Molecular Mechanism of Ischaemic Preconditioning of Skeletal Muscle In Vitro.

Abstract

IntroductionIschaemic preconditioning (IPC) is a phenomenon whereby tissues develop an increased tolerance to ischaemia and subsequent reperfusion if first subjected to sublethal periods of ischaemia. Despite extensive investigation of IPC, the molecular mechanism remains largely unknown. Our aim was to show genetic changes that occur in skeletal muscle cells in response to IPC.MethodsWe established an in vitro model of IPC using a human skeletal muscle cell line. Gene expression of both control and preconditioned cells at various time points was determined. The genes examined were hypoxia-inducible factor-1 alpha (HIF-1 alpha), early growth response 1 (EGR1), JUN, and FOS. HIF-1 alpha is a marker of hypoxia. EGR1, JUN, and FOS are early response genes and may play a role in the protective responses induced by IPC.ResultsHIF-1 alpha was upregulated following one and two hours of simulated ischaemia (p = 0.076 and 0.841, respectively) verifying that hypoxic conditions were met using our model. Expression of EGR1 and FOS was upregulated and peaked after one hour of hypoxia (p = 0.001 and <0.00, respectively). cFOS was upregulated at two and three hours of hypoxia. IPC prior to simulated hypoxia resulted in a greater level of upregulation of EGR1, JUN and FOS genes (p = <0.00, 0.047, and <0.00 respectively).ConclusionThis study has supported the use of our hypoxic model for studying IPC in vitro. IPC results in a greater upregulation of protective genes in skeletal muscle cells exposed to hypoxia than in control cells. We have demonstrated hitherto unknown molecular mechanisms of IPC in cell culture.