In a rabbit model we investigated the efficacy of a silk fibroin/hydroxyapatite (SF/HA) composite on the repair of a segmental bone defect. Four types of porous SF/HA composites (SF/HA-1, SF/HA-2, SF/HA-3, SF/HA-4) with different material ratios, pore sizes, porosity and additives were implanted subcutaneously into Sprague-Dawley rats to observe biodegradation. SF/HA-3, which had characteristics more suitable for a bone substitite based on strength and resorption was selected as a scaffold and co-cultured with rabbit bone-marrow stromal cells (BMSCs). A segmental bone defect was created in the rabbit radius. The animals were randomised into group 1 (SF/HA-3 combined with BMSCs implanted into the bone defect), group 2 (SF/HA implanted alone) and group 3 (nothing implanted). They were killed at four, eight and 12 weeks for visual, radiological and histological study. The bone defects had complete union for group 1 and partial union in group 2, 12 weeks after operation. There was no formation of new bone in group 3. We conclude that SF/HA-3 combined with BMSCs supports bone healing and offers potential as a bone-graft substitute.