2.5.1 Bone induction surgery in mice

All animal experimental protocols were approved by the Animal Ethics Committee of Chengdu University. Surgery was conducted on animals after 1 week of adaptive housing. Twenty-four 8-week-old female ICR mice (Chengdu Dashuo Experimental Animal Co., Ltd., China) were randomly divided into the following four groups (n = 6 each): 0.01 mL of CS, 0.01 mL of Sr-HA/CS, 10 μg of calcium phosphate (CAP), and 0.01 mL of CAP/CS. Calcium phosphate-based materials are very similar to the inorganic components of human bone, both in terms of chemical composition and biological properties, and are therefore widely used in bone tissue engineering and clinical medicine. In our study, we chose to increase the CAP group for comparison, thus better demonstrating the advantages of our prepared materials. Before implantation, CAP, an amorphous powder, was moistened with sterile PBS and thereafter underwent natural drying and solidification. The ratio of Sr HA/CAP: CS hydrogel was 0.05 g: 10 mL. The mice were anesthetized using isoflurane gas (Shenzhen Reward Life Science and Technology Co., Ltd.). After complete anesthesia, the hair on both outer thighs of mice was clipped, the skin was sterilized with ethanol, and a 10-mm long incision was made in the skin. Subsequently, a longitudinal muscle pocket measuring approximately 8 mm in length was promptly prepared along the skin incision. Finally, the four kinds of materials were implanted, and the muscles and skin were sutured sequentially. Normal postoperative feeding was performed and penicillin was injected for 3 consecutive days to prevent postoperative infection. Following the CO₂-inhalation-induced execution of mice at 8 and 10 weeks, the tissues were fixed for 72 h in 4% paraformaldehyde.