Upgrowth of Tibial Heterotopic Vascularized Corticoperiosteal Flap in Dog compared with Rabbit

Comparative investigation on capacity of vascularized corticoperiosteal tibial flap to produce heterotopic osteogenesis in dogs versus rabbits. Methods – Proximal tibial flap was taken and transferred in the popliteal muscle. The development of the flap and the new formed bone were evaluated by digital computer image radioopacity, bone mineral density, osseous resorption and bone quality. Results In rabbits the bone mineral density increased to values close to the reference value of normal tibial bone tissue. In dog, contrariwise, a marked decrease was observed. Histological sections in rabbit revealed that the regenerated tissue shows a high functional organization level and in the dog revealed the presence of the osseous resorption process. Clinical significance The heterotopic flaps have different behavior in dog from rabbits. Dog’s developmental model confirms the incapacity of the osteogenic reaction of the vascularized corticoperiosteal flaps in heterotopic transplantation. INTRODUCTION Periosteum’s role in tissue-engineered bone is widely recognized (Zhang et al., 2005; Zhu et al., 2006; Kaminski et al., 2009; Takeuchi et al., 2010) but the role of vascularized periosteum flap as a graft and the factors stimulating it into osteogenic activity remain obscure (Chen et al., 2009). The idea of using vascularized periosteal flaps in reconstructing bone defects is more than one hundred years old. Up to now, experimental and clinical results regarding their osteogenic capacity have been a subject of debate. In rats, the studies of ectopic bone formation in the groin and orthotopic bone formation in the femoral defect demonstrates that optimal bone formation requires four factors: bone morphogenetic protein (rh-BMP-2), a biodegradable matrix, osteoprogenitor cells, and blood supply (Kusumoto et al., 1997; Kusumoto et al., 1998; Vogelin et al., 2000; Vogelin et al., 2002). Few articles recognized periosteum’s osteogenic process without the dependence of stress stimulation or bony contact. Experimental study on rabbits reported that vascularized periosteal flaps presented strong osteogenic capacity in heterotopic conditions (Dailiana et al., 2002; Ortak et al., 2005; Chen et al., 2009). The information above was the basis of our investigations in this study, in which we aimed to evaluate comparatively the quantity and quality of bone formed in heterotopic conditions from vascularized corticoperiosteal flaps in the dogs versus rabbits. MATERIALS AND METHODS In the present study 8 white New Zeeland mature rabbits were enrolled, weighing 2.5 to 3 kg (group A), and 8 adult (mixed breed) dogs weighing 22 to 25 kg (group B). The study was approved by the ethics committee of our institution (resuming European Directive 86/609/EEC). Under general anesthesia and aseptic conditions, on both hind limbs, a longitudinal skin incision was made on the medial surface of tibia. A portion of tibial diaphyseal periosteum (approximately of 5 mm x 7 mm in rabbits, and 20 mm x 30 mm in dogs) was dissected and elevated maintaining the contact of pedicle with the tibia at its proximal portion. A bone fragment, harvested with a saw, of 10/5/3 mm in rabbits, and of 20/30/5 mm in dogs (width/ length/ thickness) remains attached at the distal portion of the pedicle. The dissection of osteo-periosteal pedicle includes a vascular pedicle originating from saphenous artery and vein. Each flap consisted of four layers: cortical bone, periosteum, muscle, and fascia. The vascular patency was checked during specimen sampling. The elevated vascularized osteo-periosteal pedicle was transferred into the adjacent musculature in the pocket created in popliteal muscle. The opposite limb served as control (reference values) for radiographic investigation and for bone mineral density (BMD) measurement. After the surgical intervention, analgesia was provided by butorphanol (Butomidor, Richter Pharma) (0.2 mg/kg, subcutaneously q4h in the first 24 hours, and q12h in the following 48 hours). The development and density of osteo-periosteal pedicle was evaluated weekly, postoperatively during 16 weeks by digital computer image (C-arm, Siremobil Compact L (Siemens, Germany), using an image intensifier of 7 inches). On a period of 0 to 8 weeks X-ray investigations were made to a total of eight individuals from both groups, and between 10 to 16 weeks to four individuals from both groups. These investigations aimed to establish the presence, formation, emplacement and radiodensity of the flap. Under general anaesthesia each animal was exposed under fixed irradiated conditions (50 kV in voltage, 0.4 mAS current, 70 cm distance – for dogs and 47 kV in voltage, 0.3 mAS current, 70 cm distance – for rabbits) in two views (cranio-caudal and oblique). On the