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15 November 1995 | Volume 123 Issue 10 | Pages 747-753
Objective: To evaluate the prevalence of moderate hyperhomocysteinemia and inherited thrombophilia disorders (congenital defects of the natural anticoagulant or fibrinolytic mechanisms) in patients with early-onset venous or arterial thromboembolic disease.
Design: Cross-sectional 2-year evaluation of consecutive unrelated patients with a history of venous or arterial occlusive disease occurring before the age of 45 years or at unusual sites, in the absence of local predisposing factors.
Setting: Thrombosis research unit of a community hospital.
Patients: 107 patients with venous thromboembolism (mean age at event, 32.9 ± 11.9 years) and 50 patients with arterial occlusive disease (mean age at event, 31.1 ± 10 years) who did not have acquired coagulation defects, overt cancer, or acquired conditions affecting methionine metabolism.
Measurements: Total plasma homocysteine (fasting levels), antithrombin III, protein C, protein S, activated protein C resistance, plasminogen, and heparin cofactor II were measured at least 3 months after the event. In 87 patients, total plasma homocysteine levels were also measured 8 hours after an oral methionine load was administered (L-methionine, 0.1 g/kg body weight). Ninety-fifth percentiles of the distribution of these variables were established in 60 apparently healthy persons; sex-specific ranges were used for protein S and total plasma homocysteine. Relatives of patients with laboratory abnormalities were studied to confirm inheritance of the defects.
Results: Moderate hyperhomocysteinemia was detected in 13.1% (95% CI, 7.6% to 21.3%) and in 19.2% (CI, 9.0% to 31.9%) of patients with venous or arterial occlusive disease. The prevalence of hyperhomocysteinemia was almost twice as high when based on homocysteine measurements done after oral methionine load as when based on fasting levels. The remaining defects were detected only in patients with venous occlusive disease (activated protein C resistance in 11.2% of patients, protein S or C deficiency in 6.6%, and plasminogen deficiency in 0.9%), with an overall prevalence of 18.7% (CI, 12.1% to 27.6%). Inheritance of hyperhomocysteinemia and of the other defects was confirmed in 26 of the 30 families studied. Event-free survival analysis showed that the relative risk for occlusive disease in patients with moderate hyperhomocysteinemia and other defects was 1.70 times (CI, 1.19 to 2.42; P < 0.01) greater than in patients without defects. After adjustment for the presence of predisposing factors (for example, use of contraceptive drugs, pregnancy, surgery, prolonged bedrest, smoking, mild hypertension or dyslipidemia) and a family history of thrombosis, the age at first event of patients with moderate hyperhomocysteinemia was similar to that of patients with the other defects (26.4 ± 11.2 years compared with 25.2 ± 10.6 years), and the 43 patients with defects were significantly younger at first event than the 114 patients without defects (25.5 ± 11.1 years compared with 31.0 ± 12.3; P < 0.005). Patients with mild hyperhomocysteinemia had a higher rate of recurrence than those without defects (52% compared with 25%; P = 0.01); among the 56 patients who had their first event more than 1 year before observation, the recurrence rate was higher (80% [CI, 51% to 95%]) in patients with defects than in patients without defects (41% [CI, 26% to 57%] P = 0.01).
Conclusions: Moderate hyperhomocysteinemia may have pathogenic significance in premature venous and arterial occlusive disease and should be included among the (inherited) disorders of venous and arterial thrombophilia.
Although the relation of hyperhomocysteinemia to the occurrence of vascular disorders is still unclear, it is of increasing interest that less severe enzymatic abnormalities of the methionine metabolic pathway may predispose to the development of premature vascular disease. Moderate hyperhomocysteinemia, which is characterized by elevations of fasting plasma homocysteine levels as high as 100 µmol/L or by abnormally high levels of plasma homocysteine after methionine load [3], has been observed with increasing frequency in young patients with arterial occlusive conditions such as myocardial infarction and cerebrovascular and peripheral occlusive diseases [4-7]. However, although venous thromboembolism accounts for 50% of the vascular complications of homocystinuria, few data are available on the relation between moderate hyperhomocysteinemia and venous thromboembolic disease [8, 9].
The term thrombophilia refers to an increased tendency to thrombosis sustained by an ongoing stimulus to thrombogenesis or by a defect of the natural anticoagulant or fibrinolytic mechanisms [10]. Genetic factors play an important role in thrombophilia because thrombosis is often familial or is associated with congenital deficiencies of the protein C anticoagulant pathway, antithrombin III, heparin cofactor II, or plasminogen. The main clinical features of patients with inherited thrombophilia are recurrent thrombosis, thrombosis at a young age, idiopathic thrombosis, thrombosis after trivial provocation, and thrombosis in an unusual site [10]. The aim of our study was to evaluate the prevalence of moderate hyperhomocysteinemia and of established disorders of inherited thrombophilia in a series of consecutive patients with early-onset venous or arterial occlusive disease.
We studied consecutive unrelated patients referred to our coagulation service from November 1992 to October 1994. Patients were enrolled in the study if they were younger than 45 years at the time of the occlusive event (n = 142) or if they had thrombosis at unusual sites (central retinal vein or artery, cerebral veins), provided that local risk factors were absent. Exclusion criteria were the presence of overt cancer, liver disease, acquired coagulation abnormalities (for example, lupus anticoagulants, anti-protein S antibodies, and inhibitors of fibrin polymerization), or risk factors known to disturb methionine metabolism (diabetes, hypertension, hyperlipidemia and renal failure). Thirty-two patients referred during the study period were excluded on the basis of these criteria. At the first study visit, patients were asked [by means of a standard questionnaire] about their personal and family history of thrombosis and whether they had predisposing factors for thrombosis. Diagnoses were confirmed by objective methods as follows: 1) Peripheral arterial occlusive disease was confirmed by Doppler ultrasonography and angiography; 2) cerebrovascular disease was confirmed by angiography and computed tomography or nuclear magnetic resonance imaging; 3) arterial or venous retinal occlusion was confirmed by retinal fluorescein angiography; 4) deep venous thrombosis was confirmed by ultrasonography or phlebography; 5) caval or pelvic venous thrombosis was confirmed by color flow Doppler imaging; 6) pulmonary embolism was confirmed by scintigraphy or angiography; and 7) myocardial infarction was confirmed by electrocardiography.
The control group consisted of 60 apparently healthy persons. These controls were either subjects, students, or members of the hospital staff (30 women and 30 men; mean age ±SD, 35.8 ± 11.8 years) who were not taking any medications and who were recruited during the study period. Patients who were receiving antiplatelet drugs were not excluded from the investigation. All patients had blood drawn at least 3 months after the occlusive event if they were not receiving heparin or oral anticoagulant treatment. If patients were receiving anticoagulant treatment, laboratory evaluation was done 6 months after the event and at least 1 week after interruption of oral anticoagulant treatment.
Therapies known to induce variations in homocysteine levels were not given to any patient before testing. All study participants gave informed consent.
Laboratory Studies
Venous blood samples were obtained in vacutainer tubes (Becton Dickinson, Rutherford, New Jersey) containing 0.129 M sodium citrate (9:1, v/v) from an antecubital vein and centrifuged within 30 minutes at 2000 g for 10 minutes at room temperature. Plasma aliquots were snap-frozen with methanol and dry ice and stored at –70 °C. Assays were done within 3 months of blood collection.
Total plasma homocysteine, which corresponds to the sum of free homocysteine, cysteine-homocysteine mixed disulfide, and protein-bound forms, was obtained after cleavage and reduction reactions with sodium borohydride followed by iodoacetic acid treatment. Homocysteine and O-phthaldialdehyde were purchased from Fluka (Buchs, Switzerland). All chemicals and solvents were obtained from BDH (Poole, United Kingdom). High-performance liquid chromatographic analysis was done using the O-phthaldialdehyde precolumn derivatization method [11]. Plasma vitamin B12 and folate levels were determined by using chemiluminometric immunoassays (Ciba Corning ACS, Medfield, Massachusetts).
Because heterozygosity for cystathionine ß-synthase deficiency in patients who have normal fasting plasma homocysteine levels may be detected by an abnormal increment in the total plasma homocysteine level after administration of an oral methionine load [1], all patients older than 14 years were asked to receive oral methionine and have their plasma homocysteine levels measured. Eighty-seven patients (55%) gave their consent. After an overnight fast, a venous blood sample was drawn at 0800 h and L-methionine (0.1 g/kg body weight) was orally administered in 200 mL of fruit juice. Methionine intake was followed by a simple Italian coffee breakfast, and 5 hours later by a regular lunch. In a previous study [12], no diurnal variation in total plasma homocysteine levels in relation to food intake had been observed. The blood specimen was taken 8 hours after methionine intake. In 12 healthy persons, the transient increase in plasma homocysteine level peaked 6 to 8 hours after methionine administration. Increments in plasma homocysteine levels after methionine load were measured in controls.
The activity of antithrombin III (Coatest Antithrombin, Chromogenix, Stockholm, Sweden) and plasminogen (Coatest Plasminogen, Chromogenix) were tested by amydolytic methods adapted to the ACL 300 (Instrumentation Laboratories, Milan, Italy). The activity of heparin cofactor II was tested as previously described [13], and protein C and protein S anticoagulant activity [14, 15] and antigens [14, 16] were measured by previously described methods. The anticoagulant response to activated protein C was tested in an activated partial thromboplastin time-based assay using a commercial reagent (Actin FSL, Baxter, Miami, Florida) and human purified protein C activated by the thrombin-thrombomodulin complex [17]. Because this assay was introduced in our laboratory in March 1993, patients who were enrolled in the study before this date were reinvestigated for activated protein C resistance.
Family Studies
When an abnormality in the laboratory variables was detected, a second blood sample was taken to confirm the observation. In addition, whenever possible, the relatives of the propositus were studied to confirm inheritance of the abnormality. First-degree relatives and siblings of 12 patients with mild hyperhomocysteinemia were studied both while fasting and after methionine intake. For 8 of these families, both parents of the proband were available for study. First-degree relatives and siblings of 11 probands were evaluated for activated protein C resistance and deficiency of protein S, protein C, and plasminogen.
Statistical Analysis
Results are expressed as mean ±SD or as median and range. Hyperhomocysteinemia was defined by a persistent elevation of fasting plasma homocysteine levels or by plasma homocysteine levels after methionine load that exceeded the 95th percentile of the distribution in the control group in the presence of normal folate and vitamin B12 plasma levels [5, 18]. In children, hyperhomocysteinemia was diagnosed when fasting plasma homocysteine levels were greater than the 95th percentile of an age-matched control group (12 µmol/L [12]). Deficiency of protein C, protein S, and plasminogen and resistance to activated protein C were confirmed by the observation of values persistently lower than the 5th percentile distribution of values in the control group. In the control group, all laboratory variables were normally distributed except total plasma homocysteine levels, which were log-transformed to approximate normal distribution. Homocysteine levels are lower in women than in men [12, 19]. Because sex-related differences in plasma homocysteine levels were also observed in our control group (P < 0.0001), separate 95th percentiles were used for men (fasting levels, 19.5 µmol/L; levels after methionine load, 35 µmol/L) and women in the control group (fasting levels, 15.0 µmol/L; levels after methionine load, 30.0 µmol/L) in computing prevalences in the patient groups. Sex-related differences were also observed in the antigen levels (P < 0.001) and in the anticoagulant activity of protein S (P < 0.02), which were lower in women than in men, and therefore separate 95th percentiles for men and women were also used for this variable. There were no sex-related differences in any of the remaining variables.
Hypotheses of no difference in the distribution of continuous variables were tested using analysis of variance and the Student t-test. A single-sample binomial test was used to test whether the prevalence of hyperhomocysteinemia and of the other defects in the patient groups did differ from 5%. Hypotheses of no difference in the distribution of discrete variables among patients with different laboratory abnormalities were tested using the Fisher exact test. Ninety-five percent CIs for discrete variables were calculated according to the binomial distribution. To assess the influence of defects on event-free survival, the Cox proportional-hazards stepwise regression model [20] was used, with sex, predisposing risk factors, and family history of thrombosis included as covariates (BMDP-2L). Analysis of covariance was done to adjust mean age at first event for the presence of predisposing risk factors and family history of thrombosis.
Fifty patients with a history of early-onset arterial occlusive disease (26 women, 24 men; mean age ±SD, 32.8 ± 10.1 years; mean age at event, 31.1 ± 10 years) and 107 patients with history of early-onset venous thromboembolic disease (60 women, 47 men; mean age, 36.4 ± 11.9 years; mean age at event, 32.9 ± 11.9 years) were enrolled in the study. Arterial events included ischemic stroke (23 patients), transient ischemic attacks (7 patients), peripheral arterial occlusive disease (7 patients), central retinal artery occlusion (6 patients), and myocardial infarction (7 patients). Venous thromboembolism included deep venous thrombosis or pulmonary embolism or both (57 patients); caval, portal, or mesenteric venous thrombosis (6 patients); superficial thrombophlebitis (10 patients); central retinal vein occlusion (25 patients); and thrombosis of the cerebral veins (7 patients). Six patients had both arterial and venous thromboembolic disease. A family history of thrombosis was reported by 41% (95% CI, 33% to 49%) of the patients and more often by patients with venous disease than by those with arterial disease (61% [CI, 51% to 70%] compared with 44% [CI, 30% to 59%]; P < 0.05). Predisposing factors for thrombosis (for example, use of contraceptive drugs, pregnancy, surgery, prolonged bedrest, smoking, mild hypertension, or dyslipidemia) were present in 58% of the patients at the time of the first occlusive event, with oral contraceptive use and pregnancy accounting for 56% of all the risk factors recorded. The interval between the first occlusive episode and observation ranged from 0.25 to 27 years (median, 0.25 years); the interval was less than 1 year in 64% of the patients and greater than 5 years in 15%. One or more recurrences had occurred in 14 of the patients with arterial occlusive disease and in 32 of the patients with venous thromboembolic disease (28% and 30%, respectively). Of the 56 patients for whom the interval between the first occlusive event and observation was greater than 1 year, 29 (22 patients with venous thromboembolic disease and 7 patients with arterial occlusive disease) had had one or more recurrences.
Prevalence of Moderate Hyperhomocysteinemia and of the Inherited Thrombophilia Disorders
In the fasting state, the prevalence of moderate hyperhomocysteinemia was 9.3% (CI, 80% to 17.0%; P < 0.05) compared with 5% in patients with venous thromboembolic disease and 8.0% (CI, 3.7% to 22.6%) in patients with arterial occlusive disease. Six additional patients had increased plasma homocysteine levels, but they also had reduced vitamin B12 or folate levels or both.
The percentage of patients with a family history of thrombosis and the recurrence rate were similar in the group who received oral methionine load and in the remaining patients (37% [CI, 27% to 48%] compared with 47% [CI, 35% to 59%] and 33% [CI, 24% to 44%] compared with 26% [CI, 17% to 38%]). Of the 87 patients who agreed to receive the methionine load (58 patients with venous thromboembolic disease and 29 patients with arterial occlusive disease; 47 women and 40 men), 7 of the patients with venous thromboembolic disease (2 with deep venous thrombosis, 2 with cerebral venous thrombosis, 2 with superficial venous thrombosis, and 1 with central retinal venous occlusion) and 3 of the patients with arterial occlusive disease (2 with ischemic stroke, 1 with central retinal artery occlusion)—5 women and 5 men—had fasting homocysteine levels that exceeded the upper limit of the normal range. After methionine load, abnormal elevations of plasma homocysteine levels were observed in 5 additional patients with arterial occlusive disease (2 with myocardial infarction, 1 with transient ischemic attacks, and 2 with ischemic stroke) and in 4 additional patients with venous thromboembolic disease (1 with caval venous thrombosis, 2 with deep venous thrombosis, and 1 with central retinal venous occlusion)—5 women and 4 men. All patients with high fasting plasma homocysteine levels also had abnormally high levels after methionine load. Thus, in both groups of patients, the prevalence of abnormal homocysteine plasma levels after methionine load was twice as high as the prevalence observed in the fasting state (Table 1). Including the 4 patients with fasting hyperhomocysteinemia who refused to receive methionine load (3 patients with deep venous thrombosis, 1 patient with myocardial infarction; 1 woman and 3 men), the resulting overall prevalence of moderate hyperhomocysteinemia was 13.1% (CI, 7.6% to 21.3%; P < 0.001) in patients with venous thromboembolic disease and 18.0% (CI, 9% to 31.9%; P < 0.001) in patients with arterial occlusive disease. ARTICLE
Prevalence of Moderate Hyperhomocysteinemia in Patients with Early-Onset Venous and Arterial Occlusive Disease
Homocysteine, a thiol-containing amino acid, is the branch point between the transsulfuration and the transmethylation pathways of methionine [1]. In the last decade, investigators have studied the role of homocysteine metabolism in the pathogenesis of occlusive vascular disease. Homocystinuria, a disorder caused by homozygous cystathionine ß-synthase (E.C. 4.2.1.22) deficiency or defects in the vitamin B12 – or folate-dependent remethylation of homocysteine to methionine and characterized by fasting plasma homocysteine levels of 200 µmol/L or greater, is frequently associated with severe vascular disease during infancy or childhood. More than half of homocystinuric patients have a first thromboembolic event before the age of 30 years, and premature atherosclerosis is a common development [2].
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Demographic Characteristics of the Patient Sample
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Sex-related differences in homocysteine levels after fasting and after methionine load in patients and controls are reported in Table 2.
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Inheritance of hyperhomocysteinemia was tested in 12 propositi (7 with a family history of thrombosis). It was confirmed by family studies in 8 (6 patients with venous thromboembolic disease and 2 patients with arterial occlusive disease) by the detection of abnormally high total plasma homocysteine levels after fasting or after methionine load in at least one first-degree relative or sibling. In the remaining four families, only one of the proband's parents was available for investigation.
The other inherited defects of thrombophilia were observed only in the patients with venous thromboembolic disease. Poor anticoagulant response to activated protein C was by far the most common abnormality; it was detected in 12 patients (8 women, 4 men), all of whom had deep venous thrombosis or pulmonary embolism or both (11.2% [CI, 6.2% to 19.1%]; P < 0.01). Poor anticoagulant response to activated protein C was observed in 4 additional patients at first examination, but it was not confirmed at repeat testing. Inheritance of the defect was confirmed in all of the 11 families investigated. Among the probands in whom inheritance of the defect was confirmed, none had a combination of activated protein C resistance and mild hyperhomocysteinemia.
Protein S deficiency was observed in five patients (4.7%; CI, 1.7% to 11.1%), protein C deficiency was observed in two patients (1.9%; CI, 0.3% to 7.2%), and plasminogen deficiency was observed in 1 patient (0.9%; CI, 0.01% to 5.8%), with an overall prevalence of 7.5% (CI, 3.5% to 14.5%). The seven patients with either protein C or protein S deficiency (four men and three women) had had deep venous thrombosis, whereas the woman with plasminogen deficiency had central retinal venous occlusion. Inheritance of the defect was confirmed in all 7 families studied.
Demographic Characteristics of Patients with Moderate Hyperhomocysteinemia and with Inherited Thrombophilia Disorders
Predisposing factors for thrombosis were similarly distributed in the 43 patients with defects (25 women, 18 men) (60% [CI, 44% to 77%]: moderate hyperhomocysteinemia, 57% [CI, 35% to 77%]; other defects, 65% [CI, 41% to 84%]) and the 114 patients without defects (57% [CI, 47% to 66%]). A family history of thrombosis was recorded in 49% (CI, 34% to 64%) of the patients with defects (moderate hyperhomocysteinemia, 43% [CI, 23% to 65%]; other defects, 55% [CI, 32% to 76%]) and in 36% (CI, 27% to 46%) of the remaining patients. Among patients with moderate hyperhomocysteinemia, predisposing factors for and family history of thrombosis were similarly distributed in the patients with venous thromboembolic disease and those with arterial occlusive disease. The time elapsed since the first occlusive event was not significantly different in patients with defects and those without defects, ranging from 3 months to 26 years in patients with defects and from 3 months to 27 years in patients without defects. Disease had recurred in 40% (CI, 19% to 65%) of patients with defects and in 25% (CI, 18% to 34%) of patients without defects; higher prevalence of venous and arterial recurrences was observed in patients with mild hyperhomocysteinemia (52% [CI, 31% to 72%]; P < 0.01 compared with patients without defects). Among patients who were observed more than 1 year after their first occlusive event, thrombosis had recurred in 8 of 10 patients with hyperhomocysteinemia (2 patients with arterial occlusive disease and 6 patients with venous thromboembolic disease), in 4 of 5 patients with other defects, and in 17 of 41 patients without defects (5 patients with arterial occlusive disease and 12 patients with venous thromboembolic disease). The combined recurrence rate for patients with defects was significantly higher than that for patients without defects in the entire subgroup of patients (80% [CI, 51% to 95%] compared with 41% [CI, 26% to 57%]; P = 0.01) and in the patients with venous thromboembolic disease only (77% [CI, 46% to 94%] compared with 38% [CI, 22% to 57%]).
The influence of hyperhomocysteinemia and the other inherited defects on event-free survival was analyzed by the Cox regression model, with sex, family history of thrombosis, and predisposing risk factors included as covariates. Event-free survival was negatively influenced by the presence of defects (P < 0.01) and predisposing risk factors (P < 0.01) and by a family history of thrombosis (P < 0.05) in the entire series of patients and in the patients with venous thromboembolic disease only. The relative risk for an occlusive event in patients with defects was 1.7 times greater (CI, 1.19 to 2.42) than in patients without defects, both in the entire series of patients (1.7 [CI, 1.19 to 2.42]) and in patients with venous thromboembolic disease only (1.7 [CI, 1.12 to 2.59]). The relative risks conferred by predisposing risk factors and a family history of thrombosis were 1.25 (CI, 1.13 to 1.38) and 1.17 (CI, 1.02 to 1.35) in the entire series of patients and 1.21 (CI, 1.07 to 1.36) and 1.22 (CI, 1.02 to 1.46) in the patients with venous thromboembolic disease only. No variable had a significant effect on event-free survival in the comparison of patients with moderate hyperhomocysteinemia and patients with the other defects. After adjustment for the covariates, the age at first event was lower in patients with defects than in patients without defects in the entire series of patients (25.5 ± 11.2 years compared with 31.0 ± 12.3 years; P < 0.005) and in the patients with venous thromboembolic disease only (25.5 ± 10.7 years compared with 30.3 ± 12.0 years; P < 0.02). No difference in age at first event was found between patients with hyperhomocysteinemia and patients with the other defects (26.4 ± 11.0 years compared with 25.2 ± 10.7 years).
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When plasma vitamin B12 and folate levels or both were normal, inheritance of the abnormality—confirmed in all eight probands (five of whom had a family history of thrombosis) who had both parents studied—indicated congenital deficiency of cystathionine ß-synthase or of methylenetetrahydrofolate reductase [23] as the main biochemical abnormality responsible for moderate hyperhomocysteinemia. Because the prevalence of heterozygosity for enzyme deficiency in the normal population is between 0.05% and 1.4% [24], these findings confirm the association of isolated moderate hyperhomocysteinemia with premature venous [9] and arterial [1, 3, 4, 7] occlusive disease.
Whereas the prevalence of mild hyperhomocysteinemia was similar in patients with arterial occlusive disease and in those with venous occlusive disease, the hemostatic defects of thrombophilia were observed only in the latter. Altogether, congenital deficiencies of protein S, protein C, and plasminogen were observed in 7.4% of the patients, which is consistent with the data reported by Heijboer and colleagues [25]. Activated protein C resistance, a recently described mechanism for thrombophilia [26], was detected in 11.0% of the patients, and inheritance of the defect was shown in all of the families investigated. In most instances, this disorder is caused by a point mutation in the factor V gene, which affects one of the activated protein C cleavage sites in factor Va [27]. Activated protein C resistance is considered the most common inherited defect predisposing to venous thromboembolism, with reported prevalences ranging from 19% to 64% in different series of patients [28-32].
Establishing a causal association between the presence of a laboratory abnormality and the occurrence of thrombosis is important because correction of the abnormality may reverse the thrombotic risk. For inherited abnormalities, biological plausibility obtained from animal models of thrombosis, strength of the association, and relation established by family studies surveying all family members with and without thrombosis and with and without the abnormality are the main criteria for proving a causal association [10]. In venous occlusive disease, unequivocal evidence for a cause-effect relation has been provided for congenital deficiencies of protein C and protein S [10] and, more recently, for activated protein C resistance [33] but not yet for moderate hyperhomocysteinemia. In vitro studies have shown that hyperhomocysteinemia may induce oxidation of low-density lipoprotein [34] and increase the incorporation of lipoprotein(a) into fibrin [35]. The toxic effect of homocysteine on endothelial cells [36] may lead to increased platelet adhesion [37, 38], induction of tissue factor [39], activation of factor V [40], inhibition of the protein C anticoagulant pathway [41, 42], suppression of heparan sulfate expression [43], and modulation of tissue plasminogen activator activity [44]. Although these procoagulant and antifibrinolytic effects have the potential to predispose hyperhomocysteinemic patients to both arterial and venous occlusive disease, they have been observed primarily when homocysteine concentrations are very high.
Nevertheless, just as there is evidence for a causal association between congenital defects of the protein C anticoagulant pathway (which represent 95% of all of the defects other than hyperhomocysteinemia detected) and venous occlusive disease, the results of our study do suggest that moderate hyperhomocysteinemia contributes to the pathogenesis of venous occlusive disease. The risk for thrombosis conferred by defects of inherited thrombophilia is usually evaluated in cross-sectional studies by analysis of event-free survival curves. Despite the a priori selection of a young patient population, the age at first event (after adjustment for the presence of predisposing risk factors and a family history of thrombosis) of patients with moderate hyperhomocysteinemia was similar to that of patients with defects of the protein C anticoagulant pathway, and these patients were significantly younger at first event than patients without defects. Event-free survival analysis showed that patients with defects had a relative risk that was 1.7 times greater than that of patients without defects. These results, obtained in the entire series of patients and in the subgroup with venous occlusive disease, suggest that the risk for thrombosis conferred by moderate hyperhomocysteinemia is similar to that conferred by the inherited defects for which a causal association has already been proved.
Recurrence of disease is another clinical indicator of risk for thrombosis that can be attributed to an underlying pathogenic defect. Patients with moderate hyperhomocysteinemia had a higher rate of recurrence than patients without defects. Because the likelihood of recurrence depends on the time elapsed between the first event and observation, we also analyzed the rate of recurrence in patients who had their first event more than 1 year before investigation. In the subgroup of patients with venous thromboembolic disease, thrombosis recurred in 75% of patients with hyperhomocysteinemia and in 80% of those with inherited defects of the protein C anticoagulant pathway but in only 38% of the remaining patients. An association between hyperhomocysteinemia (not resulting from vitamin deficiency) and recurrent thrombosis was also found in a recent study observing moderate hyperhomocysteinemia in 25% of 180 patients with a history of recurrent venous thrombosis [45]. These findings point to a causal link between moderate hyperhomocysteinemia and venous and arterial occlusive disease, and further studies are needed to determine whether reversal of hyperhomocysteinemia—such as that achieved by vitamin supplementation [46]—reduces the risk for recurrent occlusive disease.
In our study, moderate hyperhomocysteinemia and inherited thrombophilia disorders were detected in 31.0% and 18.0% of the patients with early-onset venous occlusive disease and arterial occlusive disease, respectively. The prevalence of inherited defects of thrombophilia is greater in patients referred to specialized laboratories for screening studies than in an unselected population of patients with thrombosis [10]. In our series, 36% of the patients unaffected by the disorders investigated had a family history of thrombosis, suggesting the presence of yet unknown inherited causes of thrombophilia. Thus, because mild hyperhomocysteinemia accounted for 54% of the defects identified, measurement of homocysteine should be included in the laboratory evaluation of thrombophilia.
Dr. Calori: Dipartimento di Epidemilogia, Istituto Scientifico H. S. Raffaele, Via Olgettina 60, 20132.
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