A novel in vitro model of ischaemic preconditioning: its effect on cell viability in human skeletal muscle and a possible explanation for the ‘second window of protection’

Abstract

BackgroundThe ischaemia–reperfusion (IR) injury causes significant morbidity. Ischaemic preconditioning (IPC) is a technique for limiting the effects of the IR injury. Its potential has not yet been harnessed in orthopaedics. AimsTo establish a novel in vitro IR model using a human skeletal muscle cell line. Secondly, to introduce simulated IPC to the model and examine the effect of this on cell viability. MethodsA human skeletal muscle cell line was cultured in vitro. Placing the cells in a hypoxic buffer and a closed hypoxic environment simulated ischaemia. Reversing this process simulated reperfusion. IPC was simulated by alternate cycles of ischaemia and reperfusion. Cell viability comparisons were made between control and experimental groups of cells. ResultsA reproducible in vitro IR model was established. The addition of simulated IPC is associated with increased cell death at 12 and 24h of reperfusion. Significantly greater cell survival is seen in the IPC group when measured at 72h reperfusion. ConclusionsWe hypothesise that IPC initially decreases cell number. The remaining cells are more robust. This selected cell line then expands over the course of 72h and displays greater resistance to the IR injury. This theory can help explain delayed preconditioning.