Objective To investigate the effect of mechanical stretch force on the apoptosis, proliferation and differentiation of osteoblast culture in vitro. Methods Rat calvarial osteoblasts passage 2 to 4 were used and cell apoptosis was induced by serum starvation after 2 or 4 days of static culture. The osteoblasts were then subjected to 72 hours of cyclic equibiaxial stretch forces using Flexcell 4000TM strain unit. Apoptotic levels of the osteoblasts were quantified by flowcytometer right after the stretch application. The cell number and alkaline phosphatase (ALP) activity were also evaluated. Results After 2 days culture, 6% elongation of stretch force led to a 45% decrease of apoptosis rate, with a 34% increase of cell number. After 4 days culture, 13.6% elongation of stretch force elicited 192% increase of osteoblast apoptosis, while the cell population was decreased by 64%. ALP activity was declined upon the two magnitudes of force loading. Conclusions Stretching force can affect apoptosis in osteoblast culture. This regulative effect was magnitude sensitive controlling the osteoblast activity.