Cranial Bone Defect Creation and Healing Assessment

The animals were anesthetized with an intramuscular injection of a mixture of ketamine hydrochloride (Ketalar, Yuhan Co., Seoul, Korea) and xylazine (Rompun, Bayer Korea Co., Seoul, Korea). The surgical sites were shaven and then draped with alcohol and povidone iodine, followed by local anesthesia with 2% lidocaine (Lidocaine HCl, Huons, Seoul, Korea). An incision was made along the sagittal midline from the frontal bone to the occipital bone. A full-thickness flap was elevated to expose the cranial bone. Standardized circular defects with diameters of 6, 8, 11, and 15 mm were created using trephines of the corresponding sizes under cool-saline irrigation. The soft tissues were repositioned and then sutured layer by layer with a resorbable suture material (4-0 Vicryl, Ethicon, Somerville, NJ, USA) to achieve primary closure. The stitches were removed after 10 days. The animals were sacrificed at 2, 4, 8, or 12 weeks postoperatively (Fig. 1).