Wound healing is a complex process consisting of a series of overlapping events which include inflammation, matrix deposition and remodelling. Any perturbations in this process can result in delayed or incomplete healing leading to chronic wounds such as diabetic ulcers. These wounds often fail to heal, due to a stalling of the process in the inflammatory phase, leading to increased cytokine production and breakdown of the extracellular matrix. Macrophages (M1) initially promote the inflammatory response but then switch their phenotype to an anti-inflammatory state (M2) that helps lead to a resolution of inflammation. Toll-like receptors (TLRs) are part of the innate immune system and, upon injury, initiate an immune response to remove pathogens and cellular debris from the wound site. TLR4 is expressed on macrophages and is integral in the initiation of this inflammatory response. TLR4 is activated by LPS and signals intracellularly, via the MyD88 pathway, to promote inflammation via the release of TNF. Flightless I (Flii) can influence TLR4 signalling both extracellularly, by sequestering LPS to TLR4 receptor activation and intracellularly via its inhibition of MyD88 signalling. To further investigate the role of Flii during inflammation we have used in vitro and in vivo approaches in conjunction with Flii genetic mice expressing low (Flii+/-), normal (WT) or high (FliiTg/Tg) levels of Flii. Wounds were created on the mice and the effect of Flii on inflammatory responses, TLR signalling and cytokine production determined. Macrophages collected by intraperitoneal lavage from these mice were assessed for cytokine production by bead array. Our studies showed that increased Flii prolongs inflammation, increases TNF production, decreases VEGF, delays the appearance of M2 macrophages and impairs healing. These studies illustrate that Flii has important functions in wound healing and inflammation and reducing its activity can lead to improved wound outcomes.