Exercise mode and intensity/duration are important factors for influencing muscle morphology and function as well as bone. However, it is unknown whether masters cyclists who undergo regular moderate- to high-intensity exercise maintain lower-body skeletal muscle mass (SM) and function and bone health when compared with young adults. The purpose of this study was to compare SM, areal bone mineral density (aBMD), and gait performance between masters cyclists and young adults. Fourteen male masters cyclists (aged 53-71 years) and 13 moderately active young men (aged 20-30 years, exercising less than twice a week) volunteered. The masters cyclists were all training actively (four to five times per week, ∼200 miles per week) for on average the last 17 years (range 7-38 years). Thigh SM was estimated from an ultrasound-derived prediction equation using muscle thickness (MTH). Appendicular lean mass (aLM) and aBMD were also estimated using dual-energy X-ray absorptiometry. There were no significant differences (p<0.05) in thigh SM, anterior and posterior thigh MTH ratio, or aLM between masters cyclists and young men. Maximum straight and zigzag walking times were also similar between groups. Lumbar spine (L1-L4) aBMD was not different between groups, but femoral neck aBMD was lower (p<0.05) in the cyclists than in the young men. Our results suggest that appendicular as well as site-specific thigh muscle loss with aging were not observed in masters cyclists. This maintenance of muscle mass in masters cyclists may preserve walking performance to similar levels as moderately active young adults. However, long-term cycling does not preserve femoral neck aBMD.