Bone defects, particularly large bone defects resulting from infections, trauma, surgical resection or genetic malformations, maintain a significant challenge for clinicians. In this study, the tricalcium silicate/mesoporous bioactive glass (C3S/MBG) cement scaffolds were successfully fabricated for the first time by 3D printing with a curing process, which combined the hydraulicity of C3S with the excellent biological property of MBG together. The C3S/MBG scaffolds exhibited 3D interconnected macropores (~400μm), high porosity (~70%), enhanced mechanical strength (>12MPa) and excellent apatite mineralization ability. Human bone marrow-derived mesenchymal stem cells (hBMSCs) were cultured on the scaffolds to evaluate their cell responses, and the results showed that C3S/MBG scaffolds could stimulate hBMSCs attachment, proliferation and differentiation with increasing MBG component. The critical-sized rat calvarical defects as an animal model, further in vivo results indicated both C3S and C3S/MBG30 scaffolds could induce new bone formation, but the C3S/MBG30 scaffolds significantly improved osteogenic capacity compared to the pure C3S scaffolds. Therefore, the C3S/MBG cement scaffolds fabricated by 3D printing with a curing process would be a promising candidate for bone regeneration.