[The plasma homocysteine, folic acid and vitamin B12 levels in young people with high risk for cardiovascular disease and its relation to metylentetrahydrofolate reductase (MTHFR) gene polymorphism]. / Szív- es errendszeri betegségek vonatkozásában fokozott kockázatú fiatal felnottek homocisztein-, folsav- es B12-vitamin szintjei metilén-tetrahidrofolsav-reduktáz gén polimorfizmusuk függvényében.

INTRODUCTION: Hyperhomocysteinemia is an independent risk factor for cardiovascular morbidity. AIM: The study was designed to evaluate the total homocysteine level and MTHFR C677T polymorphism frequency of 122, healthy, young adults who had increased risk for cardiovascular disease. The serum levels of folic acid and vitamin B12 were also measured. METHODS: Immunoassay, PCR-RFLP methods were used. The statistical analysis was performed by SPSS program. RESULTS: The frequency of the gene-polymorphism was not different significantly in the study group compared to a Hungarian neonatal sample: although in the increased risk group the frequency of homozygous 677TT polymorphism was higher (14.8%), and heterozygosity was smaller (41%). There was no association between MTHFR gene polymorphism and homocysteine levels. A significant negative correlation was found between the folic acid and homocysteine, and between the vitamin B12 and homocysteine levels correlating with the literature. The mean serum total homocysteine level of the group without vitamin supplementation (n: 86) was 9.8 +/- 3.3 micromol/l, while in the other group with vitamin uptake (n: 36) this level was 7.5 +/- 3.0 micromol/l. There was a significant difference between the homocysteine levels of men and women. CONCLUSION: The results of the study correlate with the literature. It would be useful to call the attention of the Hungarian population to the importance of vitamin supply.

Hyperhomocysteinemia, enzyme polymorphism and thiobarbituric Acid reactive system in children with high coronary risk family history.

OBJECTIVES: To investigate both the frequency and the genetic background of hyperhomocysteinemia and the frequency of increased plasma thiobarbituric acid reactive system (TBARS) levels in children and adolescents whose parents had premature coronary heart disease (CHD). METHODS: The study was performed on children and adolescents aged 4-18 years (105 offspring of parents with CHD before age 45 and 74 referents from families without any evidence of premature atherosclerosis). Fasting serum total cholesterol (TC), high density lipoprotein cholesterol (HDL-C), and total triglyceride (TG) levels were measured by enzymatic methods. Low density lipoprotein cholesterol (LDL-C) level was calculated by the Friedewald formula. Plasma total homocysteine (THCy) level was measured by fluorescence polarisation immunoassay. Plasma TBARS level was determined by fluorimetric method. 5,10-methylenetetrahydrofolate reductase (MTHFR) enzyme polymorphism was analyzed by polymerase chain reaction with restriction fragment length polymorphism (PCR-RFLP). RESULTS: Hyperhomocysteinemia was found in 32 cases and in 4 controls. Increased plasma THCy level was found in 10 children and adolescents from 12 cases homozygous for the C677T polymorphism of the MTHFR gene. No similar high frequency was observed in heterozygous subjects. Elevated fasting plasma TBARS levels were found in 38 cases and in 8 controls. The frequency differences were significant (p < 0.01). Allele frequency of the MTHFR polymorphism among cases and controls was similar. Significant correlation (r = 0.53, p < 0.02) was detected between plasma THCy and TBARS levels. One child had high serum TC level, 5 had low serum HDL-C level and all other children had normal serum TC, LDL-C, HDL-C and TG levels from children with hyperhomocysteinemia and/or high plasma TBARS levels. A significant correlation (r = 0.64, p < 0.01) was observed between plasma THCy levels of parents and children in the case group. CONCLUSION: The measurement of plasma THCy and TBARS levels may contribute to the detection of the risk of children and adolescents with high CHD risk family history.

Correlation between T-cell and mast cell activity in patients with chronic urticaria.

BACKGROUND: The presence of autoantibodies reacting with the high affinity IgE receptor (FcepsilonRIalpha) usually indicates a more severe form of chronic urticaria (CU). Previously, we showed an increased lymphocyte reactivity in CU patients; however, the relation between enhanced cellular immunity and the presence of anti-FcepsilonRIalpha-specific autoantibodies has not been investigated. METHODS: Cellular and humoral immune reactivity of 50 CU patients and 28 healthy controls was studied. Serum sIL-2R, neopterin, and tryptase levels were measured to assess T-cell, monocyte/macrophage and mast cell activity, respectively. Helicobacter pylori (HP)-specific IgG antibody, and IgE levels were also tested. Anti-FcepsilonRIalpha-specific autoantibody was determined by Western blotting. In vivo histamine-releasing activity of patients' sera was assessed by the autologous serum skin test (AST). RESULTS: 17/50 CU patients, who both had IgG-type anti-FcepsilonRIalpha-antibodies by Western blotting and a positive AST response, were classified as autoimmune CU. All patients with CU had significantly higher serum sIL-2R and tryptase levels than healthy controls (p = 0.000257, p = 0.000166, respectively), indicating T-cell and mast cell activation. Patients with higher sIL-2R levels also had higher tryptase levels; the strongest correlation was shown in the autoimmune subgroup of patients (rho = 0.688, p = 0.002). There was a tendency towards higher tryptase levels in the autoimmune subgroup, as compared to the nonautoimmune CU patients. While the serum IgE was significantly lower in autoimmune than in nonautoimmune CU (p = 0.000836), there was no significant difference in their sIL-2R, neopterin and HP-specific IgG antibody levels. CU patients with a positive AST response (38/50) had significantly higher tryptase levels (p = 0.0107) when compared to the negative skin test group. CONCLUSIONS: The significant correlation between sIL-2R and tryptase levels in patients with CU indicates that T cell activation is proportional to mast cell degranulation in these patients. The increased level of tryptase in autoimmune CU may suggest more severe disease.

Histamine deficiency in gene-targeted mice strongly reduces antigen-induced airway hyper-responsiveness, eosinophilia and allergen-specific IgE.

Histamine is an important mediator released from activated mast cells provoked by allergen and has a substantial role in the pathophysiology of asthma. However, several lines of evidence indicate that histamine could also have important functions in the regulation of basic cell biological processes. We have used histidine decarboxylase gene-targeted (HDC-KO) mice, lacking histamine, to investigate the effect of histamine deficiency in an animal model of asthma. Our previous investigations revealed that HDC-KO mice had fewer mast cells with reduced granular content and defective degranulation characteristics. Ovalbumin (OVA)-sensitized and challenged HDC-KO mice had significantly reduced airway hyper-responsiveness, lung inflammation, bronchoalveolar lavage eosinophilia, and OVA-specific IgE compared with congenic wild-type littermates treated in the same way. Comparing the expression profiles of cytokines, the levels of IL-1alpha, IL-1beta, IL-4, IL-5, IL-6 and IFN-gamma were significantly lower in the HDC-KO mice in asthmatic late phase, indicating a significantly altered immune response to OVA provocation and challenge. Evaluation of chemokine gene expression revealed that OVA treatment caused elevation of both T(h)1- and T(h)2-type chemokines in wild-type mice, while the chemokine expression was polarized toward a T(h)1 response in HDC-KO mice. According to our results we can suggest that the possible causes of the reduced asthma symptoms in the HDC-KO mice may be the imperfect mast and eosinophil cell system, and an altered immune response to OVA provocation and challenge.

Direct tandem mass spectrometric analysis of amino acids in dried blood spots without chemical derivatization for neonatal screening.

Neonatal screening performed by electrospray tandem mass spectrometry is a powerful technique in clinical diagnostics. In the present paper an alternative to the widely accepted method involving butylation has been developed. In the new method butylation is not required, and multiple reaction monitoring (MRM) was used instead of constant neutral loss scanning. The method was optimized for detection of 23 L-amino acids in their native form. Quantitation was based on isotope-labeled internal standards, calibration curves were linear from 0 to 500 micromol/L, and detection limits were in the range 2-42 micrmol/L. The utility of the present technique is illustrated in the case of one neonate suffering from citrullinaemia.

Severe coagulation factor V deficiency caused by 2 novel frameshift mutations: 2952delT in exon 13 and 5493insG in exon 16 of factor 5 gene.

A male infant with severe bleeding tendency had undetectable factor V activity. Sequence analysis of the proband's DNA revealed one base deletion in exon 13 (2952delT) and one base insertion in exon 16 (5493insG) in heterozygous form. Both mutations introduced a frameshift and a premature stop at codons 930 and 1776, respectively. The proband's father and mother were heterozygous for 2952delT and for 5493insG, respectively. Both mutations would result in the synthesis of truncated proteins lacking complete light chain or its C-terminal part. In the patient's plasma, no factor V light chain was detected by enzyme-linked immunosorbent assay. The N-terminal portion of factor V containing the heavy chain, and the connecting B domain was severely reduced but detectable (1.7%). A small amount of truncated factor V-specific protein with a molecular weight ratio of 236 kd could be immunoprecipitated from the plasma and detected by Western blotting. This protein, factor V(Debrecen), corresponds to the translated product of exon 16 mutant allele.