A Longitudinal Study of Exhaled Nitric Oxide in Children
A Longitudinal Study of Exhaled Nitric Oxide in Children
Five percent of children in the UK are prescribed steroid inhalers to control asthma symptoms
 but there is no test to determine whether the dose of steroids is correct. Too much steroid
 treatment has potential side effects and too little may lead to asthma attacks. Exhaled
 nitric oxide (ENO) is a gas present in everyone's breath and may be a useful "meter" for
 asthma control. In children, ENO can be measured easily and quickly, the results are
 available immediately to the doctor or nurse and for these reasons ENO is an attractive
 clinical test.

 Pioneering studies have used ENO to help clinicians treat asthmatic adults and children and
 the results are promising. Breathing tests improved among those where asthma treatment was
 guided by ENO and asthma symptoms were slightly less frequent. These studies all used a
 single ENO value to increase or reduce treatment and study authors have suggested there
 should be a range of ENO values where treatment is neither increased nor reduced; what is not
 known is what these ENO values may be. Elevated NO is associated with a number of factors
 other than asthma, including allergy and pollen exposure. What is not known is how factors
 other than asthma affect ENO measurements over time.

 The proposed study will answer two important questions: What values of ENO indicate that
 steroid treatment should be increased or reduced? And how much does ENO rise and fall
 normally? The investigators will recruit 200 asthmatic and non-asthmatic children. The
 investigators will measure ENO on six occasions over a 12-month period. The investigators
 will measure factors that may affect ENO other than asthma. For the asthmatic children, the
 investigators will also assess asthma control. The investigators' methodology is based on
 several years experience with ENO. The investigators' results will allow ENO to be used to
 monitor asthma.
Our hypotheses are that there will be different ENO values that correlate with loss of/gain
 in asthma control and that ENO variability is related to environmental exposures including
 tobacco smoke and pollen. Our research questions are:

 1. Do determine what values or % change in ENO correlates with gain and loss of asthma
 control

 2. To determine by how ENO measurements vary within an individual over a 12-month period
 among children aged 6-16

 3. To determine whether ENO measurements vary by the same degree for stable asthmatic,
 non-asthmatic atopic and non-asthmatic non-atopic children

 4. To determine whether variability in ENO correlates with environmental exposures
 including tobacco smoke, pollen, domestic animals, mould and ambient NO2

 Background Asthma is a common chronic condition that affects approximately one million
 children in the UK and 5% of all children in the UK are prescribed inhaled corticosteroids
 (ICS) to palliate asthma symptoms. The 2008 British Thoracic Society/Scottish Intercollegiate
 Guidelines Network guidelines for the management of asthma recommends a step-wise approach to
 treatment, i.e. treatment is stepped up and down as symptoms relapse and remit. There is no
 definition of which severity or duration of symptoms should prompt a step-up in treatment.
 The guideline suggests a "trial and error" approach to stepping down treatment in
 symptom-free individuals. In the 21st century, the management of asthma remains based on
 highly subjective clinical decisions and there is a pressing need for a biomarker to guide
 asthma treatment. "Priorities for respiratory research in the UK" recommends that "Rigorous
 evaluation is needed of the use of new technologies for diagnostic and monitoring purposes
 [in asthma], for example exhaled nitric oxide".

 A small number of pioneering studies have sought to use ENO to guide asthma treatment, and
 these have been reviewed recently by the lead applicant. These studies all find ENO-guided
 treatment is associated with improvements in bronchial hyperresponsiveness, spirometry or
 ENO. Although all studies found ENO-guided management was associated with improved symptom
 control, this only achieved significance in one of the four populations studied. The dose of
 ICS was often considerably higher in individuals where treatment was guided by ENO. One
 possible reason for the lack of improved symptom control despite increased ICS dose is that
 all studies used a single cut-off value for ENO to step up or step down ICS treatment; in
 this situation treatment may be stepped up in an individual with minimal/no symptoms but with
 a relatively small increase in ENO. Use of separate ENO values for stepping up and stepping
 down treatment may be more effective, and a recent study has found that changes in paired ENO
 measurements between -40% and +30% from baseline did not indicate gain or loss of asthma
 control. Additionally, approximately 10% of individuals with high ENO do not have airway
 eosinophilia and in this subgroup, who have minor asthma symptoms, ENO does not reduce with
 ICS treatment. Whilst the present literature suggests that ENO is a promising biomarker for
 asthma, the findings to date are conflicting and questions remain. In particular, what is
 still not understood is (i) what values of ENO (or change in ENO) correspond with loss or
 gain in asthma control? (ii) the variability of ENO over time independent of asthma.

 Our study will include rigorous clinical evaluation of participants and will have good
 statistical power (a weakness of some previous studies). In the proposed study we will take
 ENO measurements on six occasions over a 12-month period in well-phenotyped asthmatic,
 non-asthmatic atopic and non-atopic non-asthmatic children. We will describe changes in ENO
 in the context of asthma control. We will use the results to produce a series of cut-offs for
 change in ENO measurements in children; analogous to traffic lights there will be a green
 range (little or no change), amber range (slight increase in ENO), red range (significant
 increased in ENO). This "traffic light" approach to asthma management is analogous to current
 asthma management plans where symptoms and peak flows measurements are used. We will also
 report on changes in ENO in the context of atopy and also environmental exposures including
 tobacco smoke, pollen, domestic animals, mould and ambient nitrogen dioxide (NO2) levels. In
 future, and in collaboration with colleagues in primary care, we will use these ranges to
 design clinical trials where ENO is used to safely monitor asthma. Our data will also be made
 public and could be used by colleagues to design studies where ENO is used to step-up and
 step-down corticosteroid treatment in asthmatic children.

 c) Experimental design and methods Study design Protocol. Two hundred children aged between 6
 and 10 years at enrolment will be recruited from local schools. The initial assessment will
 take place at hospital and the remaining five will take place at school. The following will
 be included in the initial phenotyping assessment: height and weight, a standard respiratory
 questionnaire (ISAAC), Child Asthma Control Test (CACT) for asthmatics, ENO, spirometry, in
 bronchodilator response and skin prick reactivity. Longitudinal measurements of ENO will
 thereafter be taken at two monthly intervals. Asthmatic children will complete a CACT at each
 assessment where changes in asthma medication will also be recorded. The defined primary end
 point is change in ENO between assessments where asthma control changes. The defined
 secondary end point will be variability in ENO over 12 months.

 Eligibility. Children attending schools in Aberdeen city will be eligible. There is a long
 tradition of Aberdonian schools and school children participating with asthma studies and we
 are fortunate to have had the support of Aberdeen City Council, headteachers, class teachers,
 parents and children over the last 40 years. Our experience is that blanket distribution of
 letters to parents via class teachers results in relatively poor enrolment rates. We will
 therefore meet with headteachers and representative of the parent-teachers associations to
 explain the purpose of the study, gain their approval and include articles in school parent
 bulletins before approaching the parents and children. Children aged under 6 years will not
 be included since we have shown that ENO measurements cannot be obtained in the majority of
 this young age-group. Children who are unable to provide an ENO measurement on the initial
 assessment will not be included for future assessments. In the event that few children with
 severe asthma (defined as BTS/SIGN treatment step 3 and 4) are recruited from schools in the
 first year of the study, we will enrich the cohort with severe asthmatics enrolled from
 hospital clinics. At the start of the study, written consent will be obtained from parents
 and verbal assent will be obtained for each assessment from the child.

 Categorisation of participants. Children with asthma will be defined as those with
 affirmative responses to both questions "have you/your child ever had asthma?" and "have
 you/your child had wheezing or whistling in the chest during the last 12 months?" The number
 of non-atopic asthmatic children (ie asthma and negative skin prick reactivity) is
 anticipated to be small; in the final analysis, and only if the number of non-atopic
 asthmatics is sufficient, we will determine whether the variability for this group differs
 for the atopic asthmatics. A stable asthmatic will be defined as an asthmatic child where
 CACT remains ≥19 and treatment does not change over one month.

 Questionnaires. At enrolment a validated questionnaire used for the International Study of
 Allergy and Asthma in Children will be completed. Parents of primary school children (aged≤11
 years) will complete the wheezing, rhinitis and eczema modules in the core questionnaire for
 6-7 year olds. Children attending secondary schools (aged >12 years) will themselves complete
 the wheezing, rhinitis and eczema modules in the core questionnaire for 13-14 year olds.
 Asthmatic children will also complete the CACT. A CACT score of ≥19 will be defined as loss
 of asthma control. A CACT score of <16 will be used to define good asthma control and has a
 79% negative predictive value for physician-defined uncontrolled asthma. Indoor NO2 exposure
 will be ascertained using questions relating to open gas fires and hobs in the home. Current
 cat and dog exposure will be defined as an affirmative response to the questions "Do you
 currently have a cat at home?" and "Do you currently have a dog at home?" Mould exposure will
 be determined from the response to the "Is there visible damp within the home?" if so which
 room?" Home will be defined as the residence where the child spends most of the week.

 ENO measurements. A portable NO analyser (MINO®, Aerocrine, Sweden) will be used to measure
 ENO in accordance with manufacturer's recommendations. The same MINO® will be used for each
 school to eliminate the risk of potential variability for measurements between analysers. Our
 group has already validated the use of the MINO® for use in children. The child will be asked
 to inhale through the analyser and the exhale slowly, using the visual and auditory
 incentives, at 50ml/s for six seconds. In keeping with international guidelines11, we will
 report the mean ENO from three measurements within 10% or two measurements within 5% of each
 other; this approach minimises any inherent variability in ENO measurements due to the
 analyser itself. Measurements will be taken at the same time of day for each child, if
 possible, and this will eliminate any possible diurnal variability in ENO measurements.
 Exhaled measurements will not be taken in children who have a concurrent cold since this may
 affect the ENO result.

 Spirometry. Spirometry will be used to phenotype the study participants. A standard portable
 pneumotachograph (ML3500, MicroLab) will be calibrated and used in accordance with the
 manufacturer's recommendations, international recommendations will be followed. Each child
 will complete three forced expiratory manoeuvres, the highest FEV1 and FVC values will be
 identified, FEV1 and FEV1/FVC ratio will be recorded.

 Bronchodilator response. We will measure FEV1 before and 15 minutes after 200 micorg
 salbutaom (mid/spacer).

 Skin Prick Reactivity. The skin prick test will be used to determine reactivity to cat
 dander, house dust mite, hen's egg, tree pollen, timothy grass, dog dander, cat dander,
 Aspergillus and Alterneria (allergens provided by ALK, Northampton). Positive (histamine
 10mg/ml) and negative controls (0.9% saline) will be used. Atopy will be defined as at least
 one wheal that measures more than 2mm in longest diameter.

 Ambient NO2 concentration. Using the Aberdeen NO2 diffusion tube network we will assess the
 likely average weekly NO2 exposure for each child in the week in which their measurements
 were made. This will be supplemented by an assessment of indoor NO2 exposure from
 questionnaire data.

 Pollen exposure. This will be done by looking at the predictable seasonality of pollen
 production, ie. Tree pollen March/April Grass pollen June/July. Pollen concentrations are not
 measured in Aberdeen and concentrations in Dundee will be used as a frame of reference and
 provided at cost (£500) by the National Pollen and Aerobiology Research Unit. The pollen
 season difference between Aberdeen and Dundee is likely to be very small and insufficient to
 affect the timing of pollen exposures.
Inclusion Criteria:

 -  Child aged 5-10 years

 Exclusion Criteria:

 -  None