Hyperhomocysteinemia and other thrombotic risk factors in women with placental vasculopathy.

OBJECTIVE: To investigate coagulation inhibitors and abnormalities of the homocysteine metabolism, which are related to an increased thrombotic risk, as risk factors for placental vasculopathy. DESIGN: A case-control study comparing nonpregnant women with an obstetric history of placental vasculopathy (study group) with nonpregnant women (control group) matched for age and occupation. SETTING: Obstetric outpatient clinic in the University Hospital Nijmegen. SAMPLE: One hundred and one women in the study group and 92 women in a control group. METHODS: Determinations in blood samples of homocysteine concentrations; the occurrence of 677 C-->T mutation in the methylenetetrahydrofolate reductase (MTHFR) gene; protein C activities; activated protein C resistance ratios; concentrations of free protein S antigen; antithrombin III activities; and the occurrence of factor V Leiden mutation. RESULTS: Increased risk for placental vasculopathy was found in the study group with elevated homocysteine (odds ratio 2.28, 95% CI 1.18-4.39), MTHFR mutation (odds ratio 3.29, 95% CI 1.03-10.5), decreased activated protein C resistance ratio (odds ratio 2.46, 95% CI 1.06-5.72) and protein C (odds ratio 2.01, 95% CI 1.11-3.65). Any combination of two risk factors in the same individual resulted in a 3.40 (95% CI 1.80-6.42) higher relative risk for placental vasculopathy; combinations of three risk factors in a 6.83 (95% CI 1.52-30.7) higher risk. CONCLUSIONS: The thrombotic risk factors decreased levels of activated protein C resistance ratios and protein C, elevated homocysteine and the MTHFR 677 C-->T mutation are likely risk factors for placental vasculopathy. Combinations of these risk factors in one individual resulted in synergistic increase of risk.

A common mutation in the 5,10-methylenetetrahydrofolate reductase gene as a new risk factor for placental vasculopathy.

OBJECTIVE: This study was undertaken to investigate whether the cytosine-to-thymine substitution at nucleotide 677 (C677T) in the 5, 10-methylenetetrahydrofolate reductase gene is a risk factor for placental vasculopathy (abruptio placentae or placental infarction with fetal growth restriction). STUDY DESIGN: This case-control study enrolled 165 women with placental vasculopathy and 139 matched control women with normal pregnancy outcomes. Measurements included fasting total plasma homocysteine concentration, serum and red blood cell folate concentrations, serum vitamin B(12) concentration, whole-blood vitamin B(6) concentration, and analysis of the 5, 10-methylenetetrahydrofolate reductase gene C677T mutation. RESULTS: The median total plasma homocysteine concentration was significantly higher in the study group than in the control group (P <.01; odds ratio >97.5th percentile, 4.66; 95% confidence interval, 1.55-14.0). Homozygous genotype for the mutated 5,10-methylenetetrahydrofolate reductase gene was found in 12% of the study group and 5% of the control group (odds ratio, 2.45; 95% confidence interval, 1.00-6.02). CONCLUSIONS: Mild hyperhomocysteinemia was confirmed among women with placental vasculopathy, for which homozygosity for a mutated 5, 10-methylenetetrahydrofolate reductase gene was found to be a new risk factor. The risk of placental vasculopathy probably increases in conditions of low serum folate concentration.

Homocysteine metabolism in endothelial cells of a patient homozygous for cystathionine beta-synthase (CS) deficiency.

Homocystinuria due to cystathionine beta-synthase (CS) deficiency is the most common inborn error of methionine metabolism. Patients with CS-deficiency have an extremely high risk of vascular disease. The underlying mechanism is still unsolved. Dysfunction of endothelial cells could be the trigger in the formation of atherosclerosis and thrombosis. Therefore, differences in cell function were studied between normal and CS-deficient human umbilical endothelial cells (HUVECs). Total homocysteine (tHcy) concentrations in culture media as a measure of homocysteine export increased in all cell lines, including the cell line with CS-deficiency, with constant amounts of approximately 2.5 microM every 24 h. von Willebrand factor (vWF), tissue plasminogen activator (tPA) and plasminogen activator inhibitor (PAI-1) in culture media were used as markers of endothelial function and increased also with progression of culture time. The effects of additions of folate, vitamin B6 and methionine to the culture medium were studied. The homocysteine export and the markers of endothelial function did not differ between the control and the CS-deficient HUVECs under various test conditions. These data show that CS-deficient endothelial cells have normal homocysteine export and normal endothelial cell function. In CS-deficient patients the very high blood levels of homocysteine, probably due to deficient CS function in liver and kidney, seems to be the hazardous factor to endothelial cells, thus promoting atherosclerosis and thrombosis in CS-deficient patients.

Sequence analysis of the coding region of human methionine synthase: relevance to hyperhomocysteinaemia in neural-tube defects and vascular disease.

Elevated homocysteine (Hcy) levels are observed in two apparently unrelated diseases: neural-tube defects (NTD) and premature vascular disease. Defective human methionine synthase (MS) could result in elevated Hcy levels. We sequenced the coding region of MS in 8 hyperhomocysteinaemic patients (4 NTD patients and 4 patients with pregnancies complicated by spiral arterial disease, SAD). We identified only one mutation resulting in an amino acid substitution: an A-->G transition at bp 2756, converting an aspartic acid (D919) into a glycine (G). We screened genomic DNA for the presence of this mutation in 56 NTD patients, 69 mothers of children with NTD, 108 SAD patients and 364 controls. There was no increased prevalence of the GG and AG genotypes in NTD patients, their mothers or SAD patients. The D919G mutation does not seem to be a risk factor for NTD or vascular disease. We then examined the mean Hcy levels for each MS genotype. There was no correlation between GG- or AG-genotype and Hcy levels. The D919G mutation is thus a fairly prevalent, and probably benign polymorphism. This study, though limited, provides no evidence for a major involvement of MS in the aetiology of homocysteine-related diseases such as NTD or vascular disease.

The effect of folic acid on the homocysteine metabolism in human umbilical vein endothelial cells (HUVECs).

Mild hyperhomocysteinaemia is associated with increased risk for vascular disease. We studied homocysteine export from human umbilical vein endothelial cells (HUVECs) by measuring total homocysteine (tHcy) concentrations in the culture medium. Under standard culture conditions tHcy concentrations in the HUVEC culture medium increased by constant amounts after 24, 48 and 72 h [mean = 2.5 (SD +/- 0.7) mumol L-1 homocysteine every 24 h]. As the cells are the only source of homocysteine increase in the culture medium, we designate this as homocysteine export from HUVEC. Folic acid supplementation to the culture medium lowered the homocysteine export in a dose-dependent manner. Methyl-tetrahydrofolate (MeTHF) and folinic acid (a stable precursor of MeTHF) were in this respect about 10 times more effective than folic acid. A 50% reduction in the homocysteine export was seen with 10-30 nmol L-1 MeTHF supplementation; reduction to almost zero was seen with 100-300 nmol L-1 MeTHF. Additions to the culture medium of the other vitamins involved in the homocysteine metabolism, such as vitamin B12, vitamin B6 and flavin adenine dinucleotide, did not show any effect on homocysteine export. Because homocysteine export reflects an imbalance in the homocysteine metabolism, our observations showed a susceptible dependency of this metabolism on folic acid in endothelial cells.