Additional file 1 of Examining evidence of time-dependent treatment effects: an illustration using regression methods

Additional file 1: Supplementary Figures and Tables

Data_Sheet_1_Health-related heterogeneity in brain aging and associations with longitudinal change in cognitive function.docx

IntroductionNeuroimaging-based ‘brain age’ can identify individuals with ‘advanced’ or ‘resilient’ brain aging. Brain-predicted age difference (brain-PAD) is predictive of cognitive and physical health outcomes. However, it is unknown how individual health and lifestyle factors may modify the relationship between brain-PAD and future cognitive or functional performance. We aimed to identify health-related subgroups of older individuals with resilient or advanced brain-PAD, and determine if membership in these subgroups is differentially associated with changes in cognition and frailty over three to five years.MethodsBrain-PAD was predicted from T1-weighted images acquired from 326 community-dwelling older adults (73.8 ± 3.6 years, 42.3% female), recruited from the larger ASPREE (ASPirin in Reducing Events in the Elderly) trial. Participants were grouped as having resilient (n=159) or advanced (n=167) brain-PAD, and latent class analysis (LCA) was performed using a set of cognitive, lifestyle, and health measures. We examined associations of class membership with longitudinal change in cognitive function and frailty deficit accumulation index (FI) using linear mixed models adjusted for age, sex and education.ResultsSubgroups of resilient and advanced brain aging were comparable in all characteristics before LCA. Two typically similar latent classes were identified for both subgroups of brain agers: class 1 were characterized by low prevalence of obesity and better physical health and class 2 by poor cardiometabolic, physical and cognitive health. Among resilient brain agers, class 1 was associated with a decrease in cognition, and class 2 with an increase over 5 years, though was a small effect that was equivalent to a 0.04 standard deviation difference per year. No significant class distinctions were evident with FI. For advanced brain agers, there was no evidence of an association between class membership and changes in cognition or FI.ConclusionThese results demonstrate that the relationship between brain age and cognitive trajectories may be influenced by other health-related factors. In particular, people with age-resilient brains had different trajectories of cognitive change depending on their cognitive and physical health status at baseline. Future predictive models of aging outcomes will likely be aided by considering the mediating or synergistic influence of multiple lifestyle and health indices alongside brain age.</p

Physical and lifestyle measures at baseline by age group.

Physical and lifestyle measures at baseline by age group.</p

Clinical measures at baseline by age group.

ASPirin in Reducing Events in the Elderly (ASPREE), a placebo-controlled prevention trial of low dose aspirin, provided the opportunity to establish a biospecimen biobank from initially healthy persons aged 70+ years for future research. The ASPREE Healthy Ageing Biobank (ASPREE Biobank) collected, processed and stored blood and urine samples at -80degC or under nitrogen vapour at two timepoints, three years apart, from a willing subset of Australian ASPREE participants. Written informed consent included separate opt-in questions for biomarker and genetic testing. Fractionated blood and urine were aliquoted into multiple low-volume, barcoded cryotubes for frozen storage within 4 hours of collection. Specially designed and outfitted mobile laboratories provided opportunities for participation by people in regional and rural areas. Detailed, high quality demographic, physiological and clinical data were collected annually through the ASPREE trial. 12,219 participants contributed blood/urine at the first timepoint, 10,617 of these older adults provided 3-year follow-up samples, and an additional 1,712 provided saliva for DNA. The mean participant age was 74 years, 54% were female and 46% lived outside major cities. Despite geographical and logistical challenges, nearly 100% of blood/urine specimens were processed and frozen within 4 hours of collection into >1.4 million aliquots. After a median of 4.7 years, major clinical events among ASPREE Biobank participants included 332 with dementia, 613 with cardiovascular disease events, 1259 with cancer, 357 with major bleeds and 615 had died. The ASPREE Biobank houses and curates a large number of biospecimens collected prior to the clinical manifestations of major disease, and 3-year follow-up samples, all linked to high quality, extensive phenotypic information. This provides the opportunity to identify or validate diagnostic, prognostic and predictive biomarkers, and potentially study biological effectors, of ageing-related diseases or maintenance of older-age good health.</div

Biobank participant questionnaire data.

Table reports the questions (abbreviated from actual) asked of participants at the time of biospecimen collections and the numbers (and percentage of total) of participants who recorded each answer. The options were “yes” or “no” or “unsure” with the latter numbers including those questions not answered. The differences in the questions at year 3 compared with baseline collections were mainly related to use of open-label aspirin and the ASPREE clinical trial medication. (DOCX)</p

Recruitment map of ASPREE Biobank participants.

Legend: Map of Australia with rectangle of enlarged south-eastern area showing the major ASPREE Biobank recruitment sites (named) and include the number of Biobank participants recruited from each site. Processing laboratories are indicated by the dotted oval symbols, as are the regions covered by the mobile biobuses indicated by shaded oval areas. The percentages of ASPREE participants recruited to Biobank in each location, relative to the total number of ASPREE participants in each site (listed alphabetically), were Adelaide, SA (61%), Ballarat, VIC (75%), Bendigo, VIC (79%), Burnie, TAS (42%), Canberra, ACT/Southern NSW (66%), Geelong, VIC (73%), Hobart, TAS (78%), Launceston, TAS (46%), Melbourne, VIC (78%), Mildura, VIC (75%), Traralgon, VIC (77%), Warrnambool VIC/Mt Gambier, SA (83%), Wodonga/Albury, VIC/NSW (72%), Wollongong, NSW (76%). A regional site established in Shepparton, VIC closed after collecting 15 samples.</p

Sample types and numbers stored at baseline and at year 3.

Sample types and numbers stored at baseline and at year 3.</p

Timing of sample transport, processing & storage.

Bar graphs show elapsed time for different stages of sample preparation, categorised into 30 min or 60 min blocks. (A) Sample transport time defined as the elapsed time from sample collection to sample arrival at the processing laboratory (n = 12,219). (B) Sample processing time defined as the elapsed time from sample arrival at the laboratory to storage in a freezer (n = 12,218). (C) Total time from collection to storage in a freezer (n = 12,218). (DOCX)</p

Mobile laboratory used to access regional and rural/remote participants to collect and process biospecimens.

Legend: Vehicle outfitted with laboratory grade benches, refrigerated centrifuge, refrigerator, disposable equipment in secure cupboards and drawers, air conditioning, a bed readily converted to an additional bench, a chair for participants, electric step for easy access, a -80 degC portable cryoshuttle and/or LN2 resin insert cryoshipper, an external powerpoint and an external generator to provide power when not connected to main power.</p

Demographics of ASPREE Biobank participants.

ASPirin in Reducing Events in the Elderly (ASPREE), a placebo-controlled prevention trial of low dose aspirin, provided the opportunity to establish a biospecimen biobank from initially healthy persons aged 70+ years for future research. The ASPREE Healthy Ageing Biobank (ASPREE Biobank) collected, processed and stored blood and urine samples at -80degC or under nitrogen vapour at two timepoints, three years apart, from a willing subset of Australian ASPREE participants. Written informed consent included separate opt-in questions for biomarker and genetic testing. Fractionated blood and urine were aliquoted into multiple low-volume, barcoded cryotubes for frozen storage within 4 hours of collection. Specially designed and outfitted mobile laboratories provided opportunities for participation by people in regional and rural areas. Detailed, high quality demographic, physiological and clinical data were collected annually through the ASPREE trial. 12,219 participants contributed blood/urine at the first timepoint, 10,617 of these older adults provided 3-year follow-up samples, and an additional 1,712 provided saliva for DNA. The mean participant age was 74 years, 54% were female and 46% lived outside major cities. Despite geographical and logistical challenges, nearly 100% of blood/urine specimens were processed and frozen within 4 hours of collection into >1.4 million aliquots. After a median of 4.7 years, major clinical events among ASPREE Biobank participants included 332 with dementia, 613 with cardiovascular disease events, 1259 with cancer, 357 with major bleeds and 615 had died. The ASPREE Biobank houses and curates a large number of biospecimens collected prior to the clinical manifestations of major disease, and 3-year follow-up samples, all linked to high quality, extensive phenotypic information. This provides the opportunity to identify or validate diagnostic, prognostic and predictive biomarkers, and potentially study biological effectors, of ageing-related diseases or maintenance of older-age good health.</div