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, 36 (3), 94-102

Bone Healing Properties of Autoclaved Autogenous Bone Grafts Incorporating Recombinant Human Bone Morphogenetic Protein-2 and Comparison of Two Delivery Systems in a Segmental Rabbit Radius Defect

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Bone Healing Properties of Autoclaved Autogenous Bone Grafts Incorporating Recombinant Human Bone Morphogenetic Protein-2 and Comparison of Two Delivery Systems in a Segmental Rabbit Radius Defect

Eun Joo Choi et al. Maxillofac Plast Reconstr Surg.

Abstract

Purpose: This study aims to validate the effect of autoclaved autogenous bone (AAB), incorporating Escherichia coli-derived recombinant human bone morphogenetic protein-2 (ErhBMP-2), on critical-sized, segmental radius defects in rabbits. Delivery systems using absorbable collagen sponge (ACS) and fibrin glue (FG) were also evaluated.

Methods: Radius defects were made in 12 New Zealand white rabbits. After autoclaving, the resected bone was reinserted and fixed. The animals were classified into three groups: only AAB reinserted (group 1, control), and AAB and ErhBMP-2 inserted using an ACS (group 2) or FG (group 3) as a carrier. Animals were sacrificed six or 12 weeks after surgery. Specimens were evaluated using radiology and histology.

Results: Micro-computed tomography images showed the best bony union in group 2 at six and 12 weeks after operation. Quantitative analysis showed all indices except trabecular thickness were the highest in group 2 and the lowest in group 1 at twelve weeks. Histologic results showed the greatest bony union between AAB and radial bone at twelve weeks, indicating the highest degree of engraftment.

Conclusion: ErhBMP-2 increases bony healing when applied on AAB graft sites. In addition, the ACS was reconfirmed as a useful delivery system for ErhBMP-2.

Keywords: Autogenous autoclaved bone; BMP carrier; Bone morphogenetic protein 2; Critical bone defect; Rabbit radius.

Figures

Fig. 1.
Fig. 1.
Experimental procedure for the AAB graft on the rabbit radius. (A) Reinsertion of the resected segments after autoclaving, then internal fixation with the screws and plate. (B) Insertion of ErhBMP-2 along with ACS carrier material. (C) Segmental resection of the radial bone. (D) Radius removed with soft tissue cuff. (E) Radius after removal of soft tissue, including periosteum. (F) Schematic drawing of longitudinal hemisection and bone marrow removal. (G) Preparation and drape of foreleg. (H) Exposure of periosteum of radius. AAB, autoclaved autogenous bone; ErhBMP, Escherichia coli-derived recombinant human bone morphogenetic protein; ACS, absorbable collagen sponge.
Fig. 2.
Fig. 2.
Schematic drawing of ROI. Quantitative analyses: BV/TV, TbTh, TbN, and TbSp, were measured using micro-computed tomography within this ROI at each proximal end of the autoclaved autogenous bone. ROI, region of interest; BV, bone volume; TV, total volume; Tb, trabecular; Th, thickness; N, number; Sp, separation.
Fig. 3.
Fig. 3.
Method for calculating the degree of engraftment. Red line represents the autoclaved autogenous bone perimeter, within which we measured the proportion of perimeter showing osteoid, osteoblasts, and the reversal line at higher magnification (H&E, ×40).
Fig. 4.
Fig. 4.
Plain radiographic findings after graft of AAB. Six weeks after operation, discontinuity was seen in groups 1 and 3; whereas, 12 weeks after operation, continuity between the AAB and radius was seen in all groups. Only AAB reinserted (group 1), and AAB and ErhBMP-2 inserted using an ACS (group 2) or FG (group 3) as a carrier. AAB, autoclaved auto-genous bone; ErhBMP-2, Escherichia coli-derived recombinant human bone morphogenetic protein-2; ACS, absorbable collagen sponge; FG, fibrin glue.
Fig. 5.
Fig. 5.
Micro-computed tomography findings for each group at six and 12 weeks. White arrows represent both ends of autoclaved autogenous bone. Only AAB reinserted (group 1), and AAB and ErhBMP-2 inserted using an ACS (group 2) or FG (group 3) as a carrier. AAB, autoclaved autogenous bone; ErhBMP-2, Escherichia coli-derived recombinant human bone morphogenetic protein-2; ACS, absorbable collagen sponge; FG, fibrin glue.
Fig. 6.
Fig. 6.
Histologic assessment of each animal at six and 12 weeks (A∼C: six weeks after surgery, D∼F: twelve weeks after surgery). (A) Group 1, six weeks after surgery. Although woven bone originated from the adjacent ulnar bone and a reversal line, suggesting bony remodeling, was observed, the new bone did not unite with the AAB. (B) Group 3, six weeks after surgery. Although woven bone originated from the adjacent ulnar bone and a reversal line which suggests bony remodeling was observed, the new bone did not unite with the AAB. (C) Group 2, six weeks after surgery. High magnification revealed extensive cartilage formation around the AAB, active cell division of chondroblasts, and replacement of cartilage by osteoid. (D) Group 1, twelve weeks after surgery. Some osteoclasts and osteoblasts were also observed, suggestive of bony remodeling, and woven bone originated from a single end of the radial bone and restored some continuity with the AAB. Sparse new bone formation and a reversal line suggested proper engraftment of AAB. (E) Group 2, 12 weeks after surgery. Woven bone originated from the adjacent cutting ends of the radius and showed complete fusion with AAB. At low magnification, AAB had completely restored continuity with the radial bone marrow. Moreover, high magnification revealed reversal lines and osteocytes inside the woven bone, suggesting that active bony remodeling was in progress. (F) Group 3, twelve weeks after surgery. There was some evidence suggesting bony remodeling around AAB, similar to the group 1 results, although not as extensive as in group 2 (H&E; left: ×40, right: ×100/×400). Boxes with yellow line in left slides (×40) were magnified in right slides (×100/×400) to be determined whether complete bony fusion with osteoblast, osteoclast, and osteoid was done. U, ulnar bone; R, radial bone; AAB, autoclaved autogenous bone; W, woven bone; black arrows, reversal line; blue arrows, end of AAB; purple arrows, resorption of AAB; green arrow, osteoclast.

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