Laboratory methods for vitamin D status
Limitations of vitamin D assays are well documented, with significant variability in results between assay methods and laboratories noted in the literature (17). The Vitamin D External Quality Assessment Scheme (DEQAS) has been monitoring the analytical reliability of 25(OH)D assays since 1989 (18). In the Asian-Australian Health study, 25(OH)D levels were tested using liquid chromatography-tandem mass spectrometry (LC-MS/MS) in Canterbury Health Laboratories, New Zealand. Canterbury Health Laboratories participate in DEQAS, and LC-MS/MS is widely considered to be the best assay for accurate and precise measurement of 25(OH)D (19).
Statistical considerations
We used the raw 25(OH)D values to examine the determinants of vitamin D status and the association with cardio-metabolic risk factors, which also fluctuate over
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seasons. Levels of 25(OH)D vary across the year due to changes in UV radiation exposure; to ensure comparability across participants in the evaluation of the association between acculturation and vitamin D status, we removed the seasonal contribution from the measures 25(OH)D values using a sinusoidal regression (20). Methods to measure determinants of vitamin D
This thesis used several different approaches, including both subjective (e.g. sun diaries) and objective (e.g. UV radiation dosimeters) tools, to measure the most important determinant of vitamin D status: sun exposure.
Measurement of Sun exposure
There are two main categories of methods commonly used to estimate sun exposure: geographic estimates and personal estimates. In ecological epidemiological studies geographic data such as latitude of residence and measured ambient UV radiation are used as proxies for personal exposure to UV radiation. However, these geographic parameters cannot reflect the considerable variation in personal sun behaviours. This is especially important in studies with a considerable proportion of immigrant populations where sun exposure behaviour may be very different from the “average”. Here individual-level data are required. In considering which tool to use for measuring sun exposure, the relevant timing of exposure is important. In relation to vitamin D status, previous studies have shown that sun exposure over the previous 6-8 weeks is most important (21). For assessment of sun exposure or vitamin D over the lifetime, in relation to disease risk, longer-term measures of sun exposure, such as silicone skin casts or personal residence calendars may be more appropriate (14). In this thesis, the focus was on short term measurement of sun exposure, in relation to vitamin D status. Various forms of personal dosimeters for UV radiation exposure have been developed. The two most common forms are spore-containing biofilm dosimeters (22) and polysulphone dosimeters (23). More recently, electronic dosimeters have been developed (24). The electronic dosimeters used in our study are comparatively new, and are not yet widely used. They provide the most precise measure of personal exposure to UV radiation. Before the study, dosimeters were calibrated by the Australian Radiation Protection and Nuclear Safety Agency (ARPANSA) and calibration factors specific to each dosimeter were used for
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conversion between raw data and a dose that is measured in standard erythemal doses (SEDs) (4). Data from dosimeter readings are sensitive to weather variation during the short period of study and may not accurately reflect a participant’s sun exposure pattern over a relevant period of weeks. We thus calculated average UV radiation exposure standardised to the ambient UV radiation (data from ARPANSA) for the 28 days prior to blood collection. Standardising in this way to the ambient UV radiation of the previous 28 days reduces the influence on the vitamin D-relevant UV radiation exposure of rainy/cloudy days during the week of dosimeter wearing. Combining dosimeter readings with recent past levels of ambient UVR enables us to evaluate sun exposure over a longer period than usually possible, providing more stable estimates for the period relevant to vitamin D synthesis.
Daily sun diary
Diaries have been used in collecting data on a varied range of health behaviours, including sun exposure and physical activity. Sun diaries are advantageous because they make record of specific events on the day of occurrence (25) and provide data additional to that from dosimeters by including information on clothing, time of day that participants were outdoors, and use of sunscreen. When the diary is used alone (without dosimeter data), the times people report spending outdoors can be converted into personal ambient exposure and into personal dose, provided data are available for the local terrestrial doses of UV radiation (4, 25, 26).
We evaluated the daily average outdoors time and daily physical activity based on the data over the diary days. The daily sun diary has been previously validated against electronic UV radiation dosimeters (4).
Ethics
As the thesis reported on studies that conducted and collected data in Canberra, ACT, Australia, I obtained ethical clearance from the Human Research Ethics Committee (HREC) at the Australian National University. The reference number for the ANU HREC approval is 2012/191. In accordance with ANU HREC requirements, informed consent was sought and obtained from all individuals participating in the studies for which primary data were collected.
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The 45 and Up Study has overarching approval from the University of New South Wales (UNSW) Human Research Ethics Committee (HREC), including use of data from the baseline questionnaire. As this PhD is conducted using the data from the 45 and Up Study linked to administrative hospital data and death data, I also received approval to use data from this study and conduct the linked data analysis from the New South Wales Population and Health Services Research Ethics Committee and the ANU HREC.
Funding
Two sources of funding enabled the conduct of the studies reported in this thesis. The ANU National Centre for Epidemiology and Population Health provided research funds that enabled the data collection and laboratory testing of blood samples for Asian-Australian Health Study. The NSW Cardiovascular Research Network provided funds for the use of the data from the 45 and Up Study and contributed funding for some publications.
Methods
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