Objective The present study was designed to examine 1. whether hypoxic preconditioning (HP) increases the migration potential of mesenchymal stem cells (MSC) and 2. the mechanistic basis for this effect.
Methods MSCs derived from green fluorescent protein (GFP)-transgenic mice were cultured under either normoxia (N-MSC) or hypoxia (0.5% O2) (HP-MSC) for 24 h. Wound healing and transwell assays were performed to assess cell migration in vitro. Potassium channel expression and FAK phosphorylation were analysed by western blot analysis, whole cell patch clamp recording was used to measure delayed rectifier K+ currents. Interactions between the Kv2.1 potassium channel and FAK were analysed via immunoprecipitation. N-MSC or HP-MSC were intravenously injected via tail vein in myocardial rat, and migration of MSC was assessed in vivo.
Results Both wound healing and transwell assays showed that, compared with controls, shRNA-mediated knock-down of the Kv2.1 potassium channel significantly reduced the ability of N-MSC to migrate. While exposure to hypoxia enhanced MSC migration both in vitro and in vivo, this effect was inhibited in presence of either tetra-ethylammonium (TEA) or elevated extracellular potassium. The potential effects of hypoxia exposure on MSC migration were significantly reduced by Kv2.1 knockdown. Hypoxic pre-conditioning significantly increased the expression of Kv2.1. Similarly, compared to normoxic controls, delayed rectifier K+ currents in HP-MSC were much greater. Lastly, HP-mediated increases in phosphorylation of FAK576/577 and FAK397 were neutralised by either addition of TEA or elevation of extra-cellular potassium levels.
Conclusion These findings demonstrate that KV2.1 plays an important role in the regulation of MSC migration. HP enhances the ability of MSC to migrate both in vitro and in vivo, and this effect may be mediated, at least in part, through activation of Kv2.1 potassium channels and the FAK pathway.