This study aimed to investigate whether inhibition of glycogen synthase kinase-3β (GSK-3β) could promote chondrocytes proliferation. The expression pattern of GSK-3β was firstly determined by immunohistochemistry (IHC) in normal mouse. Tibias were then isolated and cultured for 6 days. The tibias were treated with dimethylsulfoxide (control) or GSK-3 inhibitor SB415286 (SB86). Length of tibias was measured until 6 days after treatment. These bones were either stained with alcian blue/alizarin red or analyzed by IHC. In addition, GSK-3β and β-catenin were analyzed by Western blot. Finally, cartilage-specific GSK-3β deletion mice (KO) were generated. Efficiency of GSK-3β deletion was determined through Western blot and IHC. After treated by inhibitor SB86, the overall length of growth plate was not changed. However, growth of tibia in SB86 group was increased by 31 %, the length of resting and proliferating was increased 13 % (P < 0.01), whereas the length of hypertrophic was decreased by 57 % (P < 0.01). Besides, the mineralized length was found to be significant longer than the control group (P < 0.05). In KO mice, growth plate and calvaria tissue both exhibit significant reduction of GSK-3β (P < 0.05) whereas the lengths of tibias in KO were almost same compared with control mice. Finally, an increase amount of β-catenin protein was observed in SB86 (P < 0.05). In addition, significantly increased β-catenin was also found in the growth plate of KO mice (P < 0.05). Inhibition of GSK-3 could promote longitudinal growth of bone through increasing bone formation. Besides, the inactivation of GSK-3β could lead to enhancing β-catenin, therefore promote chondrocytes proliferation.