Systemic effects of ZA treatment.

<p>A: No significant difference in kidney histology or BUN serum level between control and ZA-treated animals. 9B: No significant difference in liver histology or ALT activity, used to evaluate liver function between the two groups. 9C: No significant difference in body weights between the two groups.</p

A Model for Osteonecrosis of the Jaw with Zoledronate Treatment following Repeated Major Trauma

<div><p>This study aims to develop a reproducible rat model for post-traumatic bisphosphonate-related osteonecrosis of the jaw (BRONJ). In our previous studies using dental extraction as an inducing factor, only 30% - 60% of zoledronate-treated animals fulfilled the definition of clinical BRONJ. We modified the zoledronate regimen and introduced repeated surgical extraction to illicit quantifiable BRONJ in all animals. Eighty retired-breeder female Sprague-Dawley rats were divided between the treatment (IV zoledronate; 80 μg/kg/week for 13 weeks) and control (saline) groups. On week 13, the left mandibular first molar was surgically extracted, followed by the second molar a week later. Animals were euthanized at 1-week, 2-weeks, and 8-weeks following extraction. The occurrence and severity of BRONJ were scored in each animal based on gross and MicroCT analysis. Parameters of bone formation and osteoclast functions at the extraction site were compared between groups. All zoledronate-treated animals developed a severe case of BRONJ that fulfilled the clinical definition of the condition in humans. Osteoclast attachment continued to be defective eight weeks after stopping the treatment. There were no signs of kidney or liver toxicity. Our data confirmed that repeated surgical extraction (major trauma) by itself consistently precipitated massive bone necrosis in ZA-treated animals, eliminating the need to induce pre-existing infection or comorbidity. These results will be the basis for further studies examining the <i>in-vivo</i> pathogenesis and prevention of BRONJ.</p></div

Analysis of alveolar bone at the extraction site, excluding any sequestered bone.

<p>The numbers of empty lacunae over total number of osteocyte lacunae were used to calculate the percentage of dead osteocytes within the bone. There was a significant increase in the percentage of empty lacunae (arrows) on the extraction sites on ZA-treated animals. There was also a significant decline in the numbers of empty lacunae in the control group from week 1 to week 8. The un-operated sites in the ZA-treated animals did not show a significant difference in bone viability, compared to the un-operated sites in control animals, signifying that surgical extraction was necessary to illicit bone necrosis.</p

Calcein double-labeling in Control and Treatment groups within the extraction sites have shown that Mineral Formation Rate (MFR) and mineral Apposition Rate (MAR) within the extraction sites 2 weeks post extraction was diminished in the ZA-treated animals, compared to controls.

<p>Bottom panel: Staining with modified Brown and Brenn (B&B; 100x) stain showed minimal to no bacterial colonization of alveolar bone either in control or ZA-treated animals.</p

Systemic effects of ZA treatment.

<p>A: No significant difference in kidney histology or BUN serum level between control and ZA-treated animals. 9B: No significant difference in liver histology or ALT activity, used to evaluate liver function between the two groups. 9C: No significant difference in body weights between the two groups.</p

Intraoral images of extraction sites in control vs. ZA-treated rats, showing persistent exposure of necrotic bone in the ZA-treated animals 8 weeks after extraction.

<p>Untreated animals showed complete healing.</p

TRAP-labeled decalcified sections from dental extraction sites at 1-week following extraction.

<p>A. Characterization of the number of Oc (N.Oc) normalized to the bone surface perimeter (B.Pm). B. Osteoclast perimeter vs bone surface perimeter.</p

Summary of Previous BRONJ Animal Models.

<p>Summary of Previous BRONJ Animal Models.</p

MicroCT images of Control and ZA-treated mandibles, eight weeks after extractions, showing massive sequestration in a treated animal, compared to normal healing in the control.

<p>Quantifying the severity of osteonecrosis on a 0–4 scale, the ZA-treated animals received a score between 3 and 4, compared to 0–1 in control animals.</p