Chemical Pathology
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Browsing Chemical Pathology by Author "Davis, William Henry"
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- ItemDevelopment of a high-throughput genotyping assay for detection of functional polymorphisms involved in homocysteine metabolism and the methylation process implicated in multiple sclerosis(Stellenbosch : Stellenbosch University, 2013-12) Davis, William Henry; Janse van Rensburg, Susan; Kotze, Maritha J.; Stellenbosch University. Faculty of Medicine and Health Sciences. Dept. of Pathology. Chemical Pathology.ENGLISH ABSTRACT: The aetiology of multiple sclerosis (MS) remains largely unknown due to the multifactorial nature of disease susceptibility determined by both environmental and genetic factors. Progress has been made in identifying the genetic component of MS , as well as the possible interactions with the environment. In this study single nucleotide polymorphisms (SNPs) in the FTO (rs9939609, Intron 1 T>A), MTR (rs1805087, 2756 A>G), MTRR (rs1801394, 66 A>G), MTHFR (rs1801133, 677 C>T and rs1801131, 1298 A>C) and COMT (rs4680, 472 G>A) genes involved in the methylation metabolic pathway were studied in the context of MS. The overall objective of this study was to elucidate the mechanism underlying raised homocysteine levels in MS patients. The specific aims were 1) to analytically validate high throughput real-time polymerase chain reaction (RT-PCR) genotyping assays for the 6 selected SNPs against direct sequencing as the gold standard for 2) possible integration into a pathology-supported genetic testing strategy aimed at improved clinical management of MS. The study population included a total of 114 unrelated Caucasian MS patients (98 females and 16 males) and 195 unrelated Caucasian control individuals without a diagnosis of neurological disease (128 females and 67 males). A novel finding of this study was that the risk-associated FTO rs9939609 A-allele was associated with raised homocysteine levels (p=0.003) in patients diagnosed with MS, but not in controls. Furthermore, homocysteine levels correlated significantly with bo dy mass index (BMI) (p=0.046) and total cholesterol levels (p=0.048). Both homocysteine (p=0.011) and BMI (p=0.017) were significantly reduced with increasing intake of folate in the diet, while high saturated/trans fat intake correlated significantly with increased BMI (p<0.001). High physical activity correlated with reduced BMI (p<0.006) in the study population, adjusted for age, gender and disease status. Daily intake of at least five fruit and vegetable portions and the COMT rs4680 (472 G>A) AA genotype had a favourable lowering effect on MS disability as assessed by the expanded disability status scale (EDSS) (p=0.035), while smoking increased MS disability significantly (p<0.001). All SNPs studied were found to be in Hardy-Weinberg equilibrium (HWE), with no significant differences detected between patients and control individuals in genotype distribution or allele frequencies. This study has shown for the first time that the underlying disease process of MS moderates the effect of the FTO rs9939609 polymorphism on homocysteine levels , which is consistent with the role of FTO in demethylation and epigenetic changes. Identification of FTO rs9939609 reinforces the importance of adequate folate intake in the diet that can be assessed accurately with use of the Medical History and Lifestyle Questionnaire applied in this study. Finally, the finding that raised homocysteine levels and BMI are significantly influenced by lifestyle factors such as diet and physical activity in our study cohort , offers a solution to counteract the detrimental effects of genetic risk factors contributing to the development of these established vascular risk factors for MS. Combining this information with FTO rs9939609 and COMT rs4680 genotyping may in future translate into a comprehensive pathology supported genetic testing strategy aimed at improved risk management and quality of life in MS patients.