factor for thrombosis. Thromb Haemost 67:189, 1992.

305.
Prins MH, Hirsh J: A critical review of the evidence supporting a relationship between impaired fibrinolytic activity and venous thromboembolism. Arch Intern Med 151:1721, 1991.

306.
Ohlin AK, Marlar RA: The first mutation identified in the thrombomodulin gene in a 45-year-old man presenting with thromboembolic disease. Blood 85:330, 1995.

307.
Ohlin AK, Marlar RA: Thrombomodulin gene defects in families with thromboembolic disease—a report on four families. Thromb Haemost 81:338, 1999.

308.
Ohlin AK, Norlund L, Marlar RA: Thrombomodulin gene variations and thromboembolic disease. [Review] [26 refs]. Thromb Haemost 78:396, 1997.

309.
Walker ID: Factor V Leiden: should all women be screened prior to commencing the contraceptive pill? [Review] [22 refs]. Blood Rev 13:8, 1999.

310.
Hagstrom JN, Walter J, Bluebond-Langner R, et al: Prevalence of the factor V leiden mutation in children and neonates with thromboembolic disease. J Pediatr 133:777, 1998.

311.
Zenz W, Bodo Z, Plotho J, et al: Factor V Leiden and prothrombin gene G 20210 A variant in children with ischemic stroke. Thromb Haemost 80:763, 1998.

312.
DeVeber G, Monagle P, Chan A, et al: Prothrombotic disorders in infants and children with cerebral thromboembolism. Arch Neurol 55:1539, 1998.

313.
Nowak-Göttl U, Junker R, Hartmeier M, et al: Increased lipoprotein(a) is an important risk factor for venous thromboembolism in childhood. Circulation 100:743, 1999.

314.
Hirsh J, Kearon C, Ginsberg J: Duration of anticoagulant therapy after first episode of venous thrombosis in patients with inherited thrombophilia. Arch Intern Med 157:2174, 1997.

315.
Schulman S, Rhedin AS, Lindmarker P, et al: A comparison of six weeks with six months of oral anticoagulant therapy after a first episode of venous thromboembolism. N Engl J Med 332:1661, 1995.

316.
Kearon C, Gent M, Hirsh J, et al: A comparison of three months of anticoagulation with extended anticoagulation for a first episode of idiopathic venous thromboembolism. N Engl J Med 340:901, 1999.

317.
Lensing AWA, Prandoni P, Prins MH, Büller HR: Deep-vein thrombosis. Lancet 353:479, 1999.

318.
Lechner K, Kyrle PA: Antithrombin III concentrates—are they clinically useful? Thromb Haemost 73:340, 1995.

319.
Bucur SZ, Levy JH, Despotis GJ, Spiess BD, Hillyer CD: Uses of antithrombin III concentrate in congenital and acquired deficiency states. Transfusion 38:481, 1998.

320.
Menache D, O’Malley JP, Schorr JB, et al: Evaluation of the safety, recovery, half-life, and clinical efficacy of antithrombin III (human) in patients with hereditary antithrombin III deficiency. Blood 75:33, 1990.

321.
Vukovich T, Auberger K, Weil J, et al: Replacement therapy for a homozygous protein C deficiency-state using a concentrate of human protein C and S. Br J Haematol 70:435, 1988.

322.
Manco-Johnson M, Nuss R: Protein C concentrate prevents peripartum thrombosis. Am J Hematol 40:69, 1992.

323.
Gerson WT, Dickerman JD, Bovill EG, Golden E: Severe acquired protein C deficiency in purpura fulminans associated with disseminated intravascular coagulation: treatment with protein C concentrate. Pediatrics 91:418, 1993.

324.
Toglia MR, Weg JG: Venous thromboembolism during pregnancy. N Engl J Med 335:108, 1996.

325.
McColl MD, Ramsay JE, Tait RC, et al: Risk factors for pregnancy associated venous thromboembolism. Thromb Haemost 78:1183, 1997.

326.
Greer IA: Thrombosis in pregnancy: maternal and fetal issues. Lancet 353:1258, 1999.

327.
Bauer KA: Management of patients with hereditary defects predisposing to thrombosis including pregnant women. Thromb Haemost 74:94, 1995.

328.
Friederich PW, Sanson BJ, Simioni P, et al: Frequency of pregnancy-related venous thromboembolism in anticoagulant factor-deficient women: implications for prophylaxis. Ann Intern Med 125:955, 1996.

329.
Preston FE, Rosendaal FR, Walker ID, et al: Increased fetal loss in women with heritable thrombophilia. Lancet 348:913, 1996.

330.
Kupferminc MJ, Eldor A, Steinman N, et al: Increased frequency of genetic thrombophilia in women with complications of pregnancy. N Engl J Med 340:9, 1999.

331.
Goddijn-Wessel TAW, Wouters MGAJ, Molen EFvd, et al: Hyperhomocysteinemia: a risk factor for placental abruption or infarction. Eur J Obstet Gynecol Reprod Biol 66:23, 1996.

332.
Powers RW, Evens RW, Majors AK, et al: Plasma homocysteine concentration is increased in preeclampsia and is associated with evidence of endothelial activation. Am J Obstet Gynecol 179:1605, 1998.

333.
Grandone E, Margaglione M, Colaizzo D, et al: Prothrombotic genetic risk factors and the occurrence of gestational hypertension with or without proteinuria. Thromb Haemost 81:349, 1999.

334.
De Vries JIP, Dekker GA, Huijgens PC, et al: Hyperhomocysteinaemia and protein S deficiency in complicated pregnancies. Br J Obstet Gynaecol 104:1248, 1997.

335.
Ridker PM, Miletich JP, Buring JE, et al: Factor V Leiden mutation as a risk factor for recurrent pregnancy loss. Ann Intern Med 128:1000, 1998.

336.
Gardone E, Margaglione M, Colaizzo D, et al: Factor V Leiden is associated with repeated and recurrent unexplained fetal losses. Thromb Haemost 77:822, 1997.

337.
Dizon-Townson D, Meline L, Nelson LM, Varner M, Ward K: Fetal carriers of the factor V Leiden mutation are prone to miscarriage and placental infarction. Am J Obstet Gynecol 177:402, 1997.

338.
Brenner B, Mandel H, Lanir N, Younis J, Rothbart H: Activated protein C resistance can be associated with recurrent fetal loss. Br J Haematol 97:551, 1997.

339.
Quere I, Bellet H, Hoffet M, et al: A woman with five consecutive fetal deaths: case report and retrospective analysis of hyperhomocysteinemia prevalence in 100 consecutive women with recurrent miscarriages. Fertil Steril 69:152, 1998.

340.
Sibai BM: Thrombophilias and adverse outcomes of pregnancy—what should a clinician do? N Engl J Med 340:50, 1999.

341.
Pabinger I, Schneider B, GTH Study Group Natural Inhibitors: Thrombotic risk of women with hereditary antithrombin III-, protein C- and protein S-deficiency taking oral contraceptive medication. Thromb Haemost 71:548, 1994.

342.
Martinelli I, Taioli E, Bucciarelli P, Akhavan S, Mannucci PM: Interaction between the G20210A mutation of the prothrombin gene and oral contraceptive use in deep vein thrombosis. Arterioscler Thromb Vasc Biol 19:700, 1999.

343.
Trauscht-Van Horn JJ, Capeless EL, Easterling TR, Bovill EG: Pregnancy loss and thrombosis with protein C deficiency. Am J Obstet Gynecol 167:968, 1992.

344.
Rosendaal FR: Oral contraceptives and screening for factor V Leiden. Thromb Haemost 75:524, 1996.

345.
Vandenbroucke JP, Van der Meer FJM, Helmerhorst FM, Rosendaal FR: Factor V Leiden: should we screen oral contraceptive users and pregnant women? Br Med J 313:1127, 1996.

346.
Jick H, Jick SS, Gurewich V, Myers MW, Vasilakis C: Risk of idiopathic cardiovascular death and nonfatal venous thromboembolism in women using oral contraceptives with differing progestagen components. Lancet 346:1589, 1995.

347.
Vandenbroucke JP, Helmerhorst FM: Risk of venous thrombosis with hormone-replacement therapy. Lancet 348:972, 1996.

348.
Jick H, Derby LE, Myers MW, Vasilakis C, Newton KM: Risk of hospital admission for idiopathic venous thromboembolism among users of postmenopausal oestrogens. Lancet 348:981, 1996.

349.
Daly E, Vessey MP, Hawkins MN, et al: Risk of venous thromboembolism in users of hormone replacement therapy. Lancet 348:977, 1996.

350.
Douketis J, Ginsberg JS, Holbrook A, et al: A reevaluation of the risk for venous thromboembolism with the use of oral contraceptives and hormone replacement therapy. Arch Intern Med 157:1522, 1997.

351.
Schafer AI: Hypercoagulable states: molecular genetics to clinical practice. Lancet 344:1739, 1994.
Books@Ovid
Copyright © 2001 McGraw-Hill
Ernest Beutler, Marshall A. Lichtman, Barry S. Coller, Thomas J. Kipps, and Uri Seligsohn
Williams Hematology

One comment on “CHAPTER 127 HEREDITARY THROMBOPHILIA

  1. […] CHAPTER 127 HEREDITARY THROMBOPHILIA | Free Medical … Uncategorized by admin […]

Leave a Reply

Fill in your details below or click an icon to log in:

WordPress.com Logo

You are commenting using your WordPress.com account. Log Out / Change )

Twitter picture

You are commenting using your Twitter account. Log Out / Change )

Facebook photo

You are commenting using your Facebook account. Log Out / Change )

Google+ photo

You are commenting using your Google+ account. Log Out / Change )

Connecting to %s

%d bloggers like this: