Purpose: To examine the association between trajectories of physical activity (PA) over 12 years and epigenetic age acceleration (EAA) in 3600 middle-aged and older adults of the Health and Retirement Study. Methods: Latent variable mixture modeling identified subgroups with similar trajectories of vigorous, moderate, and light PA from 2004 to 2016. Six EAAs, including Horvath’s age acceleration, Hannum’s age acceleration, GrimAge acceleration, PhenoAge acceleration, DunedinPoAm acceleration, and ZhangAA were calculated by regressing epigenetic age on chronological age in 2016. Linear regression models tested associations of PA trajectories with EAAs, controlling for age, sex, race, education, smoking, alcohol consumption, and depression. Results: Five trajectories were identified for each PA type. Moderate and light PA trajectories were stable or slightly changed over time. In contrast, vigorous PA trajectories were either consistently low (27.2%), slightly increased at a low level (14.9%), decreased from moderate to low levels (25.9%), increased to a high level (11.9%), or consistently high (20.1%). Moderate PA trajectories were negatively associated with EAA across six epigenetic clocks (p < .01). Light PA trajectories were not associated with any EAA. Vigorous PA trajectories were associated with slower GrimAge acceleration (p = .004) and DunedinPoAm acceleration (p = .03). Participants that showed consistently high or increasing vigorous PA had slower EAA compared to those with consistently low vigorous PA. Conclusion: Moderate and vigorous, but not light, PA trajectories were associated with slower EAAs.

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