Abstract
Low intensity pulsed ultrasound (LIPUS) has been proven effective in promoting fracture healing but the underlying mechanisms are not fully depicted. We examined the effect of LIPUS on the recruitment of mesenchymal stem cells (MSCs) and the pivotal role of stromal cell-derived factor-1/C-X-C chemokine receptor type 4 (SDF-1/CXCR4) pathway in response to LIPUS stimulation, which are essential factors in bone fracture healing. For in vitro study, isolated rat MSCs were divided into control or LIPUS group. LIPUS treatment was given 20 minutes/day at 37°C for 3 days. Control group received sham LIPUS treatment. After treatment, intracellular CXCR4 mRNA, SDF-1 mRNA and secreted SDF-1 protein levels were quantified, and MSCs migration was evaluated with or without blocking SDF-1/CXCR4 pathway by AMD3100. For in vivo study, fractured 8-week-old young rats received intracardiac administration of MSCs were assigned to LIPUS treatment, LIPUS+AMD3100 treatment or vehicle control group. The migration of transplanted MSC to the fracture site was investigated by ex vivo fluorescent imaging. SDF-1 protein levels at fracture site and in serum were examined. Fracture healing parameters, including callus morphology, micro-architecture of the callus and biomechanical properties of the healing bone were investigated. The in vitro results showed that LIPUS upregulated SDF-1 and CXCR4 expressions in MSCs, and elevated SDF-1 protein level in the conditioned medium. MSCs migration was promoted by LIPUS and partially inhibited by AMD3100. In vivo study demonstrated that LIPUS promoted MSCs migration to the fracture site, which was associated with an increase of local and serum SDF-1 level, the changes in callus formation, and the improvement of callus microarchitecture and mechanical properties; whereas the blockade of SDF-1/CXCR4 signaling attenuated the LIPUS effects on the fractured bones. These results suggested SDF-1 mediated MSCs migration might be one of the crucial mechanisms through which LIPUS exerted influence on fracture healing.
Highlights
Millions of fractures occur annually as a result of traumatic injuries or pathological conditions
The interactions between cytokines and Mesenchymal stem cells (MSCs) in bone repair remain controversial, many studies found that MSCs expressed both SDF1 and CXCR4 genes [9,10,11], and stromal cell-derived factor-1 (SDF-1)/CXCR4 signaling is critical for the recruitment of MSCs to the fracture site during fracture healing
The present study demonstrates that the enhanced MSCs migration mediated by SDF-1/CXCR4 pathway may be one of the crucial mechanisms through which Low intensity pulsed ultrasound (LIPUS) promotes fracture healing
Summary
Millions of fractures occur annually as a result of traumatic injuries or pathological conditions. It is widely accepted that MSCs are normally retained in the special niches of different adult tissues In stressful situations such as injury, when there is a need for tissue repair and to maintain tissue homeostasis, MSCs can be recruited to the site of injury and contribute to the repair process. This process will trigger the inflammatory cascade [6] In this early inflammatory phase of fracture healing, many types of cytokines, such as interleukin 6 (IL-6) and stromal cell-derived factor-1 (SDF-1), released from the damaged bone facilitate the egress of MSCs from the periosteum and bone marrow into the blood stream, which rapidly accumulate and engraft at fracture site, and initiate bone regeneration process [3,7,8]. Kitaori et al showed that SDF-1 level was elevated in the periosteum of injured bone, which recruited MSCs homing to the graft bone at the fracture site and promoted endochondral bone formation [8]
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