| Parathyroid hormone (PTH) is under intensive investigation for its anabolic actions in bone and therapeutic potential. Elucidating PTH effects on osteoblasts is crucial for studying anabolic actions. However, conflicting results have been reported possibly due to differentiation stage-specific effects of PTH. Further, although promising as a clinical treatment of osteoporosis, use of PTH in local osseous defects is still uncertain. This dissertation was aimed at exploring PTH effects during osteoblastic differentiation in vitro and fracture healing in vivo.; PTH-stimulated increases in CAMP result in activation of protein kinase A, which is critical for anabolic actions. In order to quantify cAMP accurately, a sensitive CAMP binding protein assay was optimized and utilized to evaluate biologic activity of transfected cell lines. An in vitro osteoblastic differentiation model with C3H10T1/2 cells stably transfected with either functional or inactivated mutant PTH receptors was used to evaluate differentiation and apoptosis. Results demonstrated that overexpression of PTH receptors increased cell viability and promoted osteocalcin expression whereas inactivated mutant receptors had opposite effects. Interestingly, PTH was antiapoptotic in preconfluent and proapoptotic in postconfluent cells, effects that were both mediated by the PKA pathway.; The use of PTH in local osseous defects was evaluated using an in vivo fracture model combining a local gene therapy delivery with systemic PTH administration. Systemic PTH increased bone mineral density (BMD) of tibia and vertebrae, and increased serum osteocalcin levels during fracture healing. Furthermore, fracture sites in rats receiving systemic PTH demonstrated trends of greater bone mass than those in vehicle-injected rats using histological and radiographic analyses. The combination of systemic and local PTH led to higher BMD and bone area using DEXA analyses, a trend for greater radiographically detectable bone area, and higher expression of osteocalcin in fracture sites when compared to the individual or control treatment groups.; In summary, we conclude that PTH, mediated by PKA, promotes cell viability early, but cell departure from the differentiation program later. The accelerated osteoblast turnover may contribute to its anabolic actions. Additionally, our results support a combined systemic and local PTH approach in treating local osseous defects. |
