The goal of this study was to use a segmental defect model in the rat femur to determine if osteogenic protein-1 (OP-1) is capable of inducing bone formation in the presence of bacterial contamination. A 6 mni segmental defect was surgically created and stabilized with a polyacetyl plate and Kirschner wires in one femur in each of 126 Sprague-Dawley rats. The animals were divided into eight groups in which the defect was either left untreated, or subjected to various combinations of OP-l (1 I or 50 pg), lyophilized bovine type I collagen (carrier for the OP-I), and 10' colony-forming units of Strcphy1ococcu.r ciureus. The animals were euthanized at either 2, 4, or 9 weeks. Quantitative radiographic and histologic analyses were performed on the harvested tissue. The initial contamination progressed to infection in all animals receiving bacteria, as determined by qualitative bacteriology. There was very little, if any, bone formation in the untreated defects, and in the contaminated defects with or without collagen carrier. Bone formation was significantly greater in contaminated defects with either dose of OP-1, compared with contaminated defects without 01'-1. The 50 pg dose of OP-1 induced significantly more bone formation than the 11 pg dose, both with and without bacteria. This investigation has demonstrated that OP-l maintains its osteoinductive capability in a contaminated segmental defect. OP-1 may potentially be used in the clinical management of Contaminated fractures.

The aim of this study was to characterize a new model of chronic osteomyelitis with clinically relevant features. A segmental defect of critical size was surgically created in the rat femur, stabilized with a polyacetyl plate and Kirschner wires, and contaminated with bacteria. The animals were allowed to recover while the contamination progressed to a chronic infection. At a later point in time, the defect was surgically debrided without removing the implant. Further treatments of interest, such as antibiotic therapy or application of an osteogenic agent, could be introduced at this time. To implement this model, an initial experiment was performed to determine the bacterial inoculum and time from contamination that would reliably result in an infected defect without causing excessive bone damage by the time debridement surgery was performed. The number of recovered bacteria, degree of radiographic bony lysis, and torsional stiffness of the defect fixation were measured in 192 rats as a function of 4 inocula of Staphylococcus aureus (lo', lo4, 10' or lo6 CFUs) and 4 times from contamination (1,2, 3 or 4 weeks). A lo4 CFU inoculum over 2 weeks was found to consistently create an infection without severe lysis and loss of fixation stability. Based on these values, a second experiment was performed in 96 rats to characterize the debrided defect over time (2,4,8 and 12 weeks after debridement), with and without 4 weeks of the antibiotic ceftriaxone, in terms of the same outcome variables. Infection was persistent in all animals in spite of debridement and antibiotic therapy. Antibiotic therapy did not reduce the degree of bony lysis. Compared with animals not given antibiotic, bacterial counts significantly decreased during the period of antibiotic therapy, but then rebounded to significantly higher levels at 12 weeks. This model allows us to perform further studies on differing regimens of antibiotic therapy and their relationship to surgical dtbridement, and on the efficacy of osteogenic agents in the presence of infection.

Chronic alcohol abuse is a major risk factor for osteoporosis but the effects of moderate drinking on bone metabolism are largely uninvestigated. Here, we studied the long-term dose-response (0, 3, 6, 13, and 35% caloric intake) effects of alcohol on cancellous bone in the proximal tibia of 8-month-old female rats. After 4 months of treatment, all alcohol-consuming groups of rats had decreased bone turnover. The inhibitory effects of alcohol on bone formation were dose dependent. A reduction in osteoclast number occurred at the lowest level of consumption but there were no further reductions with higher levels of consumption. An imbalance between bone formation and bone resorption at higher levels of consumption of alcohol resulted in trabecular thinning. Our observations in rats raise the concern that moderate consumption of alcoholic beverages in humans may reduce bone turnover and potentially have detrimental effects on the skeleton.

Several studies were performed in female rats to determine dose and time course changes in mRNA levels for matrix proteins in bone after a single administration of ethanol. As expected, dose-dependent transient increases in blood ethanol were measured. Additionally, there was mild hypocalcemia with no change in immunoreactive parathyroid hormone. Coordinated dose-dependent increases in mRNA for type 1 collagen, osteonectin, and osteocalcin were noted in the proximal tibial metaphysis 6 hr after ethanol was given, with the peak values occurring at a dose of 1.2 g/kg (0.4 ml). Similar increases in mRNA levels for matrix proteins were noted in lumbar vertebrae after ethanol treatment. The changes were specific for bone; ethanol had no effect on mRNA levels for matrix proteins in the uterus or liver, although the mRNA concentrations tended to be reduced in uterus. Message levels for several cytokines implicated in the regulation of bone turnover were also assayed; mRNA levels for transforming growth factor-beta1, transforming growth factor-beta2, interferon-gamma, and interleukin-6 were unchanged at doses ranging from 0.14 to 1.7 g/kg. At the highest dose of ethanol, the mRNA level for tumor necrosis factor-alpha was elevated while the level for insulin-like growth factor-1 was reduced. The time course effects of ethanol (0.4 ml dose) were determined in a separate experiment. Ethanol resulted in a transient increase in mRNA levels for the three bone matrix proteins assayed. However, matrix protein synthesis, as determined by incorporation of 3H-proline into the proximal tibial metaphysis, was not changed after 6 hr. The changes in mRNA levels for the matrix proteins were preceded by brief, transient decreases in mRNA levels for interleukin-1beta, interferon-gamma, and migration inhibitory factor, and followed by a more prolonged decrease in the mRNA level for insulin-like growth factor-1. A subsequent study was performed to determine the effects of repetitive daily treatment with ethanol on rat bone. After 7 days, there were highly significant decreases in the mRNA level for type 1 collagen, as well as decreased bone formation. These results suggest that ethanol may alter bone metabolism by disturbing signal transduction pathways that regulate the expression of genes for bone matrix proteins, skeletal growth factors, and cytokines.