Variation in lipid mediator and cytokine levels during mouse femur fracture healing.

Fracture healing is regulated by a variety of inflammatory mediators and growth factors which act over time to regenerate the injured tissue. This study used a mouse femur fracture model to quantify the temporal expression pattern of lipid mediators, cytokines, and related mRNAs during healing. Cyclooxygenase (COX-1 and COX-2) and 5-lipoxygenase (5-LO) derived lipid mediators, cytokines, and mRNA levels were quantified using mass spectrometry (LC-MS/MS), bead-based multiplex assays (xMAP), and quantitative PCR of cDNA (RTqPCR), respectively. Our analysis found that, the early inflammatory response (between 0 and 4 days after fracture) in the mouse femur fracture model coincided with elevated levels of COX-derived lipid mediators and inflammatory cytokines but with decreased levels of 5-LO-derived lipid mediators. Further, the COX-derived lipid mediators remained elevated for at least 7 days after fracture, suggesting that the COX-derived lipid mediators have additional functions during later phases of the fracture healing response. Differences were also found between mRNA levels and corresponding cytokines and lipid mediator levels, supporting a role for post-transcriptional regulation of gene expression. The temporal changes in fracture callus lipid mediator levels and inflammatory cytokines support a general positive role for inflammatory cytokines and COX-derived lipid mediators on fracture healing and a general negative role for 5-lipoxygenase derived lipid mediators during the initial stages of repair. © 2016 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 34:1883-1893, 2016.