Dose-dependent Effect of Zoledronic Acid on the Development of Medication-Related Jaw Osteonecrosis (MRONJ)-Like Lesion in C57Bl/6 Mice: A Histological Preliminary Study

Abstract

Medication-related osteonecrosis of the jaw (MRONJ) is a potentially painful condition affecting patients with previous or continuous treatment with antiresorptive or antiangiogenic agents. In this context, the cumulative use of zoledronic acid (ZL) is important duration medication-related risk factors for patients needing tooth extraction. In this preliminary study, we investigated the development of MRONJ-type lesions in C57Bl/6 male mice (ethical committee approval n°00262-2019) at the late time point of socket healing after continuous and cumulative administration of zoledronic acid (ZL) in 2 different dosages. Mice at age of 4 to 6 months were divided, receiving intraperitoneal injection of saline solution (SS, control, n = 3) or ZL at a concentration of 250ug/Kg (n = 3) and 500ug/Kg (n = 4) once a week, starting 4 weeks before extraction of the right upper incisor and continuing until the 30-day post-surgical period. At the end of 30 days, bone samples were collected for histological (H&E and Goldner Trichrome [GT] staining), histomorphometric and immunohistochemical analyzes for bone (Rankl), and inflammatory (COX-2) markers. While the control group exhibit a dental socket filled with mature bone, the ZL-250 and ZL-500 groups presented a significant and gradual increase in the area density of inflammatory infiltrate dependent of each dosage. Bone matrix area density was significantly lower in ZL-250 and ZL-500 groups, and GT staining revealed a gradual impairment in bone matrix in ZL-250 and ZL-500 groups compared to control (Figure 1). Area density of COX2+ cells, an inflammatory parameter positively related to bone neoformation, was also decreased in both treated groups compared to the control. On the other hand, area densities of fibroblasts, RANKL+ cells, and empty osteocyte lacunae were increased in ZL-250 and ZL-500 groups. Interestingly, the area density of detached osteoclasts, a typical characteristic of ZL effect, was higher in the ZL250 group compared to the SS, while ZL500 group presented a lower density of attached osteoclasts compared to the others. In summary and despite of limitations of the present study, the final outcome of alveolar bone healing was significantly affected in C57BL/6 mice using both dosages used in this study, but with increasing effects on histological MRONJ parameters at the higher dosage.