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Objective To explore the suitable level and action time of l7-β estradiol and fluid shear stress (FSS) and their combined effect on the proliferation of rat osteoblasts in Vitro. Methods MC3T3-E1 osteoblasts were adopted after subcultured and different concentrations of 17-β estradiol and FSS values were applied respectively on MC3T3- E1, the suitable level of 17-β estradiol and FSS were selected through MTT and alkaline phosphatase (ALP). Then the two factors at the suitable level were applied simultaneously to MC3T3-E1 to detect the proliferation activity. Results Seventeen-β estradiol(10-18 mol L-1 ) for 5 d and 12x10-5 N FSS for 60 mm exhibited better effects on the proliferation activity than the other groups respectively, and the combined effect of both factors was better than any single-factor treated group. Conclusion Both l7-β estradiol and FSS have a suitable threshold in promoting proliferation of osteoblasts, and two-factor treated group exhibits better effect than any other single-factor treated groups. Therefore 17-β estradiol and FSS have a synergetic action on differentiation and proliferation of osteoblasts. [ABSTRACT FROM AUTHOR]