Novel mutation in coagulation factor VII (Carmel mutation): Identification and characterization.

BACKGROUND: Measurement of factor VII (FVII) activity does not enable prediction of bleeding tendency in individuals with inherited FVII deficiency. OBJECTIVE: To characterize the molecular and functional features of FVII in a family with FVII deficiency and correlate them with the bleeding tendency. PATIENTS/METHODS: We studied 7 family members with very low FVII activity using prothrombin time (PT), activated factor VII (FVIIa), FVII activity level, and thrombin generation. The factor 7 gene was sequenced and the mutation was analyzed by prediction software. RESULTS: The proband has very low FVII activity (0%-4%), with PT ranging between 5% to 18% depending on the tissue factor (TF) origin. Direct sequencing demonstrated a single homozygous nucleotide substitution G > A in exon 6, predicting a novel missense mutation Cys164Tyr. Three members of the family were found to be heterozygous carriers of this mutation. One of them was a compound heterozygote, carrying both the Cys164Tyr and Ala244Val mutation (linked to Arg353Gln polymorphism). Her FVII activity and antigen levels were 3%-7% and 8%, respectively. The other heterozygous carriers demonstrated FVII activity of 41%-54%, FVII antigen of 46%-66%, and FVIIa activity of 30%. FVIIa was undetectable in the homozygous and compound heterozygous subjects. Thrombin generation was normal in the presence of calcium, but no response to TF addition was observed in the homozygous proband, and a reduced response was observed in the compound heterozygous subject. CONCLUSION: The patient homozygous for the "Carmel" mutation has mild clinical manifestations despite very low FVII activity, which correlates with thrombin generation results.

Association of Vitamin B12 Deficiency with Homozygosity of the TT MTHFR C677T Genotype, Hyperhomocysteinemia, and Endothelial Cell Dysfunction.

BACKGROUND: Hyperhomocysteinemia is associated with increased cardiovascular risk, but treatment with folic acid has no effect on outcome in unselected patient populations. OBJECTIVES: To confirm previous observations on the association of homozygosity for the TT MTHFR genotype with B12 deficiency and endothelial dysfunction, and to investigate whether patients with B12 deficiency should be tested for 677MTHFR genotype. METHODS: We enrolled 100 individuals with B12 deficiency, tested them for the MTHFR C677T polymorphism and measured their homocysteine levels. Forearm endothelial function was checked in 23 B12-deficient individuals (13 with TT MTHFR genotype and 10 with CT or CC genotypes). Flow-mediated dilatation (FMD) was tested after short-term treatment with B12 and folic acid in 12 TT MTHFR homozygotes. RESULTS: Frequency of the TT MTHFR genotype was 28/100 (28%), compared with 47/313 (15%) in a previously published cohort of individuals with normal B12 levels (P = 0.005). Mean homocysteine level was 21.2 ± 16 µM among TT homozygotes as compared to 12.3 ± 5.6 µM in individuals with the CC or CT genotype (P = 0.008). FMD was abnormal ( 6%) in 9/13 TT individuals with B12 deficiency (69%), and was still abnormal in 7/12 of those tested 6 weeks after B12 and folic treatment (58%). CONCLUSIONS: Among individuals with B12 deficiency, the frequency of the TT MTHFR genotype was particularly high. The TT polymorphism was associated with endothelial dysfunction even after 6 weeks of treatment with B12 and folic acid. Based on our findings we suggest that B12 deficiency be tested for MTHFR polymorphism in order to identify potential vascular abnormalities and increased cardiovascular risk.

[Laboratory evaluation of lupus anticoagulant in Israel].

Lupus anticoagulants (LAC) are antibodies which are detected by a prolongation of phospholipid-dependent coagulation assays, and are associated with thrombotic events and pregnancy complications in patients with the antiphospholipid syndrome. The antiphospholipid syndrome is defined by arterial or venous thrombosis and/or pregnancy morbidity and by laboratory diagnosis of antiphospholipid antibodies. The laboratory diagnosis is based on LAC and/or anticardiolipin and/or anti-beta2-glycoprotein I antibodies present in plasma, on two or more occasions at least 12 weeks apart. ALthough the presence of LAC correlates best with thrombosis, the Laboratory testing of LAC is not well standardized. In this article, the Laboratory evaluation of LAC will be explained, including the different tests that are recommended by the Israeli Sub-committee of Thrombosis and Hemostasis Laboratories, the possibility to evaluate LAC in patients treated with antithrombotic therapy, and how to report and interpret the results.

Platelet function recovery after cessation of aspirin: preliminary study of volunteers and surgical patients.

BACKGROUND AND OBJECTIVE: Recent evidence indicates that platelet function may recover more rapidly after cessation of aspirin therapy than previously thought. The present study evaluated the effect of aspirin on platelet function using platelet aggregometry in healthy individuals and in aspirin-treated patients scheduled for surgery. METHODS: Platelet aggregation in response to arachidonic acid, epinephrine, and adenosine diphosphate was determined in 14 male volunteers during and after 10 days' aspirin administration (100 mg) and in 58 aspirin-treated patients during intake, on days 3, 4 or 6 after drug cessation, and on day 10 after drug cessation, prior to elective surgery. Urine thromboxane (11-dehydro-thromboxane B2) concentrations were also measured. RESULTS: Platelet aggregation in response to arachidonic acid and epinephrine was significantly decreased in both volunteers and patients during aspirin administration. The aggregation normalized within 3 days of aspirin cessation in the volunteers and within 4-6 days in the patients. Urine concentration of 11-dehydro-thromboxane B2 was about three times lower with aspirin treatment than without, although in two patients concentrations were higher with aspirin. CONCLUSION: Platelet aggregometry with arachidonic acid is a sensitive test for the evaluation of the effects of aspirin on platelet function. In most aspirin-treated patients, platelet function recovers 4 days after drug cessation, although the process is sometimes prolonged. Therefore, the time of aspirin cessation before scheduled surgery should be determined individually.

Endothelial cell dysfunction in women with cardiac syndrome X and MTHFR C677T mutation.

BACKGROUND: The etiology of chest pain with normal epicardial coronary arteries (cardiac syndrome X) seems to be related to endothelial cell dysfunction. Multiple factors are implicated in the pathophysiology, including elevated levels of homocysteine in the blood. Mutations in the MTHFR gene are associated with elevated levels of homocysteine. OBJECTIVES: To test whether abnormal homocysteine metabolism is associated with syndrome X. METHODS: Forty-two women with chest pain, positive stress test and normal coronary arteries (syndrome X) and 100 asymptomatic women (controls) were studied for the C677T mutation. Vitamin B12, folic acid, and plasma levels of homocysteine were also measured. Endothelial cell function was studied in 10 patients with syndrome X and homozygosity for C677T mutation, and in 10 matched healthy controls. Folic acid (5 mg daily) was prescribed to syndrome X patients after initial measurements of ECF. Following 13 weeks of treatment, ECF and blood tests were repeated and compared to baseline measurements. RESULTS: Homozygosity for C677T mutation was doubled in syndrome X vs. control (33%, 14/42 vs. 16%, 16/100, P < 0.02), and homocysteine levels were increased (9.16 +/- 2.4 vs. 8.06 +/- 2.6 pmol/L, P = 0.02). In the 10 homozygous patients, homocysteine levels decreased significantly after treatment with 5 mg/day folic acid (10 +/- 3.3 vs. 5.4 +/- 1.1 micromol/L, P = 0.004). Abnormal baseline ECF improved after treatment with folic acid: flow-mediated dilatation was greater (11.3 +/- 7.9% vs. 0.7 +/- 4.5%, P < 0.002), as was nitroglycerin-mediated dilatation (15.2 +/- 9.0% vs. 5.6 +/- 6.4%, P < 0.003). Frequency of chest pain episodes was significantly reduced after 13 weeks of folic acid treatment. CONCLUSION: Our findings establish the association between the C677Tmutation, endothelial cell dysfunction and cardiac syndrome X, and provide a novel and simple therapy for a subset of patients with syndrome X and homozygosity for the C677T mutation.

Autonomous growth of committed hematopoietic progenitors from peripheral blood of workers exposed to low levels of benzene.

We investigated whether exposure to low levels of benzene (geometric mean of exposures = 0.28-0.41 parts per million) results in perturbations of the hemopoietic system found in patients with myeloproliferative diseases. Erythroid (BFU-E) and granulocyte-monocyte (CFU-GM) colonies from peripheral blood were grown in methylcellulose with and without the addition of cytokines (erythropoietin and granulocyte-stimulating factor). Colony growth from 17 workers exposed to low levels of benzene was compared with 20 healthy control subjects. Exposed workers had significantly increased growth of autonomous BFU-E and unstimulated CFU-GM when compared with the control subjects. Unexposed smokers had increased colony growth without the addition of cytokines. Colony growth was not significantly different between the groups after the addition of cytokines. Workers exposed to low concentrations of benzene and unexposed smokers have increased cytokine-independent hematopoiesis.