Fish and seafood are regarded healthy components of a varied diet, particularly important for pregnant women so that they can pass on essential nutrients needed for fetal development of the brain and other organs1. On the other hand, fish may contain environmental pollutants, such as methylmercury, that can put fetal brain development at risk2. A balance has been sought by state and federal agencies in advising the public 3, but certain subpopulations (i.e. pregnant women or women of childbearing age) generally eat less fish than desirable in order to obtain the beneficial effects of fish 1. At the same time, U.S.EPA reports that over 300,000 children born every year exceed the Reference Dose (RfD) for methylmercury and may therefore be at risk at developing adverse effects4.

Tragic pollution episodes have clearly documented that the fetal brain is particularly susceptible to methylmercury toxicity; the adverse effects on the nervous system appear to be irreversible and are much more widespread and serious in children than in adults, especially when exposures have occurred prenatally5,6. While the existence of adverse effects at low exposure levels has been affirmed by national and international regulatory agencies,7-10 certain scientific questions remain unanswered. They relate to the possible occurrence of neurotoxic effects close to the U.S.EPA Reference Dose, the persistence of the cognitive deficits through adolescence, and the possible compensation of adverse effects by essential nutrients present in fish and seafood. The proposed research aims at developing new and more definite information on the benefits and risks associated with seafood intakes at levels relevant to the US general population.

This proposal builds upon the extensive data already collected on a cohort of 14,138 children born between April 1st 1991 and December 31st 1992 in Avon, England, as part of the Avon Longitudinal Study of Parents and Children (ALSPAC) study11. The focus was to identify features of the environment that affect the health and development of children. Detailed information and biological samples were collected at the time of birth and continue to be collected during the course of follow-up11. A10% sample randomly selected was seen at 4, 8 and every 6 months until age 5 years with data including motor and intellectual development. From age 7 years, all children were invited annually for neurobehavioral assessment (including IQ). A pilot project was carried out to measure cord-mercury wet-weight concentrations12, and frozen cords are available from 7,500 subjects to link to the outcome parameters already available.

Because cord tissue has been collected from over half of the children, and because the mercury concentration in the umbilical cord is an excellent biomarker of prenatal exposure levels13, the opportunity exists to assess the developmental mercury exposure level to determine its possible impact on highly relevant neurodevelopmental functions, and the possible interaction with beneficial effects of maternal fish intake.

The specific aims of this project are therefore: to determine if methylmercury-associated cognitive deficits are present at low-level prenatal methylmercury exposures and remain detectable through to adolescence;

• to examine whether intake of fish containing beneficial nutrients affect the same outcomes and counterbalance the methylmercury-associated deficits; and
• to identify other behavioral outcomes that are linked to prenatal methylmercury exposure for possible consideration in future research.

No outline received

No outline received

The Objective

Understanding the basic molecular mechanisms for growth in utero will eventually lead to prevention or therapy for intrauterine growth restriction (IUGR). Our group and others have shown that imprinted genes have an important role in fetal growth both in mouse and man (1). Some of these genes are imprinted in the placenta and their role appears to be linked to successful nutrient transfer, which is vital for normal fetal growth (2). The objective of this proposal is to follow up our study on the maternally expressed PHLDA2 imprinted gene and its promoter region. The expression of PHLDA2 in the placenta has been negatively correlated with growth (3). Recently we have found a deletion polymorphism in the PHLDA2 promoter region that exhibits a trend implicating a role in its growth suppressing function.

Aim:

Using existing ALSPAC data this project will model social, economic and demographic measures as risk factors for having another baby over the following decade. ALSPAC provides an unusually detailed dataset to consider these questions, providing regular time varying data on a range of relevant variables. Accordingly, multilevel event history models will be utilised to gain a uniquely sophisticated longitudinal analysis of the determinants of fertility. The project aims to appraise current theories of reproductive decision making grounded in evolutionary and economic theories of fertility (e.g. Kaplan et al., 2002; Mace, 2007) and further promote integration between these two frameworks.

The project would be two genetic association studies assessing the effects of variations in two genes (BDNF and ACE) on normal variation in cognitive function in ALSPAC. This would be an extension of our previous, successful collaboration in which we have investigated the effects of SNPs in the COMT, 5HTTP and MAOA genes (Barnett et al 2007). Analyses are hypothesis-driven (see below). Outcome measures would be cognitive scores from ages 8 (attention and IQ) and ten years (working memory). Genotype data would be required for the ACE and BDNF genes, which we understand has already been completed in the cohort.

Brain-derived neurotrophic factor (BDNF) is one of the group of neurotrophins which affect cell survival, axonal and dendritic growth and synaptic plasticity among dopaminergic and other neurons (Bath and Lee 2006). The BDNF gene on 11p13 contains a common Val-Met substitution at codon 66. The Met allele is associated with smaller hippocampal volume (Pezawas et al 2004; Szeszko et al 2005) and better memory (Egan et al 2003; Hariri et al 2003; Dempster et al 2005; Tan et al 2005; Ho et al 2006) but not attention, processing speed, language or planning abilities (Ho et al 2006). The relative rarity of Met/Met homozygotes (about 4% of the US population, (Shimizu et al 2004)) means that the cognitive status of Met/Met individuals is effectively unknown. There have been no studies, to our knowledge, of the effect of BDNF genotype on children's cognitive function, despite the obvious importance of BDNF in neurodevelopment.

Angiotension-converting enzyme (ACE) is a central enzyme of the renin-angiotensin system (RAS) and is primarily involved in blood pressure regulation. It is widely distributed throughout the brain, and particularly associated with cerebral blood vessels, and astrocytes of the periventricular nuclei. Raised RAS activity through elevated ACE is thought to impair cellular metabolism (Williams et al 2000) and amplify free radical and pro-inflammatory responses (Harding et al 2005). ACE may therefore play a role in neurodevelopmental progress and subsequent cognitive development (Harding et al 2005) as well as late-life cognitive decline (Amouyel et al 1996). This hypothesis is supported by demonstrations that ACE inhibitors improve cognitive function (Amenta et al 2002; Tzourio et al 2003).

The ACE gene on 17p23 contains a 287bp insertion/deletion polymorphism in intron 16. Both tissue and circulating ACE levels are higher in individuals with two copies of the deletion allele (Tiret et al 1992). The insertion allele appears to confer a slightly increased risk for Alzheimer's disease (Narain et al 2000) while the deletion allele has been repeatedly associated with increased risk for age-related cognitive impairment and dementia (Amouyel et al 1996; Palumbo et al 1999; Bartres-Faz et al 2000; Richard et al 2000; Stewart et al 2004). Deletion allele carriers are reported to show greater cognitive impairment in moderate and severe traumatic brain injury (Ariza et al 2006), less improvement after shunt surgery in normal pressure hydrocephalus (del Mar Matarin et al 2005), and reduced cognitive impairment in alcoholism (Bartres-Faz et al 2002). However three epidemiological studies have found no associations between ACE genotype and normal cognitive ageing (Visscher et al 2003, Frederiksen et al 2003, Yip et al 2002). Two studies have investigated the effects of ACE genotype on cognitive function in children (Gao et al 2006, Harding et al 2005). Both studies were negative, however both were limited by sample size and by the relatively crude cognitive measure they employed.

The overall aim is to investigate the role that language and literacy play in the educational and social functioning of young people as they enter adulthood and to identify factors and processes that are associated with positive and negative outcomes. Four work streams will focus on particular objectives as follows: 1. To identify risk and protective factors associated with language and literacy outcomes. In particular we will identify: how speech, language, literacy and cognitive variables cluster and change during childhood; to what extent do language learning and literacy difficulties persist to school leaving age predictors of language and literacy performance at school leaving age; mechanisms and characteristics by which social class impacts upon children's language and literacy during preschool, primary school and at school leaving. 2. To identify the relationship between young people's current and past performance in language and literacy and their decisions about social activities, further education and future employment as they leave school. We aim to identify the patterns of social, educational and employment outcomes at school leaving age that are related to different language learning and literacy trajectories; social conditions that predict young people's perceptions of their own functional language and literacy performance and outcomes. 3. To identify the prevalence of conduct and behaviour disorders, risk taking and criminal offending behaviours in relation to young people's current and past performance in language and literacy. In particular we will identify any subgroups of speech, language and literacy deficits that are associated with raised occurrence of psychopathology and /or conduct disorders and at what age the mental health impact become apparent. 4. To identify the current and predicted social costs and return on investment to society as a whole from support of children's language and literacy skills development through professional involvement of speech and language services. Particular objectives include: What is the relationship between GCSE performance outcomes and the level of support in childhood for language and literacy skills development and how does this translate to economic resource savings in the UK economy? To identify whether children with language and literacy difficulties who have received support in childhood through the professional involvement of speech and language services choose higher skill occupation destinations than who have not received such support and, if so, what are the social costs incurred? To identify the economic benefits in monetary terms of professional involvement in language and literacy skills support for emotionally and socially vulnerable young people. Finally it is also an objective of the project to create an archive of the language samples from 5, 8 and from the young people at 17+.

This study aims to use the ALSPAC data to examine the association between risk of a 'high depressive symptom score' and levels of physical activity during pregnancy. The ALSPAC data is ideally suited to address this question as it has an appropriate measure of exposure, at more than one time point, an outcome measure that has been accepted in previous publications and a longitudinal, prospective design.

Appetite is tightly controlled by internal physiological mechanisms in younger children but a decline in the degree of innate appetite control is observed with age 1-3. It has been postulated that external factors, such as palatability of food, begin to influence internal controls and ultimately override satiety signals 4. The energy density of food has been shown to increase energy intake and short term weight gain in experimental studies 5. Energy density is closely related to the palatability of a food and may disrupt appetite control by over stimulating sensory centres in the brain, which process information on texture and taste of food. An association between fat mass and the FTO gene has recently been identified 6. No functional information about the FTO gene is currently available. Human tissue panel expression studies showed that the FTO gene was most highly expressed in the brain, specifically the cerebral cortex, which may implicate a role in the sensory centres, located in the prefrontal cortex.

I hypothesise that the relationship between dietary energy density and obesity, which we have shown to vary by age in the ALSPAC sample 7, may also vary by the FTO genotype, such that children carrying the high-risk allele A will have a stronger relationship between energy density and obesity than those without. Since the relationship between the FTO gene and BMI z score also appeared to get stronger with age this may be explained by age related changes in appetite control in response to energy dense foods. So I would hypothesise further that the interaction between FTO and energy density on obesity may also get stronger with age.

Data requirements:

This analysis would require access to existing data at ALSPAC from direct assessment and biological samples including:

• Dietary energy density from diet diaries at age 10 years
• Genotype for FTO gene
• Fat mass from DXA at age 13 years
• Height measured at age 13 years (In order to adjust fat mass for height)

The proposed work would be split between MRC HNR and ALSPAC. Laura Johnson, at MRC HNR, would generate the dietary energy density variable from the dietary data at age 10 years. This variable would be checked and cleaned by Laura and sent to ALSPAC. Nick Timpson, at ALSPAC, would perform the analysis of association between the FTO genotype, dietary energy density and fat mass. Laura Johnson would then write up the results of the analysis in a paper.

The plan of analysis to test the hypothesis would be to initially inspect the data for potential associations using cross tabs. If there is evidence for an association then a linear regression should be performed with fat mass (adjusted for height) as the dependent variable and the FTO gene, dietary energy density (DED) and FTO*DED interaction term as independent variables.

If there is evidence for an interaction then a linear regression analysis of fat mass and DED stratified by genotype would be done in order to test the hypothesis that the relationship between DED and obesity is stronger in children with the A allele. Further analysis could include a logistic regression with obesity (the top quintile of fat mass adjusted for height) as the dependent variable and the FTO gene, dietary energy density (DED) and FTO*DED interaction term as independent variables. Followed by stratification by genotype if evidence of an interaction is found.

Calculations in Quanto (http://hydra.usc.edu/gxe/) indicate that using a sample of 5000 children there is a power of 0.99 to detect a true positive effect, using the following parameters to estimate power:

For the continuous outcome fat mass index (kg/m5.8);

Risk allele (A) frequency = 0.39

Mode of inheritance = additive

Dietary energy density standard deviation = 1.5 kJ/g

Fat Mass Index mean = 1.21 kg/m5.8, standard deviation = 0.63

Main effect sizes

Gene B = 0.1 kg/m5.8 per A allele

Diet B = 0.05 kg/m5.8 per 1kJ/g

Gene*Diet B = 0.05 to 0.15

N = 5000

Power for all G*E effect sizes between 0.05 and 1.5 was 0.99, meaning that it should be possible to detect an interaction effect as small as 0.05 kg/m5.8