Modulation of Macrophage Activity During Fracture Repair Has Differential Effects in Young Adult and Elderly Mice

Abstract

Objectives Advanced age is a factor associated with altered fracture healing. Delays in healing may increase the incidence of complications in the elderly, who are less able to tolerate long periods of immobilization and activity restrictions. The following study sought to determine if fracture repair could be enhanced in elderly animals by inhibiting macrophage activation, blocking the M-CSF receptor c-fms, and inhibiting monocyte trafficking using CC chemokine receptor-2 (CCR2) knockout mice. Methods Closed, unstable tibial shaft fractures were produced in mice aged four, 12 and 78 weeks. Mice were then fed a diet containing PLX3397 or a control diet from days 1–10 post-injury. Fractures were similarly made in CCR2−/− mice aged 78 weeks. The fracture callus was collected during fracture healing and was assessed for its size and the presence of macrophages, both of which were evaluated using the Mann-Whitney U test. Results PLX3397 treatment resulted in a decrease in the number of macrophages in the fracture callus at Day 5. Calluses in juvenile mice trended towards being smaller compared to elderly mice (p=0.08). There was also a trend toward larger callus size and increased bone formation in PLX3397-treated elderly animals compared to those of the control animals (p=0.12). Similar increases in bone formation (p=0.013) and decreases in cartilage within the callus (p=0.03) were seen at Day 10 in CCR2−/− mice. Conclusions The inhibition of macrophages in elderly mice may lead to an acceleration of fracture healing. Altering macrophage activation after fracture may represent a therapeutic strategy for preventing delayed healing and nonunion in the elderly.

Macrophage-based therapeutic strategies in regenerative medicine

“…Similarly, another study reported that macrophages in aged mice secreted lower levels of pro-angiogenic factors than macrophages from young mice, resulting in impaired wound healing [125]. In addition, inhibiting M-CSF receptor kinase activity reduced macrophage numbers and induced a trend of increased bone formation in old but not young mice [126]. These data indicate that the function of macrophages may change with age, and thus age and other contextual factors should be considered when designing macrophage-targeted approaches.…”

Macrophages: Their Emerging Roles in Bone

“…Consequently, cautious but optimistic progression of this therapeutic avenue is justified. Additional remaining questions include whether perturbations in macrophage function and/or subset dominance underlie impaired fracture healing or systemic bone disease, including bone fragility associated with chronic inflammation, infection, anti‐inflammatory treatment, and age …”

Tissue engineering strategies for promoting vascularized bone regeneration

“…179 The recruitment of inflammatory cells to sites of injury is mediated by four different subfamilies of chemokines (CC, CXC, CX3C and C) based on their biochemical, structural and functional properties 180 . In a Ccr2 −/− mouse model of decreased macrophage recruitment, it was shown that disruption of Ccr2 signaling impaired vascularization, decreased formation of callus, and delayed maturation of cartilage and bone remodeling up to 21 days after injury 181,182 . M2 macrophages contribute to the resolution of inflammation and are important to the progression bone healing.…”

“…Dysregulation of macrophage polarization, specifically persistence of the M1 phenotype or dampened M2 response, may contribute to age related delays in bone healing. 180–182 …”