Background: Metaphyseal fracture healing presents special biomechanical challenges in orthopaedic surgery. The void typically created by damage to the metaphyseal cancellous bone must usually be filled in order to recover the biomechanical integrity of the bone. While autologous bone grafting is a standard treatment for these fractures, bone graft substitutes delivered with or without pharmacologic agents may augment healing.

Hypothesis: Tricalcium phosphate (TCP) is a known osteoconductive bone filler and OP-1 an osteoinductive bone morphogenetic protein; both have been used in the past in diaphyseal fractures with success. PTH (parathyroid hormone) has been recently shown to enhance osteoblastic activity, to have a net anabolic effect on bone mass, and to enhance healing of diaphyseal fractures. Each of these agents may also enhance healing of metaphyseal fractures.

Objective: The potential of all above factors to accelerate metaphyseal fracture healing has been evaluated in a new metaphyseal fracture model developed in our laboratory in a rabbit model.

Material and Methods: A metaphyseal wedge osteotomy was created in the distal tibia of 16-week-old female New Zealand White rabbits (n=20). The osteotomy was bridged with a custom-made external fixator. The osteotomy gap was filled with TCP containing OP-1 (n=4), TCP alone with daily subcutaneous injections of 10μg/Kgr BW PTH (n=4), or TCP alone with daily subcutaneous administration of 40μg/Krg BW PTH (n=4). Two control groups, TCP alone (n=4) and normal healing (n=4), were also included. Assessment methods included biomechanical testing in both compression and torsion, radiographic examination, and QCT scans.

Results: Healing was observed in both PTH treated groups as well as in the OP-1 group at 4 weeks post-surgery. PTH appeared to have a systemic effect on bone formation, whereas the effect of OP-1 was local to the osteotomy site. In comparison, healing was delayed in the normal healing and TCP alone groups.

Conclusion: PTH and OP-1 both enhance metaphyseal fracture healing. The different systemic vs. local effects of these two agents, suggest that PTH and OP-1 may have potential synergism in accelerating healing of metaphyseal fractures.

Endochondral ossification involves a well ordered sequence of cellular events. Chondrocytes change their morphology and functions and are ultimately removed by the process of apoptosis. A variety of apoptotic-related signals have been characterised. These include Fas receptor (FasR)/Fas ligand (FasL), p53 and Bcl family. However, there is little known regarding the activity of these signals in the process of fracture healing. The purpose of this study was to investigate mRNA expression of apoptotic signals using RNase protection assay (RPA) and immunohistochemistry in endochondral bone formation.

BALB/C mice aged 8 to 10 weeks were used for this study. First, a transverse fracture was made in the right tibia. Mice were euthanised at 1, 2 and 3 weeks postfracture. The calluses were harvested and studied for the expression of caspase-8, a key enzyme of apoptosis, and apoptosis inducers: tumour necrosis factor-alpha (TNF-α) and its receptor p55, FasL and Fas receptor (FasR), and TNF-related apoptosis-inducing ligand (TRAIL). Four mice at each timepoint were used for immunostaining of fracture callus. Sections were incubated with primary antibody then labelled by avi-din-biotin complex method. Another four to ten tibiae were used for RPA. Fracture callus were harvested and snap frozen in liquid nitrogen. RNA was isolated by TRI reagent and BCP, and mRNAs expression of apoptotic signals were detected.

At each timepoint, mRNA of caspase-8, TNF-α, p55, FasL,FasR and TRAIL were detected by RPA. Immunostainings clearly showed that those apoptotic-related proteins were expressed by callus chondrocytes. Cartilaginous callus is replaced by woven bone in endochondral ossification. In this process, chondrocytes should be removed by the process of apoptosis in which death factors are elaborated directly in both an autocrine and paracrine manner.