Globalisation has resulted in several anthropometric, reproductive, and lifestyle factors and/or exposures exhibiting secular trends. These same factors and exposures have been implicated in modifying breast cancer risk and have been shown to influence DNA methylation in at least one publication. In this thesis, I conduct a preliminary and exploratory investigation to examine whether socioeconomic status, nutritional factors, weight and height, age at menarche, menopausal status, age at menopause, and estimated lifetime estrogen exposure (ELEE) - all factors displaying secular trends and influencing breast cancer risk - are associated with DNA methylation. The LUminometric Methylation Assay (LUMA) was used to assess global methylation in 376 female subjects from the EPIC cohort and the Infinium HumanMethylation 450 BeadChip, was used to assess genome-wide methylation in 667 male and female subjects from the EPIC Italy subcohort. Genome wide, probe-type specific, CpG locus specific, clustering and single enrichment analyses were performed. Total caloric intake, folate intake, total dietary methyl intake, and age at menarche were found to be associated with global methylation using LUMA. In addition, the 450k BeadChip results suggest that education, household density, highest household occupational position, total caloric intake, folate intake, dietary methyl intake, adherence to the Italian Mediterranean diet, body mass index, age at menopause and ELEE influence methylation patterns, but the power of these investigations did not permit identification of biologically relevant, differentially methylated CpG sites. The present findings indicate that DNA methylation may be one way through which these exposures may become biologically embedded and influence disease risk, and especially breast cancer risk. Mechanisms involving epigenetic changes provide a possible mechanism though which even early life exposures can affect adult disease risk. However, more detailed and more powered epidemiological study designs and tools need to be implemented in order to confirm these associations and also examine causality.

Obesity is a highly heritable and genetically heterogeneous disorder. Reports have shown that the number of obese individuals will project to worldwide high levels in the next couple of years. In this study I tried to identify missing heritability in obesity by looking at extreme phenotypes that are likely to be enriched with rare variants. This will improve the power of their discovery by identifying the loci that may reveal more rare variants. I employed the strategy of using 'extreme' sub-groups of obese patients to identify known and novel loci for obesity in the population of Saudi Arabia. Two different approaches were used. The first was to investigate the contribution of copy number variation (CNV) to obesity in Saudi adults/children that have mental retardation, dysmorphic features and obesity or morbid obesity: for adults, this was defined as body mass index (BMI) ≥ 30 and ≥ 40 kg.m-2 respectively; for children, BMI respectively above the 90th and 97th percentiles. The second was to look for known and new Mendelian forms of obesity in multiplex consanguineous families in children that have extreme obesity (z-BMI above 97th centile). Genome-wide genotyping was performed using the Affymetrix SNP 6.0 Chip platform (AFFYMETRIX, Santa Clara, CA) to discover novel copy number variants using PennCNV. I also used a whole exome sequencing in combination with the detection of runs of homozygosity (ROH) and linkage analysis to screen for known monogenic and possibly novel obesity genes in multiplex Saudi consanguineous families. Two rare obesity causing CNVs were identified from this study. The first was the known chromosome 16p11.2 heterozygous deletion that has been previously shown to be associated with obesity. The second is a novel rare homozygous CNV deletion that encompasses TRIB3 gene that has been previously associated with diabetes. A chromosome 16p11.2 duplication was also found within the known chromosome 16p11.2 deletion. This duplication helped in narrowing down the obesity-causing region within the chromosome 16p11.2 deletion region. Additionally, rare CNV duplication with a size above 500kb were found to be slightly enriched (P=0.017) in cases versus controls. In this study we have also shown the presence of a novel homozygous missense MC4R mutation in combination with a heterozygous missense LPIN3 mutation within the same family. The carriers of MC4R mutation have developed an extreme obesity phenotype on the other hand the carriers of LPIN3 mutation also carried a heterozygous MC4R mutation and were found to be lean suggesting a reverse effect of the LPIN3 gene on the development of obesity. In another family a heterozygous deletion was found in a possibly novel obesity-causing gene CHRNA7. This gene mainly mediates fast signalling transmission at synapses and is found particularly in the lateral hypothalamus that regulates food appetite. In summary, this thesis presents multiple novel CNVs and genes that are found as promising obesity causing variants. It is hoped that the work from this thesis may contribute towards elucidation of the role of CNVs and novel genes in the development of obesity.

Obesity in children is a major public health concern in Malta with prevalence statistics placing 11-, 13- and 15- year old Maltese children at the top of the scale, when compared to other Westernised countries. Obtaining valid information on diets is therefore vital to improve aspects of dietary behaviour in this age group. The main aim of this project was to carry out a feasibility study to improve sugar and water intakes in Maltese school children using a validated dietary assessment tool, REALITYMALTA™. This is a novel internet-based tool originally developed at the Rowett Institute for Nutrition and Health (RINH) and further upgraded, by incorporating local food and drink, for use in Maltese school children aged 9-11 years. Comparative results obtained using the new tool and a one day 24-hr MPR showed non-significant differences between the two tools for sugars, NMES and water, suggesting that it can be used with reasonable confidence to measure these nutrients in Maltese school children. Focus groups carried out in the pre-intervention period guided towards the development of a combined educational and environmental feasibility study carried out for a 12-week period in three local state schools. Educational sessions for parents and children were held, an educational leaflet was distributed, and free bottled water was supplied to the participating classrooms in the intervention schools. Nutrient results comparing the pre-intervention and post-intervention periods using the REALITYMALTA™ tool showed a significant reduction in caloric intake and a decrease, although non-significant, in sugars and NMES consumption. There were no statistically significant changes for water consumption. These encouraging results could provide baseline information for a future full scale trial. Further development work of REALITYMALTA™ is required to ensure the active participation of all those involved and to successfully improve dietary behaviour and reduce obesity in Maltese school children.

No description available.

No description available.

No description available.

No description available.

No description available.

No description available.

No description available.