ABSTRACT / PROJECT SUMMARY The understanding of processes that lead to age-related decline in muscle mass and its consequences has been fundamentally limited by imperfect methods of assessing total muscle mass. This has slowed the development of interventions to prevent skeletal muscle loss. The long-term goal of this research is to gain a better understanding of the causes and consequences of low total muscle mass in older adults. The objectives of this project are to measure total muscle mass via the D3-creatine dilution method and determine its association with genetic and non-genetic risk factors, and their relation with falls, injurious falls and fractures in two large, community-based cohorts of older adults. This technique provides a direct and accurate estimate of total muscle mass from a single, fasting urine specimen. The central hypotheses are that lower total muscle mass is associated with novel genetic variants, which when used as instrumental variables in a Mendelian randomization analysis will demonstrate that lower total muscle mass directly increases the risk of incident falls, injurious falls and fractures. Furthermore, association of lifestyle predictors (diet and physical activity) with total muscle mass and with accelerated loss of total muscle mass will be partly mediated by inflammation marker interleukin-6 (IL- 6). Guided by strong preliminary data this hypothesis will be tested by pursuing three specific aims using up to 3,200 participants from two well-characterized cohorts, the Framingham Heart Study (FHS) and the Osteoporotic Fractures in Men (MrOS) Study. Aim 1 will identify genetic variants associated with total muscle mass estimated by D3-creatine dilution in the FHS and MrOS cohorts by performing a genome-wide association study (GWAS).The availability of additional cohorts with D3-creatine and genotyping will bring the sample size to 8,400. Aim 2 will determine the causal relation between total muscle mass estimated by D3-creatine dilution and incident falls, injurious falls and fractures by using SNPs identified in Aim 1 as instrumental variables for total muscle mass in a Mendelian Randomization analysis in the FHS and MrOS cohorts. Aim 3 will determine the cross- sectional associations of physical activity, dietary protein, essential amino acids (EAA), branched chain amino acids (BCAA) and n-3 polyunsaturated fatty acids (n3-PUFA) with baseline total muscle mass in the FHS and MrOS cohorts as well as associations with the change in total muscle mass over an 18 month period in the FHS cohort. To minimize the chance of confounding, Aim 3 will also use a previously published GWAS on accelerometry derived physical activity to perform a Mendelian Randomization analysis of accelerometry derived physical activity and total muscle mass. Lastly, Aim 3b will determine the implied indirect effect of activity and diet on muscle mass along a pathway delineated by IL-6. This study has the potential to transform the field in terms of defining the impact of reduced muscle mass measured using a valid technology, D3-creatine dilution, which could also ultimately serve as an endpoint in drug trials.

PROJECT NARRATIVE and Relevance to Public Health Our proposed work will estimate total body creatine pool size by D3-creatine dilution, a novel and previously validated method that provides a direct and accurate estimate of total muscle mass from a single, fasting urine sample. Our objective is to measure total muscle mass (via the D3-creatine dilution method) in the Framingham Heart Study (FHS) and determine its association with genetic and non-genetic risk factors, and their relation with falls, injurious falls and fractures in two large, community-based cohort of older adults, FHS and the Osteoporotic Fractures in Men (MrOS) Study, thus providing new and important public health insights.