Clinical Trial Details
— Status: Completed
Administrative data
| NCT number |
NCT02590848 |
| Other study ID # |
CP14/00108 |
| Secondary ID |
|
| Status |
Completed |
| Phase |
N/A
|
| First received |
|
| Last updated |
|
| Start date |
April 2016 |
| Est. completion date |
June 5, 2019 |
Study information
| Verified date |
July 2019 |
| Source |
Barcelona Institute for Global Health |
| Contact |
n/a |
| Is FDA regulated |
No |
| Health authority |
|
| Study type |
Interventional
|
Clinical Trial Summary
Background: Adolescence is an important period for brain development as a result of increased
synaptic plasticity. Omega-3 polyunsaturated fatty acids (n-3 PUFAs) are essential nutrients
for brain development and protection against oxidative stress. Walnuts have the highest n-3
PUFA concentrations of all edible plants.
Objective: The investigators hypothesize that walnut intake will increase n-3 PUFA
availability in the body to a level that enhances the development of the brain during
adolescence. The technicians will conduct a long-term (6 months) population-based randomized
controlled trial in teenagers (n=400 in each arm), and determine the effectiveness of the
intervention (30 g of walnut kernels per day, ~1.5g of n-3 PUFA) in enhancing brain
development.
Methods: Fieldwork team will contact teenagers and families in collaboration with 20 high
schools. Families will receive a basic guide on following a healthy diet in order to ensure
implementation and adherence. Brain development outcomes will be measured at baseline and
after the intervention. The use of computerized neuropsychological tests will provide the
precision required to detect even subtle changes in brain development resulting from the
nutritional intervention. Behavioral (socio-emotional) development will be assessed in order
to cover a wider picture of brain development. Blood samples will be collected to measure n-3
PUFA levels before and after the intervention in a randomized subsample from both groups
(control/ intervention). Linear regression models adjusted for baseline neuropsychological
scores will be used to analyze the intervention effect.
Implications: The outcomes of this Project are expected to be a greater understanding of the
role of n-3 PUFA intake (walnut) in brain development. A major goal in public health research
is to develop cost-effective health recommendations to teenagers.
Description:
Adolescence, an important window of opportunity and vulnerability for brain development
Although less well studied, adolescence is also a critical period of brain development. The
brain undergoes important organizational changes, with the re-emergence of
gonadotropin-releasing hormone triggering a cascade of hormone dependent processes. Other
biological processes involve epigenetic factors, which are highly sensitive to the
environment and may therefore make this period of growth more vulnerable to external insults
(Morrison, 2013). Moreover, the prefrontal cortex, which carries out important functions such
as internally guided behaviors (control of emotion), logical thinking, working memory and
organizing skills (executive function), is the last region of the brain to mature
(early-twenties). The synaptic plasticity of the prefrontal cortex is accentuated during
adolescence, a process that involves loss of grey matter density, and an increase in white
matter volume, cerebral blood flow and synaptic pruning. Adolescence is also a time of
refinement of brain connectivity and complex behaviors (Selemon, 2013). It is widely
recognized that the synaptic plasticity of the brain decreases with age (Morrison, 2013;
Selemon, 2013), but this pattern does not seem to follow a linear trend, and adolescence is
an important period during which brain development can be enhanced and protected from
environmental hazards with long-term consequences (everything from air pollution to social
stress). There is a need for greater understanding of the biological pathways through which
the environment affects human brain development, particularly during adolescence, when the
most complex behaviors are refined to adulthood sophistication.
Long-chain polyunsaturated fatty acids, walnuts and brain development Nutrition during
pregnancy, lactation, childhood and adolescence has a fundamental influence on overall
development. Essential fatty acids are long chain polyunsaturated acids (LCPUFAs) that the
body cannot synthesize by itself, and must be obtained through the diet (mainly from oily
fish, seeds and nuts). Three of these fatty acids play an essential role in brain
development: omega-3 (docosahexaenoic (DHA) and eicosapentaenoic (EPA) acids) and omega-6
(arachidonic acid (AA)) (McCann, 2005). LCPUFAs comprise approximately 15 to 30% of the
brain's dry weight (Hallahan, 2005), and are involved in the function and architecture of the
central nervous system throughout various stages of life. Studies show that DHA regulates
neurotransmission systems such as serotonergic, dopaminergic, norepinephrinergic, and
acetylcholinergic systems (Fontani, 2005). Of all edible plants, walnuts have the highest
level of the omega-3 fatty acid, alpha-linolenic acid (ALA). ALA is the precursor for DHA and
EPA, the two most essential fatty acids for the brain (Carey, 2013). Walnuts are also rich in
fiber, vitamins, minerals, and other bioactive substances, such as phenolic antioxidants and
phytosterols (Bao, 2013). Several studies have confirmed an increment in blood n-3 PUFA
levels and improvements in lipid profile after a short-term intervention of a few walnuts per
day (Marangoni, 2007; Maranhao, 2011). While most human studies have shown the cardiovascular
benefits of walnut intake (Poulose, 2014), there is a need to investigate the potential
beneficial effects on brain development, some incipient research on adults show positive
results in cognition, these results are further described in Background section (Pribis,
2012; Sánchez-Villegas, 2011). These findings in adults provide a proof of concept for the
potential brain development benefits of walnuts, which never has been studied among young
teenagers.
This question could be investigated by using a population-based randomized controlled trial
to assess whether walnut intake shows beneficial effects on teenage brain function (using
neuropsychological assessments). It is ethical and feasible to recommend and facilitate
walnut intake (30 g/day; Marangoni, 2007; Maranhao, 2011) over a long period of time (e.g. 12
months; de Ruyter, 2012). A Californian walnut contains ~0.40g of n-3 PUFAs (USDA National
Nutrient Database for Standard Reference, 2012), such that a few walnuts (kernel 30g) per day
would meet recommendations to consume at least 1g of n-3 PUFAs per day (Food and Nutritional
Board, Institute of Medicine of the National Academies, US, 2014). Young Spanish adults
(16-25 years) eat about 10g of nuts per day on average (Salas, 2013). Sea food also contains
high n-3 PUFA concentrations, but sea food interventions are contra-indicated due to their
methylmercury content, an important neurotoxic environmental pollutant (Grandjean, 2014). A
limitation of using walnuts is that intake is less consistent than with a daily pill
supplement of 1g of DHA, although this specific food recommendation is socially
cost-effective and would be complied with by a wider range of people in the global health
context.
Neuropsychological and behavioral assessment, using repeated measurements Recent experience
in incorporating neuropsychological assessment into our cohort studies from INMA (Environment
and Childhood, Guxens, 2012) has resulted in fruitful exchanges of knowledge between
neuropsychologists and epidemiologists. Computerized neuropsychological tests performed in
repeated exams are the most sensitive tool for studying high-order brain function and
development in the general population (Julvez, 2010). Moreover, these tests have been shown
in population-based studies to be very sensitive to the effects of environmental and
nutritional determinants (Julvez, 2013). The investigators will build on this experience to
efficiently select the appropriate range of tests to cover the complex neurofunctions that
rapidly develop during adolescence, such as computer tests of working memory, executive
function, motor speed, visual reasoning, selective and sustained attention and inhibitory
functions (Selemon, 2012). Furthermore, the neuropsychological tests will be complemented by
behavioral rating scales aimed at covering the socio-emotional endpoints that are so
important during early adolescence (Selemon, 2012; Morrison, 2013). The fact to evaluate
other behavioral areas (i.e., social competence and ADHD symptoms) may enrich the
interpretation of the findings, and, possibly it contributes to further understand the
intrinsic mechanisms. One limitation is the risk of losing outcome consistency because of
selecting much dispersed behavioral areas without taking into account the link between them.
The best option is a careful selection of neuropsychological and behavioral tools that
efficiently embrace several areas of teenage brain development. Only a few trials of n-3 PUFA
among adolescents over 11 years have used computer-based neuropsychological tests (Janssen,
2014). A recent large randomized controlled trial among healthy young adults using
computerized cognitive tests found that reaction time latencies and working memory were
improved after 6 months of DHA supplementation (Stonehouse, 2013). A double-blind,
placebo-controlled trial among children with ADHD (7-12 years) found no treatment
associations, but in the observational data, a positive association between DHA levels and
computerized test scores of divided attention (Milte, 2012) was reported. Large trials
measuring neuropsychological functioning using repeated computerized tests within the same
subjects have not yet been carried out among healthy young adolescents, and this is required
to better understand `normal' brain development.
The specific hypotheses are:
1. Walnut intake intervention for six months will enhance neuropsychological development
among teenagers from general population. The investigators expect to find differences
between the two groups (control/walnut) after the intervention.
2. Walnut intake intervention for six months will enhance behavioral (socio-emotional)
development among teenagers from general population. The investigators expect to find
differences between the two groups after the intervention.
3. Walnut intake intervention for six months will increase n-3 PUFA availability in the
body that will be measured in a blood subsample of 200 teenagers at least. The
investigators expect to find differences between the two groups (100/100) after the
intervention.
4. n-3 PUFA levels in blood will be associated with brain development.
Global Objective The global objective of this population-based randomized controlled trial is
to investigate whether the intake of 30 g of walnut kernels (or ~1.5g of n-3 PUFA) per day is
beneficial for brain development among teenagers. The combination of sophisticated tools for
assessing brain functions, such as computerized neuropsychological tests, will provide the
precise data required to detect even subtle changes in brain development resulting from the
effects of the nutritional intervention. Further assessments using behavioural rating scales
will give us a wider look of the potential effects of the intervention.
Specific objectives
1. Determine the effectiveness of long-term intervention (30g of walnut kernels per day for
6 months) in enhancing neuropsychological development. This project will provide a new
understanding of the role of walnut intake (as a surrogate of n-3 PUFA intake) in the
brain development of teenagers. This has an important impact for global health
interventions due to the affordability of such interventions in diverse societal groups.
A major goal in public health research is to develop effective, cost-effective and
feasible health recommendations.
2. Determine the effectiveness of long-term intervention (30g of walnut kernels per day for
6 months) in enhancing behavioural development. In combination with neuropsychological
(or cognitive) development, teenage behavioural (socio-emotional) development is
essential for a wider assessment of brain development.
3. Investigate the effect of the intervention to n-3 PUFA levels in blood and investigate
the association between n-3 PUFA levels and brain development. The importance here
relies on understanding the potential biological pathway between the intervention and
brain development.
Secondary Objectives To assess the association of diet measured by FFQ and adolescent
neuropsychological function.
To assess the association of lifestyle habits assessed by general health questionnaire and
adolescent neuropsychological function.
To assess the association of ambient air pollution estimated by LUR models and adolescent
neuropsychological function.
To assess the association between different behavioral areas during adolescence.
To assess the association between the walnut intervention and biomarkers related to pathways
of oxidative stress and inflammatory system and brain function. These biomarkers are related
to candidate circulating miRNAs from brain and a panel of metabolites from 5 different
classes (acylcarnitines, amino acids, hexoses, phospho-, and sphingolipids and biogenic
amines).
METHODOLOGY WALNUTB scientific approach consists of an ambitious randomized controlled trial
in a large population-based sample. Sample: The trial will include 400 healthy teenagers in
each arm (12-15 year old) from Barcelona city (1,900,000 inhabitants), the participants will
be randomly assigned to two groups (intervention & control) and followed up for 6 months (de
Ruyter, 2012) with two identical series (before and after the intervention) of multimodal
assessments of brain function, and biomarkers of n-3 PUFA. Power calculation for
neuropsychological outcomes: Assigning 400 participants each to the intervention and control
groups, and six primary outcomes were considered, with a correlation between them of 0.25.
The outcomes were considered to have mean 100 and standard deviation 15, as many
neuropsychological scores in the general population. The intervention effect to be detected
was 3 points. We considered a type-I error of 0.05 and corrected calculations for
multiplicity using Benjamini-Hochberg method. A 10% loss of follow-up was assumed.
Additionally, we assumed the final models had an R2 of 20%. With all these considerations,
the resulting study would have 95% power to detect the association with at least one outcome,
90% to detect at least two, 80% to detect at least 3, 70% to detect at least 4, 55% to detect
at least 5 and 31% to detect the association with all six outcomes.
WP 1: The walnut intervention: Design and setting: This trial cannot be blinded due to the
nature of the intervention. The control group will receive no intervention, apart from
general healthy eating recommendations (as in the intervention group) and follow-up
assessments. Recruitment: Nurses will recruit teenagers (n=400 in each arm) in collaboration
with 20 high schools from Barcelona. Implementation and Adherence: The investigators will
perform age- and gender-stratified random sampling to assign subjects to one of the two
groups. All families will receive a basic guide on following a healthy diet in order to
ensure implementation and adherence. Families in the intervention group will receive
additional instructions on how to encourage the adolescent to eat 30 g of walnuts per day
during 6 months, which the investigators will supply for free. The nurses will contact the
families every 3 months for a short checking-interview and to administer a food frequency
questionnaire (FFQ).
WP 2: Fieldwork and neuropsychological assessment: The investigators will validate and
standardize computerized tests to reduce inter-observer variability during assessment and
ensure that data are recorded automatically without errors. Moreover, self-reported and
internationally validated psychometrical scales will be used to further assess adolescent
socio-emotional behaviour. In this project, the investigators will exploit the interaction
between neuropsychologists and epidemiologists to test a range of neuropsychological
functions that cover the various complex behaviours that are under development during
adolescence.
WP 3: Study the intervention effects and the role of omega-3 polyunsaturated fatty acids: In
order to investigate the effects of the intervention, valid measures of the outcomes are
necessary (WP2), as are valid measures of the intermediate factors, in this case plasma n-3
PUFA levels. Blood samples will be collected at baseline and after the intervention,
centrifuged, and plasma samples stored at -80°C until analysis. Statistical Analysis: The
following statistical analyses will be carried out: 1) Assess the effect of the intervention
on the ratio of n-6 PUFA to n-3 PUFA (n6-n3 ratio) using linear regression models adjusted
for baseline levels. 2) Assess the effect of the intervention on brain development, using the
method described in 1) above; 3) Assess the effect of n6-n3 ratio on brain development at
baseline, and of n6-n3 ratio change on brain development change from baseline to follow-up.
4) Assess the proportion of the effect of the intervention on brain development that is
mediated by n6-n3 ratio change. This proportion can be calculated by comparing results from
models with and without the mediator. Analyses 1), 2) and 3) (i.e. those involving the
intervention) will be based on intention-to-treat, i.e. using intervention (no/yes)
regardless of compliance, in order to estimate the effects of the policy (i.e. to encourage
eating 30 g of walnuts per day), rather than the effect of walnut consumption.
Time Frame: The investigators expect to do the recruitment and design the intervention and
perform a pilot during the first 12 months (ongoing since January 2015), and to implement the
intervention and perform follow-up during 12-24 months. The baseline and final assessments
will be completed after 24 months (if the investigators include 2015), the total duration of
fieldwork is two years and a third year will be needed to perform data analyses.
