ANABOLIC HORMONES IN BONE: BASIC RESEARCH AND THERAPEUTIC POTENTIAL

Participants: E.P. Frankenburg, H. Chen, K.A. Sweet, B.T. Nolan, K. Lucas, R.L. Taylor, M.W. Stock, L.K. McCauley, S.A. Goldstein

Keywords: parathyroid hormone, fracture repair, animal model

Introduction

Bone fragility disorders such as osteoporosis, osteogenesis imperfecta, and many others are disorders that result from the incorrect control or functioning of bone forming cells. Successful determination of the mechanisms by which bone cells are stimulated to function should lead to substantially new and innovative therapies for treating bone disorders and fractures. PTH is an important hormone that regulates the formation or resorption of bone, although the mechanism by which it works is unclear. The specific objectives of this project are to determine whether fracture healing can be enhanced by local, systemic, or a combination of local and systemic delivery of PTH.

Materials and Methods

The model used in this study consisted of a five millimeter osteotomy in the femoral diaphysis of normal adult male Sprague-Dawley rats. The femur was stabilized with a customized external fixator. Before closing the wound, the gaps were filled with either a degradable implant or the same implant that had been infiltrated with PTH-1 receptor cDNA (R15B). The limb which received the gene/matrix was randomized, and the contralateral limb received the alternate implant type. A total of twenty animals were included into the study, and assigned to one of four groups. Animals from two of the groups were injected with rat parathyroid hormone (1-34) on a daily basis over a six week period. The other two groups were injected with a vehicle of 0.15 NaCl, 0.001 N HCL, and 2% heat inactivated rat serum. Within each injection group, the animals were either assigned for RNA isolation by Northern Analysis or for histomorphometry. The animals were allowed to ambulate freely, and weekly radiographs were taken of each animal to determine fracture site healing and pin site health. At six weeks post-surgery, the animals are euthanized, urine and blood samples as well as the experimental femurs were taken for in vitro analysis. In addition to either histomorphologic or Northern analyses, the femora had microradiographs taken, as well as DEXA scans to determine bone density and fracture healing quality. Finally, L4 vertebrae and tibiae from each specimen were micro-radiographed to determine the systemic effects of the injected PTH.

Progress

All animals have been surgically manipulated and sacrificed after six weeks. Urine and serum samples have begun to be analyzed. All specimens for northern analysis have been snap-frozen. The histomorphometry specimens have been dissected, fixed in Bouin’s, and stored in 70% EtOH. They have had microradiographs taken and have been DEXA scanned and prepared for light microscopy evaluation.