Structural allograft healing is limited because of a lack of vascularization and remodeling. To study this we developed a mouse model that recapitulates the clinical aspects of live autograft and processed allograft healing. Gene expression analyses showed that there is a substantial decrease in the genes encoding RANKL and VEGF during allograft healing. Loss-of-function studies showed that both factors are required for autograft healing. To determine whether addition of these signals could stimulate allograft vascularization and remodeling, we developed a new approach in which rAAV can be freeze-dried onto the cortical surface without losing infectivity. We show that combination rAAV-RANKL-and rAAV-VEGF-coated allografts show marked remodeling and vascularization, which leads to a new bone collar around the graft. In conclusion, we find that RANKL and VEGF are necessary and sufficient for efficient autograft remodeling and can be transferred using rAAV to revitalize structural allografts.In contrast to soft tissue organ transplantation (i.e., heart, liver, kidney), which must be live from a histocompatible donor, structural musculoskeletal grafts (i.e., bone, ligament) are often derived from allogenic cadavers. Although this convenience makes structural allografts readily available, the utility of these transplants is limited by their lack of viability. This is most evident from experimental and clinical studies showing that fresh vascularized autogenous grafts are vastly superior to allograft in terms of healing and remodeling 1,2 . Structural bone grafts used to heal critical defects and bone fusions undergo a repair and remodeling process that closely resembles fracture healing 3 . In live autograft healing, cells from both the graft and the host contribute to mediate bony union 4,5 . In contrast, healing of a diaphyseal defect that has been allografted can only be accomplished by invasion of the graft by host tissue to obtain a cortexCorrespondence should be addressed to E.M.S. (edward_schwarz@urmc.rochester.edu).. COMPETING INTERESTS STATEMENT The authors declare competing financial interests (see the Nature Medicine website for details). to-cortex union 6 . Following union, autografts continue to remodel as a result of osteoclastic resorption of necrotic or disused cortical bone that is followed by osteoblastic formation of new woven bone, which is later remodeled into stronger lamellar bone. In this way, autografts are sustained through normal bone homeostasis. In contrast, once creeping callus from the host calcifies on the cortex of an allograft, healing ceases, leaving a large segment of necrotic bone that is unable to respond to normal mechanical loading. Thus, structural allografts have a limited life span because microfractures that occur in them over time cannot be remodeled and repaired, and negative outcomes include a 25-35% failure rate from infection, nonunion and fracture 7,8 . NIH Public AccessTwo central issues that must be addressed to improve structural allografting are elucidatio...

To further understand the cellular and molecular mechanisms underlying cortical bone graft healing, we have developed a novel mouse femur model that permits quantitative and molecular analysis of structural bone graft healing. A 4 mm mid-diaphyseal femoral segment was removed and replaced by either immediate implantation of a fresh autograft, a frozen, genetically identical isograft or a frozen allograft from a different strain of mouse, which was secured with a 22-gauge metal intramedullary pin. Healing was evaluated by radiology, histomorphometry, and in situ hybridization. Autograft repair occurred by endochondral bone formation at the host-graft junction and by intramembranous bone formation along the length of the graft bed at 2 weeks, with maturation and remodeling apparent by 4 weeks. Bone repair in allografts and isografts completely relied on endochondral bone formation at the host-graft cortical junction, with absence of periosteal bone formation along the length of the graft, suggesting that live periosteal cells from the donor tissue are necessary for this response. This small animal model of structural bone grafting can be used to evaluate tissue-engineered allografts and novel bone graft substitutes using quantitative and molecularly defined outcome measures.

Lead exposure continues to be a significant public health problem. In addition to acute toxicity, Pb has an extremely long half-life in bone. Individuals with past exposure develop increased blood Pb levels during periods of high bone turnover or resorption. Pb is known to affect osteoblasts, osteoclasts, and chondrocytes and has been associated with osteoporosis. However, its effects on skeletal repair have not been studied. We exposed C57/B6 mice to various concentrations of Pb acetate in their drinking water to achieve environmentally relevant blood Pb levels, measured by atomic absorption. After exposure for 6 weeks, each mouse underwent closed tibia fracture. Radiographs were followed and histologic analysis was performed at 7, 14, and 21 days. In mice exposed to low Pb concentrations, fracture healing was characterized by a delay in bridging cartilage formation, decreased collagen type II and type X expression at 7 days, a 5-fold increase in cartilage formation at day 14 associated with delayed maturation and calcification, and a persistence of cartilage at day 21. Fibrous nonunions at 21 days were prevalent in mice receiving very high Pb exposures. Pb significantly inhibited ex vivo bone nodule formation but had no effect on osteoclasts isolated from Pb-exposed animals. No significant effects on osteoclast number or activity were observed. We conclude that Pb delays fracture healing at environmentally relevant doses and induces fibrous nonunions at higher doses by inhibiting the progression of endochondral ossification.

Object. The authors evaluated the effects of pilot hole preparation technique on insertional torque and axial pullout resistance in osteoporotic thoracic and lumbar vertebrae. Methods. Using a probe technique and fluoroscopy, 102 pedicle screws were placed in 51 dual-energy x-ray absorptiometry—proven osteoporotic thoracic and lumbar levels. Screws were inserted using the same-size tapping, one-size-under tapping, or no-tapping technique. Insertional torque and axial pullout resistance were measured. Analysis of variance, Fisher exact test, and regression analysis were performed. Same-size tapping decreased pullout resistance in the lumbar spine. There was no effect on pullout resistance in the thoracic spine. Pullout resistance values were lower for all insertion techniques in the upper thoracic spine. Insertional torque and bone mineral density correlated with pullout resistance in the thoracic and lumbar spine. Conclusions. Tapping decreased pedicle screw pullout resistance in the osteoporotic human lumbar spine, although it did not affect pullout strength in the thoracic spine. Tapping decreased insertional torque in upper thoracic levels. Surgeons should optimize overall construct rigidity when placing thoracic pedicle screws in patients with spinal segment osteoporosis.

Level II.