published_or_final_version / abstract / toc / Pharmacology / Master / Master of Philosophy

Methylenetetrahydrofolate reductase (MTHFR) synthesizes 5-methyltetrahydrofolate, a methyl donor for conversion of homocysteine to methionine. A common thermolabile variant causes mild MTHFR deficiency, induces mild hyperhomocysteinemia when plasma folate levels are low and increases risk for neural tube defects (NTD) and pregnancy loss. To increase our understanding of Mthfr regulation, the 5' and 3' regions of the mouse cDNA and gene were characterized. These studies revealed two major promoters, an internal coding exon in the 5'UTR, alternative transcriptional and translational start sites and alternative splicing and polyadenylation. These data suggest that Mthfr regulation is likely to be complex. To investigate the role of Mthfr in NTD, several approaches were taken. First, folate and MTHFR co-factor, flavin adenine dinucleotide, were shown to stabilize normal and thermolabile MTHFR during heat inactivation, suggesting that folate might prevent hyperhomocysteinemia in individuals with thermolabile enzyme through protein stabilization. Next, in situ hybridization of neurulating mouse embryos showed that Mthfr is expressed in the forebrain, hindbrain, branchial arches, blood vessels, gut, and importantly, in the ventral part of the neural tube. Mthfr+/- mice were then used as a model of mild deficiency to address the effects of maternal and embryonic Mthfr deficiency on development. When combined with inadequate dietary folate, Mthfr +/- pregnant females showed a two-fold higher rate of pregnancy loss than Mthfr+/+ pregnant females. As well, a percentage of day 10.5 embryos from only the Mthfr+/- pregnant females were underdeveloped by 2 days. These effects were not apparent when dietary folate was sufficient, consistent with a genetic-nutritional interactive effect. Finally, folate metabolism was investigated in an NTD model, the curly-tail (ct) mouse, since the ct defect and Mthfr were mapped in close proximity. However, Mthfr sequence in ct mice was simila

Background and hypothesis. Edentulous patients may be at risk for malnutrition and cardiovascular disease even with well made dentures. Improvements in methods used to assess nutritional status suggest that functional biomarkers such as plasma homocysteine (tHcy), in addition to traditional methods, will provide a better quality assessment. The hypothesis is that there is an association between functional biomarkers of nutritional status and traditional cardiovascular disease risk factors in a community dwelling edentulous elderly population. / Methods. This was a cross-sectional study of the baseline characteristics of a convenient sample of 254 edentulous community dwelling elderly over 65 years. Measurements included anthropometric, body composition, homocysteine, vitamins and relevant blood components. Correlations, multiple regression models and Adjusted Odds Ratios (AOR) for tertiles were used to assess the relationship between vitamins and other parameters with hyperhomocystenemia, defined as THcy value of ≥ 14mmol/L. The independent effect of edentulism on hyperhomocysteinemia was sought using the NHANES III data. / Results. The prevalence of hyperhomocysteinemia was 49.2%. Effect of folate on hyperhomocysteinemia was significant (p = 0.037). Low folate tertile group had a 2.45 times higher odds (CI: 1.23, 4.87) of hyperhomocysteinemia than patients in the higher tertile (p = 0.019). Groups with normal folate did not have higher odds of hyperhomocysteinemia regardless of levels of vitamin B6 or B12. AOR for vitamin B12 tertiles in relation to hyperhomocysteinemia were 2.36 (CI: 1.18, 4.75) and 2.12 (1.07, 4.22) for lower vs. high. A weak inverse relationship existed between tHcy and vitamin C (r= -0.11, p= 0.045). A borderline negative correlation existed between HDL and THcy adjusted for age (r = - 0.12; P = 0.05). Significant correlation existed between THcy and BMI (r = 0.15, P = 0.02), small waist circumference (r = 0.22; p = 0.0004) and waist/hip ratio (r = 0.23, p = 0.0003). In a multivariate analyses, edentulism was associated with hyperhomocysteinemia (p = 0.012). / Conclusions. In this sample, homocysteine levels appeared much higher than one would expect with folate fortification, and were related to several cardiovascular risk factors. Using data from NHANES III, edentulism was found to independently predict hyperhomocysteinemia. The inverse relationship between homocysteine and vitamin C and the effect of folate on hyperhomocysteinemia suggests that increasing dietary intake of fruit and vegetables in edentulous individuals might be beneficial. Furthermore, the latter are at risk of developing hyperhomocysteinemia, a condition associated with cognitive impairment, dementia and coronary artery disease risk.

1. Ištyrus homocisteino koncentraciją kraujo serume ir trombocitų agregaciją kraujo plazmoje su pagrindiniais natūriniais agregantais (adenozino difosfatas, adrenalinas, kolagenas) sergančiųjų ūminiais galvos smegenų kraujotakos sutrikimais, nustatyta koreliacija tarp:  padidėjusios homocisteino koncentracijos ir insulto su dideliu neurologiniu deficitu, išeminės širdies ligos, kreatinino ir C-reaktyviojo baltymo koncentracijų;  vidinės miego arterijos stenozės laipsnio ir homocisteino koncentracijos kraujo serume;  homocisteino koncentracijos kraujo serume ir amžiaus;  padidintos homocisteino koncentracijos ir pasikartojančio insulto, todėl homocisteino koncentracijos lygis gali būti panaudotas kaip prognostinis rodiklis naujam ar kartotiniam ūminiam galvos smegenų kraujotakos sutrikimui išsivystyti. 2. Trombocitų agregacija turtingoje trombocitais plazmoje ūminio išeminio galvos smegenų kraujotakos sutrikimo periode statistiškai patikimai padidėja insultu sergančiųjų grupėje, nepriklausomai nuo susirgimo sunkumo. 3. Profilaktinės aspirino dozės hiperhomocisteinemijos fone dalį ligonių neapsaugo nuo išeminio insulto išsivystymo. Aspirino vartojimas turi veiksmingesnį antiagregacinį poveikį moterims, nei vyrams: moterų plazmoje trombocitų agregacijos intensyvumas buvo žemesnis su visais agonistais, o vyrų trombocitai, atvirkščiai, į juos reagavo viršnorminiu atsaku. 4. Išeminiu insultu sirgusių ligonių kraujyje dažniau randami policitemija, padidėjęs leukocitų... [toliau žr. visą tekstą] / 1. When examining homocysteine concentration in blood serum and platelet aggregation in plasma using nature aggregants (adenosine diphosphate, adrenalin, collagen) in patients with acute cerebrovascular disorders, the positive corelation was found between the following parameters:  the elevated amount of homocysteine and stroke with the high neurological deficit, ischemic heart disease, creatinin and C-reactive protein concentrations;  the degree of internal carotid artery stenosis and homocysteine concentration in blood serum;  homocysteine concentration in blood serum and age;  elevation of homocysteine concentration and recurrent stroke, thus, homocysteine concentration may be a predisposing indicator to the development of new or recurrent acute cerebrovascular disorder. 2. Platelet aggreagation in platelet-rich plasma during the period of acute cerebrovascular disorder statistically significantly increases in stroke group independent upon severity of illness. 3. Preventive aspirin doses in the light of hyperhomocysteinemia do not protect a certain part of patients from ischemic stroke development (among the debatable causes of this outcome may be aspirin resistance). Aspirin intake has a more effective antiaggregate influence on women than men: the intensity of platelet aggregation in women plasma was lower using all agonists, while in men, on the contrary, platelets responded by reaching levels above normal. 4. The patients, who experienced ischemic stroke... [to full text]

Hyperhomocysteinemia is a risk factor for cardiovascular disease and stroke. Nutritional and/or genetic disruptions in homocysteine metabolism can cause hyperhomocysteinemia. Mild methylenetetrahydrofolate reductase (MTHFR) deficiency due to the 677C → T mutation in the MTHFR gene is the most common genetic cause of hyperhomocysteinemia. The 677C → T variant is associated with an increased risk for neural tube defects, pregnancy complications, schizophrenia and Down syndrome, and with a decreased risk for colon cancer and leukemia. This variant is also a potential risk factor for vascular disease. Severe MTHFR deficiency results in homocystinuria, an inborn error of metabolism with neurological and vascular complications. We have generated mice with a knockout of the Mthfr gene. The Mthfr-deficient mice exhibit hyperhomocysteinemia and decreased methylation capacity. The Mthfr+/- mice appear normal, whereas the Mthfr-/- mice are smaller and have reduced survival. Abnormal external granule neuron development associated with increased cell death in the cerebellum was observed in the Mthfr-/- mice. / Evidence for cardiovascular pathology was obtained in several ways. Impaired aortic relaxation response to acetylcholine was seen in the Mthfr +/- mice fed a high methionine diet. Both Mthfr+/- and Mthfr-/- mice fed a low folate high methionine diet developed myocardial fibrosis in the left ventricle. Abnormal lipid deposition in the proximal portion of the aorta was observed in older Mthfr+/- and Mthfr-/- mice. After crossing Mthfr -deficient mice with apoE-null mice, we demonstrated that MTHFR deficiency promoted atherogenesis and its progression in the apoE-null mice. / Gene expression in brain of Mthfr-deficient mice was investigated via microarray analysis. Five genes with altered expression in the brain of Mthfr-/- mouse were validated by RT-PCR. In biochemical studies of human MTHFR, both FAD and folate were shown to stabilize the purified recombinant wild type and mutant MTHFRs from the baculovirus expression system against heat inactivation. The effect of folate appeared to be secondary to that of FAD, and S-adenosylmethionine (SAM) inhibited purified wild type and mutant MTHFRs with similar efficiency. / This dissertation will significantly contribute to our understanding of the role of MTHFR in human disease.

Various hormones have been studied as regards prediction of pregnancy outcome after infertility treatment, but no ideal candidate has been found. Folate and genetic variations in folate metabolism have also been associated with infertility, but it remains unclear how these factors affect IVF pregnancy outcome. It is known that infertility is associated with active folic acid supplement use, but the effect of socioeconomic and lifestyle factors on folic acid supplement use in infertile women has not been well investigated. The overall aim of this work was to obtain information on the prediction of live birth, and to study factors affecting the role of folate and folic acid intake in relation to IVF pregnancy outcome. Infertile women with various infertility diagnoses were studied. Healthy, fertile non-pregnant women were used as controls in three of the studies. Blood samples were taken for assay of eight different hormones, folate and homocysteine, and for genomic DNA extraction. A questionnaire was used to assess background data and use of folic acid supplements. Twenty-four-hour recall interviews were performed for validation of the questionnaire. The studied hormones were not good predictors of live birth. The best predictor was age of the women, together with ovulatory menstrual cycles, and thyroid-stimulating hormone and anti-Müllerian hormone (AMH) status. Well-educated women, high-status employed women, and married and infertile women used the most folic acid supplements. Infertile women had better folate status than fertile women. However, pregnancy outcome after infertility treatment was not dependent on folic acid intake, folate status, genetic variation of 5,10-methylenetetrahydrofolate reductase or socioeconomic status. In conclusion, AMH levels vary less than those of other hormones during the menstrual cycle, and AMH could be used as a predictive marker of live birth together with age and ovulation. Folate might play a minor role in IVF pregnancy outcome, but the importance of folate as regards other health perspectives should not be forgotten.

Background: Impaired homocysteine metabolism is associated with a number of vasculopathies including extracranial aneurysms. We analyzed the possible association of nine genetic variants of homocysteine metabolism with the occurrence of intracranial aneurysms. Methods: Caucasian patients (n = 255) treated at two German hospitals for intracranial aneurysms and local controls (n = 348) were genotyped for the following polymorphisms: methionine synthase (MTR) c.2756A→G, methylenetetrahydrofolate reductase (MTHFR) c.677C→T, MTHFR c.1298A→C, cystathionine β-synthase (CBS) c.844_855ins68, CBS c.833T→C, dihydrofolate reductase (DHFR) c.594 + 59del19bp, glutathione S-transferase Ω-1 (GSTO1) c.428C→A, reduced folate carrier 1 (RFC1) c.80G→A and transcobalamin 2 (Tc2) c.776C→G. Results: The G-allele of the missense polymorphism Tc2 c.777C→G was found to be underrepresented in patients, suggesting that this variant may protect from the formation of cerebral aneurysms [odds ratio per two risk alleles (OR) 0.48; 95% confidence interval (CI) 0.30–0.77; p = 0.002]. We obtained borderline results for the G-allele of RFC1 c.80G→A (OR 1.64; 95% CI 1.01–2.65; p = 0.051) and the insertion allele of DHFR c.594 + 59del19bp (OR 1.61; 95% CI 1.00–2.60; p = 0.059), which were found to be overrepresented in patients. Conclusion: Polymorphisms of homocysteine metabolism are possible risk factors for the formation of intracranial aneurysms. / Dieser Beitrag ist mit Zustimmung des Rechteinhabers aufgrund einer (DFG-geförderten) Allianz- bzw. Nationallizenz frei zugänglich.

Motivation: Cardiovascular disease (CVD) is one of the leading causes of mortality and morbidity in South Africa and worldwide. Various investigations have confirmed the hypothesis that elevated plasma homocysteine (Hcy) levels may be linked to vascular disease, and it has become clear that hyperhomocysteinemia is an independent risk factor for atherosclerosis and atherothrombosis. Extensive research on the influence of vitamin supplementation leading to the lowering of homocysteine levels has been done, but extensive research on the effect of physical activity on high homocysteine levels is lacking. The interaction of vitamin supplementation in combination with physical activity has also not been investigated. If a conditioning exercise programme can demonstrate a lowering effect on elevated homocysteine levels, it will confirm the importance of physical activity as a less expensive alternative for a better lifestyle that can also continue to lower morbidity and mortality rates. Objective: This study examined the effect of a conditioning program, vitamin supplement and a combination of both on Hcy levels in men with coronary heart disease risk factors. Methods: In a randomized, placebo-controlled, blinded crossover study, 84 men matched for physical activity (PA) levels, age and risk factors were randomly assigned to one of 4 groups [A = physical conditioning, 20-30 min; 70-80% (THR), 8 = physical conditioning + supplement, C = supplement (12,5 ug vitamin 812; 200 ug folic acid) or D = control). Groups A, B, and C were crossed over according to the Latin square design. Total plasma homocysteine, maximal oxygen consumption (V02max) and body composition (BMI & Fat %) were measured before and after each 12-week intervention period. A 6-week washout period separated the crossovers. Results: The experimental and control groups presented similar baseline characteristics and the profile analysis of the V02max values and Hcy concentrations indicated positive results (multivariate p-value <0.0001), due to the fact that the four groups repeated measurements, presented different patterns. A phase effect for the V02max values and a phase and interaction effect for the Hcy concentrations were indicated, though all the subjects were requested to maintain their normal daily routine (eating pattern, PA levels and alcohol consumption) for the duration of the study. The lack of compliance to the conditioning programme makes it impossible to draw conclusions for V02max values. The poor compliance lead to a small sample size that eventually leads to less statistical power. Conclusion: This study found that a 12-week conditioning programme had no effect on Hcy concentrations. The results of this study make it impossible, due to poor compliance, to suggest that the effect of increased PA on homocysteine may play an important role in the prevention and treatment of CVD. It is, therefore, recommended that more studies should be conducted to further investigate the effect of PA and vitamin supplements on tHcy levels. / Thesis (M.Sc. (Human Movement Science))--North-West University, Potchefstroom Campus, 2006.

Thesis (M.A. (Human Movement Science))--North-West University, Potchefstroom Campus, 2009.

Thesis (M.A. (Human Movement Science))--North-West University, Potchefstroom Campus, 2009.