Preconditioning of Mesenchymal Stem Cells with Non-Toxic Concentration of Hydrogen Peroxide Against Oxidative Stress Induced Cell Death: The Role of Hypoxia-Inducible Factor-1

Purpose: To investigate the protective effect of preconditioning with non-toxic dose of hydrogen peroxide (H2O2) as a possible cell signaling molecule, against cell death induced by toxic concentration of H2O2 or by serum deprivation in human Wharton's jelly-derived mesenchymal stem cells (HWJ-MSCs) and underlying mechanisms. Methods: HWJ-MSCs were isolated and identified using flow cytometry. After finding non-toxic concentration of H2O2, cells preconditioning was performed by H2O2 (20 mu M) for 12 h and cell tolerance against serum deprivation or toxic levels of H2O2 was assayed by MTT test. Effect of preconditioning on mRNA and protein expression of Akt-1, Bcl-2 and Bax were examined using reverse transcription polymerase chain reaction (RT-PCR) and western blotting respectively. Role of hypoxia-inducible factor (HIF)-1 alpha was explored in presence HIF-1 alpha inhibitor. Results: Preconditioning with 20 mu M H2O2 for 12 h was non-toxic and decreased cell death induced by oxidative stress and serum deprivation in MSC cultures. However, the increased tolerance reversed in the presence of inhibitor of HIF-1 alpha. By regards to RT-PCR and western blotting data, although expression of Akt-1, Bcl-2 and Bax was not change considerably but phosphorylated Akt-1 (pAkt-1) was up regulated after treatment with 20 mu M H2O2 compared to control group. Moreover after exposure to 100 mu M H2O2, western blotting analysis showed that cell pretreatment with 20 mu M H2O2, decremented Bax/Bcl2 ratio and up-regulated HIF-1 alpha and pAkt-1 compared to the control group. Conclusion: Increased tolerance of H2O2-pretreated cells led to the suggestion that transplantation of H2O2 preconditioned MSCs may improve therapeutic potential of stem cells in cell therapy procedures.