A successful bone tissue engineering strategy entails producing bone-scaffold construct with adequate mechanical properties. Apart from the mechanical properties of the scaffold itself, the forming bone inside the scaffold also adds to the strength of the construct. In this study, we investigated the role of in vivocyclic loading on mechanical properties of a bone scaffold. We implanted PLA/β-TCP scaffolds in the distal femur of six rats, applied external cyclic loading on the right leg, and kept the left leg as a control. We monitored bone formation at 7 time points over 35 weeks using time-lapsed micro-computed tomography (CT) imaging. The images were then used to construct micro-finite element models of bone-scaffold construct, with which we estimated the stiffness for each sample at all time points.We found that loading increased the stiffness by 60% at 35 weeks. The increase of stiffness was correlated to an increase inbone volume fraction of 18% in the loaded scaffold compared to control scaffold. These changes in volume fraction and related stiffness in the bone scaffold are regulated by two independent processes, bone formation and bone resorption. Using time-lapsedmicro-CT imaging and anewly-developedlongitudinal image registration technique, we observed that mechanical stimulation increases the bone formation rate during 4-10 weeks, and decreases the bone resorption rate during 9-18 weeks post-operatively. For the first time, we report that in vivo cyclic loading increases mechanical properties of scaffold by increasing the bone formation rate and decreasing the bone resorption rate.