Osteoclast depletion with clodronate liposomes delays fracture healing in mice

Abstract

Osteoclasts are abundant within the fracture callus and also localize at the chondro-osseous junction. However, osteoclast functions during fracture healing are not well defined. Inhibition of osteoclast formation or resorptive activity impairs callus remodeling but does not prevent callus formation. Interestingly, though anti-osteoclast therapies differentially affect resolution of callus cartilage into bone. Treatments that inhibit osteoclast formation or viability tend to impair callus cartilage resolution, while treatments that target inhibition of bone resorption generally do not affect callus cartilage resolution. Here, we tested whether depletion of osteoclasts by systemic treatment with clodronate liposomes would similarly impair callus cartilage resolution. ICR mice were treated by intraperitoneal injections of clodronate-laden liposomes or control liposomes and subjected to closed femur fracture. Femurs were resected at multiple times after fracture and analyzed by radiography, histology, and mechanical testing to determine effects on healing. Clodronate liposome treatment did not prevent callus formation. However, radiographic scoring indicated that clodronate liposome treatment impaired healing. Clodronate liposome treatment significantly reduced callus osteoclast populations and delayed resolution of callus cartilage. Consistent with continued presence of callus cartilage, torsional mechanical testing found significant decreases in callus material properties after 28 days of healing. The results support a role for osteoclasts in the resolution of callus cartilage into bone. Whether the cartilage resolution role for osteoclasts is limited to simply resorbing cartilage at the chondro-osseous junction or in promoting bone formation at the chondro-osseous junction through another mechanism, perhaps similar to the reversal process in bone remodeling, will require further experimentation. © 2016 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 35:1699-1706, 2017.

Clodronate-Liposome Mediated Macrophage Depletion Abrogates Multiple Myeloma Tumor Establishment In Vivo

“…injection of clo-lip or a PBS-lip control, and the extent of macrophage depletion was investigated. As clo-lip administration is known to induce apoptosis of all phagocytic cells, including osteoclasts [24], zoledronate was administered to a third group of mice as an osteoclast-depletion control [33]. Flow cytometric analysis revealed a 70% reduction in the total CD11b + F4/80 + monocyte/macrophage population within the BM of clo-lip–treated animals compared with controls (Figure 1 A ).…”

“…Encapsulation of clodronate in lipid vesicles specifically targets clodronate to phagocytic macrophages that engulf and degrade the liposome, leading to clodronate accumulation and subsequent cellular apoptosis [23]. Unlike free clodronate, clodronate-liposomes (clo-lip) globally deplete macrophages and other phagocytic cells [23], [24].…”

Tiludronate and clodronate do not affect bone structure or remodeling kinetics over a 60 day randomized trial

“…However, reports regarding effects of clodronate are conflicting, including no changes in bone mineral density in a callus [ 11 ], 30% increase in bone mineral density [ 10 ], increased calcium content within the callus [ 12 ], and decreased healing callus strength [ 13 ]. Recently, it has been shown that osteoclasts are a necessary component of efficient endochondral ossification during fracture repair in mice [ 14 ]; therefore, it is possible that clodronate may impair normal bone healing in horses, which calls for careful consideration when used in a clinical setting.…”

Adjuvant Drug-Assisted Bone Healing: Advances and Challenges in Drug Delivery Approaches

“…In contrast, liposomal drug delivery can also be detrimental on bone healing as shown by Lin and O'Connor. The authors detected impaired fracture healing of femoral defects after treatment with clodronate-encapsulated liposomes ( Figure 2) over an investigation period of 28 days [166]. Liposomes or micelles are suitable site-directed drug carriers, particularly for water-insoluble drugs due to surface-decorated bone-targeting moieties.…”