Inflammatory processes are putative mechanisms underlying the cardioprotective effects of physical activity. An inverse association between physical activity and inflammation has been demonstrated, but no long-term prospective data are available. We therefore examined the association between physical activity and inflammatory markers over a 10-year follow-up period. Participants were 4289 men and women (mean age, 49.2 years) from the Whitehall II cohort study. Self-reported physical activity and inflammatory markers (serum high-sensitivity C-reactive protein and interleukin-6) were measured at baseline (1991) and follow-up (2002). Forty-nine percent of the participants adhered to standard physical activity recommendations for cardiovascular health (2.5 h/wk moderate to vigorous physical activity) across all assessments. Physically active participants at baseline had lower C-reactive protein and interleukin-6 levels, and this difference remained stable over time. Compared with participants who rarely adhered to physical activity guidelines over the 10-year follow-up, the high-adherence group displayed lower log(e) C-reactive protein (β=-0.07; 95% confidence interval, -0.12 to -0.02) and log(e) interleukin-6 (β=-0.07; 95% confidence interval, -0.10 to -0.03) at follow-up after adjustment for a range of covariates. Compared with participants who remained stable, those who reported an increase in physical activity of at least 2.5 h/wk displayed lower log(e) C-reactive protein (β coefficient=-0.05; 95% confidence interval, -0.10 to -0.001) and log(e) interleukin-6 (β coefficient=-0.06; 95% confidence interval, -0.09 to -0.03) at follow-up. Regular physical activity is associated with lower markers of inflammation over 10 years of follow-up and thus may be important in preventing the proinflammatory state seen with aging.