Craig Godfrey

Optimal Timing of a Single Dose of Zoledronic Acid to Increase Strength in Rat Fracture Repair

We hypothesized that ZA treatment would bolster fracture repair. In a rat model for closed fracture healing, a single dose of ZA at 0, 1, or 2 wk after fracture significantly increased BMC and strength of the healed fracture. Delaying the dose (1 or 2 wk after fracture) displayed superior results compared with dosing at the time of fracture.

Manipulation of the Anabolic and Catabolic Responses With OP‐1 and Zoledronic Acid in a Rat Critical Defect Model

Bone repair involves both anabolic and catabolic responses. We hypothesized that anabolic treatment with OP‐1 (BMP‐7) and anti‐catabolic treatment with zoledronic acid could be synergistic. In a rat critical defect, this combination therapy produced significant increases in new bone volume and strength.

Bolus or weekly zoledronic acid administration does not delay endochondral fracture repair but weekly dosing enhances delays in hard callus remodeling

INTRODUCTION It has been widely assumed that osteoclasts play a pivotal role during the entire process of fracture healing. Bisphosphonates (BPs) are anti-catabolic agents commonly used to treat metabolic bone diseases including osteoporosis, minimizing fracture incidence. Yet, fractures do occur in these patients and the potential for negative effects of BPs on healing has been suggested. We aimed to examine the effect of different dosing regimes of the potent BP zoledronic acid (ZA) on early endochondral fracture repair and later callus remodeling in a normal bone healing environment. METHODS Saline, a Bolus dose of 0.1 degrees mg/kg ZA or 5 weekly divided doses of 0.02 degrees mg/kg of ZA commenced 1 week post operatively in a rat closed fracture model. Samples at 1, 2, 4 and 6 weeks post fracture were used to analyze initial fracture union, and 12 and 26 weeks post fracture to investigate the progress of remodeling. RESULTS ZA did not alter the rate of endochondral fracture union. All fractures united by 6 weeks, with no difference in the progressive reduction of cartilaginous soft callus between control and treatment groups over time. ZA treatment increased hard callus bone mineral content (BMC), volume and increased callus strength at 6 and 26 weeks post fracture. Hard callus remodeling commenced at 4 weeks post fracture with Bolus ZA treatment but was delayed until after 6 weeks in the Weekly ZA group. By 12 and 26 weeks, Bolus ZA had equivalent callus content of remodeled neo-cortical bone to the Saline controls, whereas Weekly ZA remained reduced compared to Saline controls at these times (P<0.01). Callus material properties such as peak stress were significantly reduced in both ZA groups at 6 weeks. At 26 weeks, Bolus ZA-treated calluses generated peak stress equivalent to control values, whereas Weekly ZA callus peak stress remained significantly reduced, indicating remodeling delay. CONCLUSIONS Osteoclast inhibition with ZA does not delay endochondral fracture repair in healthy rats. Bolus ZA treatment increased net callus size and strength at 6 weeks while allowing hard callus remodeling to proceed in the long term, albeit more slowly than control. Prolonged bisphosphonate dosing during repair does not delay endochondral ossification but can significantly affect remodeling long after the drug is ceased.

Intermittent PTH((1-34)) does not increase union rates in open rat femoral fractures and exhibits attenuated anabolic effects compared to closed fractures.

Intermittent Parathyroid Hormone (PTH)((1-34)) has an established place in osteoporosis treatment, but also shows promising results in models of bone repair. Previous studies have been dominated by closed fracture models, where union is certain. One of the major clinical needs for anabolic therapies is the treatment of open and high energy fractures at risk of non-union. In the present study we therefore compared PTH((1-34)) treatment in models of both open and closed fractures. 108 male Wistar rats were randomly assigned to undergo standardized closed fractures or open osteotomies with periosteal stripping. 27 rats in each group were treated s.c. with PTH((1-34)) at a dose of 50 mug/kg 5 days a week, the other 27 receiving saline. Specimens were harvested at 6 weeks for mechanical testing (n=17) or histological analysis (n=10). In closed fractures, union by any definition was 100% in both PTH((1-34)) and saline groups at 6 weeks. In open fractures, the union rate was significantly lower (p<0.05), regardless of treatment. In open fractures the mechanically defined union rate was 10/16 (63%) in saline and 11/17 (65%) in PTH((1-34)) treated fractures. By histology, the union rate was 3/9 (33%) with saline and 5/10 (50%) with PTH((1-34)). Radiological union was seen in 13/25 (52%) for saline and 15/26 (58%) with PTH((1-34)). Open fractures were associated with decreases in bone mineral content (BMC) and volumetric bone mineral density (vBMD) on quantitative computerized tomography (QCT) analysis compared to closed fractures. PTH((1-34)) treatment in both models led to significant increases in callus BMC and volume as well as trabecular bone volume/total volume (BV/TV), as assessed histologically (p<0.01). In closed fractures, PTH((1-34)) had a robust effect on callus size and strength, with a 60% increase in peak torque (p<0.05). In the open fractures that united and could be tested, PTH((1-34)) treatment also increased peak torque by 49% compared to saline (p<0.05). In conclusion, intermittent PTH((1-34)) produced significant increases in callus size and strength in closed fractures, but failed to increase the rate of union in an open fracture model. In the open fractures that did unite, a muted response to PTH was seen compared to closed fractures. Further research is required to determine if PTH((1-34)) is an appropriate anabolic treatment for open fractures.

Models of Tibial Fracture Healing in Normal and Nf1 -Deficient Mice

Delayed union and nonunion are common complications associated with tibial fractures, particularly in the distal tibia. Existing mouse tibial fracture models are typically closed and middiaphyseal, and thus poorly recapitulate the prevailing conditions following surgery on a human open distal tibial fracture. This report describes our development of two open tibial fracture models in the mouse, where the bone is broken either in the tibial midshaft (mid‐diaphysis) or in the distal tibia. Fractures in the distal tibial model showed delayed repair compared to fractures in the tibial midshaft. These tibial fracture models were applied to both wild‐type and Nf1‐deficient (Nf1+/−) mice. Bone repair has been reported to be exceptionally problematic in human NF1 patients, and these patients can also spontaneously develop tibial nonunions (known as congenital pseudarthrosis of the tibia), which are recalcitrant to even vigorous intervention. pQCT analysis confirmed no fundamental differences in cortical or cancellous bone in Nf1‐deficient mouse tibiae compared to wild‐type mice. Although no difference in bone healing was seen in the tibial midshaft fracture model, the healing of distal tibial fractures was found to be impaired in Nf1+/− mice. The histological features associated with nonunited Nf1+/− fractures were variable, but included delayed cartilage removal, disproportionate fibrous invasion, insufficient new bone anabolism, and excessive catabolism. These findings imply that the pathology of tibial pseudarthrosis in human NF1 is complex and likely to be multifactorial.

Optimal duration of cooling for an acute scald contact burn injury in a porcine model.

The Australian and New Zealand Burn Association recommend 20 minutes of cold running tap water as burn first aid. Scientific evidence for the optimal duration of treatment is limited. Our aim was to establish the optimal duration of cooling using cold running tap water to treat the acute burn. Partial thickness contact scald burns were induced at five sites in each of 17 pigs. Treatments with cold running tap water for 5, 10, 20, and 30 minutes were randomly allocated to different sites together with an untreated control site. In the running water 5 and 10 minute treatments intradermal temperatures rose by 1°C per minute when cooling was stopped, compared with 0.5°C per minute for 20 and 30 minutes duration. No differences in the surface area of each burn were noted between the five treatments on day 9. Histological analysis of burn depth on days 1 and 9 revealed that a higher proportion of burns treated for 20 and 30 minutes showed improvement compared with those treated for 5 and 10 minutes only. This difference reached statistical significance (P < .05) only in the cold running water for 20 minutes treatment arm. There was a statistically significant (P < .05) improvement in burn depth in a porcine acute scald burn injury model when the burn was treated with cold running tap water for 20 minutes as opposed to the other treatment durations. This study supports the current burn first aid treatment recommendations for the optimal duration of cooling an acute scald burn.

Delayed Cooling of an Acute Scald Contact Burn Injury in a Porcine Model: Is it Worthwhile?

The current Australia and New Zealand Burn Association recommended burns first aid treatment is to place the burn under cool running water for 20 minutes. Immediate cooling of a burn wound has been shown to reduce the depth of the injury. Cooling has also been recommended as beneficial for up to 3 hours after the burn. No scientific data currently exist to support this recommendation. The aim of this study was to identify the effect of delayed cooling of an acute scald contact burn wound in a porcine model. Four partial-thickness contact scald burn injuries were induced in 12 piglets each. First aid treatment consisting of cool running water for 20 minutes was instituted randomly to each wound at different time points: immediately and at time delays of 5, 20, and 60 minutes. The group receiving immediate first aid with cool running water for 20 minutes served as the control group. At day 1 and day 9, biopsies were obtained and assessed in a blinded manner. Histologic analysis of burn depth on days 1 and 9 demonstrated no significant difference in the depth of the burn in the various treatment groups in comparison to the control group receiving immediate first aid. No significant differences in the surface areas of each burn were noted between the various treatment groups on day 9. Core body temperature did not fall below 35°C throughout the cooling process. This study provides scientific evidence that in an animal model delayed cooling for up to 60 minutes postacute contact scald burn is still effective compared with immediate cooling at reducing burn depth.

The correlation between time to skin grafting and hypertrophic scarring following an acute contact burn in a porcine model

One sequelae of burn injury remains the development of hypertrophic scarring. This appears more likely when the healing has been prolonged. Early excision of deep dermal burns and subsequent split skin grafting (SSG) may provide a more favorable result. The optimal timing of grafting for deeper dermal burns remains controversial. This study sought to establish evidence for the optimal grafting time using a porcine model. Five Large White female pigs were exposed to four contact burn injuries for duration of 20 seconds at 92°C. Each site was randomized to a treatment arm: dressing only as the control, SSG day 3, SSG day 14, and SSG day 21. Burn wound biopsies were obtained at days 0, 3, 14, 21, and 99 after the burn injury, together with microbiological swabs. Digital photographs were taken to assess scarring using the Vancouver scar scale. All biopsies were subject to histological and immunohistochemical analysis. Vancouver scar scale scores and histopathological analysis indicated that areas grafted on day 3 had the least fibrosis and scarring (P = 0.031). There was a strong correlation between the histological evaluation of the degree of fibrosis and &agr;-smooth muscle actin levels (r = .60, P = .014). A greater degree of fibrosis was observed in the presence of infection (P = .028). Sites grafted on day 3 consistently exhibited the best clinical and histological scar outcome. The increased fibrosis observed in delayed grafting may have been be related to progression of burn depth and infection. These results suggest that early grafting of deep dermal burns may be preferential.

Bisphosphonate‐laden acrylic bone cement: Mechanical properties, elution performance, and in vivo activity

Cemented total hip replacements generally fail after 10-20 years, often due to implant loosening from bone resorption. Bisphosphonates such as zoledronic acid (ZA) and pamidronate (PAM) are potent inhibitors of bone resorption. The local delivery of bisphosphonates via acrylic bone cement could decrease osteolysis and prolong implant lifespan. Conflicting studies suggest that bisphosphonate loading may or may not reduce the mechanical properties of acrylic bone cement. We assayed acrylic bone cement laden with ZA or PAM at different concentrations and diluent volumes. Four-point bend testing and compressive testing indicated that high volumes of diluent (with or without bisphosphonate) significantly reduced bending modulus and compressive strength. Radiography and electron microscopy indicated that high diluent volumes generated abnormal acrylic bone cement structure. After 6 weeks of incubation in saline, only 0.9% w/w of the total bisphosphonate incorporated in acrylic bone cement eluted in vitro, indicating a slow elution rate. In vivo testing was performed using a rat model. Cement cylinders were inserted into incisions in rat distal femora and ZA delivered locally (via elution from acrylic bone cement) or systemically (via injection). At 4 weeks postoperatively, dual energy X-ray absorptiometry demonstrated no significant increase in local bone mineral density (BMD) adjacent to ZA-laden implants. In contrast, systemic ZA delivery (0.1 mg/kg) led to a large (48.6%) and significant increase in BMD. Thus, systemic delivery appears more effective than local delivery.