Hypoxia Preconditioning Promotes Survival And Clonogenic Capacity Of Human Umbilical Cord Blood Mesenchymal Stem Cells

Background: In recent decade, human umbilical cord blood derived mesenchymal stem cells (hUCB-MSCs) provide enormous potential for appropriate cell therapy, but they have limited growth potential and cease to proliferate due to cellular senescence, so providing a strategy for increasing the stem cell survival is necessary.  Methods: In this investigation, MSCs characterized by flow cytometry were isolated from umbilical cord blood. Hypoxia preconditioning (HPC) was induced in a water-saturated gas mixture of 2.5% O2 and 5% CO2 for 15 min and then reoxygenation at 21% O2 for 30 min at 37 °C.  Subsequently, hypoxia preconditioned hUCB-MSCs were exposed to 2.5% O2 and 5% CO2 for 24, 48 and 72 hr (HPC + hypoxia groups). We examined the proliferation capacity of hUCB-MSCs after HPC in comparison with normoxia status, and we determined the best duration time of being under hypoxia (24, 48 or 72 hr of hypoxia). In order to assess the role of HPC on the expression of surface markers, cells were analyzed by flow cytometry. Proliferation of cells was evaluated using MTT assay, and doubling time and colony-forming unit-fibroblast (CFU-F) was calculated in each group.  Results: The MTT results showed that cell viability of HPC-UCB-MSCs significantly increased in comparison with UCB-MSCs under normoxia condition. Our study revealed that HPC reduces the doubling time of UCB-MSCs remarkably after passaging 48 hours of hypoxia. Our results proved that HPC can significantly increase the CFU-F colony numbers of hUCB-MSCs without any alteration on cell surface marker expression.   Conclusion: Our results suggested that HPC of umbilical cord blood derived mesenchymal stem cells along with induction of hypoxia can provide a suitable culture condition for rapid proliferation of mesenchymal stem cells with no effect on their immunophenotype features and it could be a potential therapeutic option for increasing the capability of MSCs.