Objective To explore the effect of hypergravity on morphology and osteogenesis function of preosteoblast MC3T3-E1 cells. Methods The cultured MC3T3-E1 cells under hypergravity by different loading forces were divided into five groups, including control group, 5 g group, 10 g group, 15 g group and 20 g group. The experimental groups were loaded for 30 min each time in the three successive days, and the control group was synchronously exposed to the same surrounding except for difference in g-value. The morphology of cytoskeletal protein was observed by phalloidin staining, The alkaline phosphatase (ALP) content was examined by ALP activity assay kit, the gene expression of ALP, collagen Ⅰ(ColⅠ), osteocalcin (OC), runt-related transcription factors (Runx2) was measured by real-time quantitative PCR, and the protein expression of ColⅠ and OC was tested by Western blot. Results Under the condition of hypergravity, cell body of osteoblast became thinner, but its surface area increased significantly; with the structure of skeletal arrangement becoming loose, actin microfilament structure reduced so that arrangement of actin-like dispersion orderly lowered. The gene expressions of related indicators of osteogenic differentiation including ALP, ColⅠ, OC, Runx2 loaded by hypergravity were significantly up-regulated, which was the same as ColⅠ protein and OC protein after hypergravity loading. There was only a very minute quantity of small red-orange nodules in the control group, while the cells after hypergravity loading in experimental groups obviously formed various sizes of red-orange nodules. Conclusions Under hypergravity, changes in osteoblast morphology can be triggered by rearrangements of skeletal structure. Furthermore, osteoblast maturation and differentiation can be stimulated effectively by up-regulating differentiation-related gene and protein expressions.