Tara Aghaloo

American Association of Oral and Maxillofacial Surgeons position paper on medication-related osteonecrosis of the jaw--2014 update.

Strategies for management of patients with, or at risk for, medication-related osteonecrosis of the jaw (MRONJ) were set forth in the American Association of Oral and Maxillofacial Surgeons (AAOMS) position papers in 2007 and 2009. The position papers were developed by a special committee appointed by the board and composed of clinicians with extensive experience in caring for these patients and basic science researchers. The knowledge base and experience in addressing MRONJ has expanded, necessitating modifications and refinements to the previous position paper. This special committee met in September 2013 to appraise the current literature and revise the guidelines as indicated to reflect current knowledge in this field. This update contains revisions to diagnosis, staging, and management strategies and highlights current research status. The AAOMS considers it vitally important that this information be disseminated to other relevant health care professionals and organizations.

Osteonecrosis of the Jaw in a Patient on Denosumab

Osteonecrosis of the jaws (ONJ) clinically presents as exposed, necrotic bone in the maxilla or mandible of at least eight week duration, with or without the presence of pain, infection, or previous trauma in a patient who has not received radiation to the jaws (1–3). Although necrotic bone exposure has been reported in the jaws of a variety of patients not receiving bisphosphonates (BPs;4–9), the number of BP related ONJ cases has continued to steadily increase since the first report in 2003 (10). To date, a direct causal relationship between BP use and ONJ has not been established (11,12). However, many retrospective and prospective analyses have identified cases of ONJ where BP therapy, especially the more potent intravenous preparations, was the only consistent variable, strongly suggesting that BPs play a significant role in ONJ pathophysiology (13–24). Potential mechanisms underlying bisphosphonate related osteonecrosis of the jaws (BRONJ) pathophysiology have generated great debate in the literature (25,26). It is not surprising that many hypotheses attempt to explain the unique localization of BRONJ exclusively to the jaws, including altered bone remodeling, angiogenesis inhibition, constant microtrauma, soft tissue BP toxicity, and bacterial infection (15,18,25,27–29). Importantly, ONJ incidence correlation with BP potency suggests that inhibition of osteoclast function and differentiation might be a key factor in the pathophysiology of the disease. Currently other inhibitors of osteoclast differentiation and function are entering the pharmacologic armamentarium for the treatment of diseases with increased bone turnover. The association of these new therapies with ONJ is uncertain. We report a case of ONJ in a patient receiving Denosumab, a human RANKL monoclonal antibody currently in clinical trials for the treatment of osteoporosis, primary and metastatic bone cancer, giant cell tumor, and rheumatoid arthritis (30–33).

Periodontal disease and bisphosphonates induce osteonecrosis of the jaws in the rat

Bisphosphonates (BPs) are medications used commonly to treat primary and metastatic bone cancer, as well as osteoporosis. Although BPs improve bone mineral density, reduce fracture risk, and reduce hypercalcemia of malignancy, some patients develop BP‐related osteonecrosis of the jaws (BRONJ). This devastating complication is defined as clinically exposed bone in the maxillofacial region for more than 8 weeks. Despite an increasing number of BRONJ cases since first reported, the disease pathophysiology remains largely unknown. Since published studies suggest a significant role for dental disease in the pathophysiology of BRONJ, we developed a BRONJ animal model where aggressive periodontal disease is induced by ligature placement around the crown of the right maxillary first molar in the presence of vehicle (veh) or zoledronic acid (ZA), a potent BP. Ligature placement induced significant alveolar bone loss, which was attenuated by ZA treatment. Osteonecrosis was observed associated with ligature‐induced periodontitis in the ZA‐treated group. This was seen as sequestration and extensive periosteal alveolar bone formation on micro–computed tomography (µCT) in the ligated site of BP‐treated animals. Histologic examination confirmed these findings, seen as necrotic bone with diffuse loss of osteocytes and empty lacunae, rimming of the necrotic bone by squamous epithelium and inflammation, and exposure to the oral cavity. Importantly, the rat lesions were strikingly similar to those of BRONJ patients. Our data suggest that dental disease and potent BP therapy are sufficient for BRONJ development in the rat.

Osteogenic Potential of Mandibular vs. Long-bone Marrow Stromal Cells

Although fundamentally similar to other bones, the jaws demonstrate discrete responses to developmental, mechanical, and homeostatic regulatory signals. Here, we hypothesized that rat mandible vs. long-bone marrow-derived cells possess different osteogenic potential. We established a protocol for rat mandible and long-bone marrow stromal cell (BMSC) isolation and culture. Mandible BMSC cultures formed more colonies, suggesting an increased CFU-F population. Both mandible and long-bone BMSCs differentiated into osteoblasts. However, mandible BMSCs demonstrated augmented alkaline phosphatase activity, mineralization, and osteoblast gene expression. Importantly, upon implantation into nude mice, mandible BMSCs formed 70% larger bone nodules containing three-fold more mineralized bone compared with long-bone BMSCs. Analysis of these data demonstrates an increased osteogenic potential and augmented capacity of mandible BMSCs to induce bone formation in vitro and in vivo. Our findings support differences in the mechanisms underlying mandible homeostasis and the pathophysiology of diseases unique to the jaws.

Elevated Vascular Endothelial Growth Factor in Keloids: Relevance to Tissue Fibrosis

Excessive scar or keloid shares common features of a benign dermal growth. Yet, in contrast to malignant tumor, a keloid does not expand beyond the dermis. What triggers the continuing growth of a benign lesion? Deficient or overabundant levels of vascular endothelial growth factor have been reported to contribute to impaired or excessive wound healing. Although numerous studies have examined the pathophysiology of impaired wounds, little information has been provided on mechanisms of exuberant healing. The molecular basis of keloid formation is governed by the interplay of cellular signaling pathways, specific target gene activation, and the nature of the microenvironment. Recent works have demonstrated an accumulation of hypoxia-inducible factor-1α protein in freshly biopsied keloid tissues, thus providing first evidence that a local state of hypoxia exists in keloids. Our findings and the findings of others support at least two plausible mechanisms implicated in the development of fibrotic wounds, a state of ongoing fibroplasia or inflammation and an excessive accumulation of extracellular matrix. This article will review recent works examining the potential role of vascular endothelial growth factor in keloid pathogenesis with particular focus on its involvement in the two proposed pathological processes, a prolonged inflammation and an altered balance in extracellular matrix metabolism.

Periapical disease and bisphosphonates induce osteonecrosis of the jaws in mice.

Osteonecrosis of the jaw (ONJ) is a well‐recognized complication of antiresorptive medications, such as bisphosphonates (BPs). Although ONJ is most common after tooth extractions in patients receiving high‐dose BPs, many patients do not experience oral trauma. Animal models using tooth extractions and high BP doses recapitulate several clinical, radiographic, and histologic findings of ONJ. We and others have reported on rat models of ONJ using experimental dental disease in the absence of tooth extraction. These models emphasize the importance of dental infection/inflammation for ONJ development. Here, we extend our original report in the rat, and present a mouse model of ONJ in the presence of dental disease. Mice were injected with high dose zoledronic acid and pulpal exposure of mandibular molars was performed to induce periapical disease. After 8 weeks, quantitative and qualitative radiographic and histologic analyses of mouse mandibles were done. Periapical lesions were larger in vehicle‐treated versus BP‐treated mice. Importantly, radiographic features resembling clinical ONJ, including thickening of the lamina dura, periosteal bone deposition, and increased trabecular density, were seen in the drilled site of BP‐treated animals. Histologically, osteonecrosis, periosteal thickening, periosteal bone apposition, epithelial migration, and bone exposure were present in the BP‐treated animals in the presence of periapical disease. No difference in tartrate‐resistant acid phosphatase (TRAP)+ cell numbers was observed, but round, detached, and removed from the bone surface cells were present in BP‐treated animals. Although 88% of the BP‐treated animals showed areas of osteonecrosis in the dental disease site, only 33% developed bone exposure, suggesting that osteonecrosis precedes bone exposure. Our data further emphasize the importance of dental disease in ONJ development, provide qualitative and quantitative measures of ONJ, and present a novel mouse ONJ model in the absence of tooth extraction that should be useful in further exploring ONJ pathophysiological mechanisms.

Adverse Effects of Hyperlipidemia on Bone Regeneration and Strength

Hyperlipidemia increases the risk for generation of lipid oxidation products, which accumulate in the subendothelial spaces of vasculature and bone. Atherogenic high‐fat diets increase serum levels of oxidized lipids, which are known to attenuate osteogenesis in culture and to promote bone loss in mice. In this study, we investigated whether oxidized lipids affect bone regeneration and mechanical strength. Wild‐type (WT) and hyperlipidemic (Ldlr−/−) mice were placed on a high‐fat (HF) diet for 13 weeks. Bilateral cranial defects were introduced on each side of the sagittal suture, and 5 weeks postsurgery on the respective diets, the repair/regeneration of cranial bones and mechanical properties of femoral bones were assessed. MicroCT and histological analyses demonstrated that bone regeneration was significantly impaired by the HF diet in WT and Ldlr−/− mice. In femoral bone, cortical bone volume fraction (bone volume [BV]/tissue volume [TV]) was significantly reduced, whereas cortical porosity was increased by the HF diet in Ldlr−/− but not in WT mice. Femoral bone strength and stiffness, measured by three‐point bending analysis, were significantly reduced by the HF diet in Ldlr−/−, but not in WT mice. Serum analysis showed that the HF diet significantly increased levels of parathyroid hormone, tumor necrosis factor (TNF)‐α, calcium, and phosphorus, whereas it reduced procollagen type I N‐terminal propeptide, a serum marker of bone formation, in Ldlr−/−, but not in WT mice. The serum level of carboxyl‐terminal collagen crosslinks, a marker for bone resorption, was also 1.7‐fold greater in Ldlr−/− mice. These findings suggest that hyperlipidemia induces secondary hyperparathyroidism and impairs bone regeneration and mechanical strength.

The effect of NELL1 and bone morphogenetic protein-2 on calvarial bone regeneration.

PURPOSE Most craniofacial birth defects contain skeletal components that require bone grafting. Although many growth factors have shown potential for use in bone regeneration, bone morphogenetic proteins (BMPs) are the most osteoinductive. However, supraphysiologic doses, high cost, and potential adverse effects stimulate clinicians and researchers to identify complementary molecules that allow a reduction in dose of BMP-2. Because NELL1 plays a key role as a regulator of craniofacial skeletal morphogenesis, especially in committed chondrogenic and osteogenic differentiation, and a previous synergistic mechanism has been identified, NELL1 is an ideal molecule for combination with BMP-2 in calvarial defect regeneration. We investigated the effect of NELL1 and BMP-2 on bone regeneration in vivo. MATERIALS AND METHODS BMP-2 doses of 589 and 1,178 ng were grafted into 5-mm critical-sized rat calvarial defects, as compared with 589 ng of NELL1 plus 589 ng of BMP-2 and 1,178 ng of NELL1 plus 1,178 ng of BMP-2, and bone regeneration was analyzed. RESULTS Live micro-computed tomography data showed increased bone formation throughout 4 to 8 weeks in all groups but a significant improvement when the lower doses of each molecule were combined. High-resolution micro-computed tomography and histology showed more mature and complete defect healing when the combination of NELL1 plus BMP-2 was compared with BMP-2 alone at lower doses. CONCLUSION The observed potential synergy has significant value in the future treatment of patients with craniofacial defects requiring extensive bone grafting that would normally entail extraoral autogenous bone grafts or doses of BMP-2 in milligrams.

Nell-1 Enhances Bone Regeneration in a Rat Critical-Sized Femoral Segmental Defect Model

Background: Effective regeneration of bone is critical for fracture repair and incorporation and healing of bone grafts used during orthopedic, dental, and craniofacial reconstructions. Nel-like molecule-1 (Nell-1) is a secreted protein identified from prematurely fused cranial sutures of craniosynostosis patients that has been found to specifically stimulate osteogenic cell differentiation and bone formation. To test the in vivo osteoinductive capacity of Nell-1, a critical-sized femoral segmental defect model in athymic rats was used. Methods: A 6-mm defect, which predictably leads to nonunion if left untreated, was created in the left femur of each rat. Three treatment groups (n = 8 each) were created consisting of rats treated with (1) 1.5 mg/ml Nell-1, (2) 0.6 mg/ml Nell-1, and (3) phosphate-buffered saline only as a Nell-free control. Phosphate-buffered saline or Nell-1 was mixed with demineralized bone matrix as a carrier before implantation. All animals were euthanized 12 weeks after surgery, and bone regeneration was evaluated using radiographic, three-dimensional micro–computed tomographic, and histologic analysis. Results: Both Nell-1–treated groups had significantly greater bone formation compared with the Nell-free group, with bone volume increasing with increasing Nell-1 concentration. Conclusions: Nell-1 in a demineralized bone matrix carrier can significantly improve bone regeneration in a critical-sized femoral segmental defect in a dose-dependent manner. The results of this study demonstrate that Nell-1 is a potent osteospecific growth factor that warrants further investigation. Results also support the potential application of Nell-1 as a bone graft substitute in multiple clinical scenarios involving repair of critical bone loss when autograft bone is limited or unavailable.

RANKL inhibitors induce osteonecrosis of the jaw in mice with periapical disease.

Antiresorptive medications are essential in treating diseases of pathologic osteoclastic bone resorption, including bone cancer and osteoporosis. Bisphosphonates (BPs) are the most commonly used antiresorptives in clinical practice. Although inhibition of bone resorption is important in regulating unwanted malignant and metabolic osteolysis, BP treatment is associated with potential side effects, including osteonecrosis of the jaws (ONJ). Recently, non‐BP antiresorptive medications targeting osteoclastic function and differentiation, such as denosumab, have entered the clinical arena. Denosumab treatment results in a similar rate of ONJ as BPs. Animal models of ONJ, using high‐dose BP treatment in combination with tooth extraction or dental disease, provide valuable tools and insight in exploring ONJ pathophysiology. However, the ability of other antiresorptives to induce ONJ‐like lesions in animal models has not been explored. Such studies would be beneficial in providing support for the role of osteoclast inhibition in ONJ pathogenesis versus a direct BP effect on oral tissues. Here, we tested the ability of the receptor activator of NF‐κB ligand (RANKL) inhibitors RANK‐Fc (composed of the extracellular domain of RANK fused to the fragment crystallizable [Fc] portion of immunoglobulin G [IgG]) and OPG‐Fc (composed of the RANKL‐binding domains of osteoprotegerin [OPG] linked to the Fc portion of IgG) to induce ONJ in mice in the presence of periapical disease, but in the absence of dental extractions. We demonstrate radiographic evidence of ONJ in RANK‐Fc–treated and OPG‐Fc–treated mice, including inhibition of bone loss, increased bone density, lamina dura thickening, and periosteal bone deposition. These findings closely resembled the radiographic appearance of an ONJ patient on denosumab treatment. Histologic examination revealed that RANK‐Fc treatment and OPG‐Fc treatment resulted in absence of osteoclasts, periosteal bone formation, empty osteocytic lacunae, osteonecrosis, and bone exposure. In conclusion, we have successfully induced ONJ in mice with periapical disease, using potent osteoclast inhibitors other than BPs. Our findings, coupled with ONJ animal models using high‐dose BPs, suggest that osteoclast inhibition is pivotal to the pathogenesis of ONJ.