Chondrocytes, the sole resident cell-type in cartilage are responsible for the maintenance of the extracellular matrix and as they do not divide it is vital to find a protective mechanism against impact trauma. A single impact load to a joint causes subchondral bone fracture resulting in permanent cartilage damage to the ECM, including surface fissures, loss of proteoglycan, cell death and pre-disposition to osteoarthritis (Bush et al., 2005, Scott & Athanasiou, 2006). Full-depth cartilage was excised from bovine metacarpal-phalangeal joints (18-24 months of age, obtained from a local abattoir) under aseptic conditions. Explants were incubated with 5μM calcein-AM and 1μM propidium iodide and impact experiments were performed by drop-tower with a single impact of 0.131J. Samples were incubated alone or in the presence of REV5901 (50 µM), Wortmannin (10nM) and Uridine (100µM). Cell viability was determined time points of 0, 5, 10, 20 & 30 mins post impact by confocal laser scanning microscopy (CLSM). Data for cell volume was determined in non-impacted samples and 30 min post impact. Western blot analysis were used to determine alterations in actin binding proteins. Protein bands were analysed by Image J software and changes in expression quantified. All data are expressed as Mean ± s.e.m. *P<0.05 vs. control, n = 4 determination in triplicate. Impact led to cell death which was significantly reduced by a pre-incubation with REV5901 from 10.92±1.02% to 5.44%±0.59% (p<0.001). Samples treated with Uridine caused a reduction of cell death that was significantly lower from the control samples 8.10±0.40% (p<0.05) but couldn’t reach the chondro-protective effect offered by REV5901 (p<0.01). In contrast, samples treated with Wortmannin increased cell death to 12.02±1.83%. Cell death was significantly reduced in samples treated in combination of Wortmanin and REV5901 8.58±0.54%. Data for volume changes showed significant decrease in volume for cell treated with Wortmannin 359.9±17.67μm3, compared to 716.136±37.81μm3 and 552.68±27.26 μm3 for control (p<0.001) and REV5901 (p<0.001). Nevertheless, cell volume for cells treated with Uridine significantly decreased in comparison to the control 608.3±24.32μm3 (p<0.05). In addition, when cells were treated with a combination of Wortmannin and REV5901 cell volume size was restored to 538.09±30.6μm3. Western blot analysis showed that only cofilin and gelsolin from the actin binding proteins were expressed in all the samples, no profilin or phospho-cofilin were detected. These data confirm the therapeutic opportunities offered by REV5901 and it’s chondro-protective properties in part due to the polymerisation of the actin cytoskeleton. We suggest that the pathway involved via the phosphoinositide 3-kinases (PI3Ks) could offer novel therapeutic opportunities for prevention of irreversible cartilage damage from acute impact trauma.