The Effect Of Hypoxia Preconditioning On The Proliferation And Differentiation Potentials Of Human Urine-derived Stem Cells

Objective:To study the effect of hypoxia preconditioning on the proliferation and differentiation potentials of human urine-derived stem cellsMethods:Collecting fresh clean urine samples from health adult and hUSCs were isolated from the urine,cultured with mixed medium of KSFM and EFM.The cell surface markers?CD29,CD44,CD54,CD73,CD90,CD105?were analyzed by flow cytometry,then hUSCs were cultured under normoxia?20%O₂?or hypoxia(3%O_2,24h/48h),after that,observing the cell morphology by inverted microscope,the growth rate was examined by cck-8 assay and Edu assay.The osteogenic and adipogenic potential of the hUSCs were identified with ALP expression,Alizarin red,Von kassa staining,oil red and Semi-quantitative analysis.Results:1.Flow cytometry analysis showed that the cell have high expression of the typical mesenchymal stem cell surface markers CD44?CD73?CD90?CD105,2.no change in cell morphology between the three groups,3.the cck-8 assay showed that the growth curve of hUSCs was the s-shaped,and the hypoxia group?both 24h and 48h?has higher cell proliferation ability than normoxia group,4.Edu assay showed the same result,the hypoxia group?both 24h and 48h?have high expression of DNA synthesis rate,5.After being cultured with osteogenic and adipogenic medium,both group differentiated into osteogenic and adipogenic lineage.And the results illustrate that hypoxia preconditioning increased osteogenic differentiation of hUSCs,and maintain their adipogenic differentiation,ALP staining and quantitative detection,Alizarin red,Von kassa staining,Semi-quantitative analysis showed significantly inhibited cell osteogenic differentiation in hypoxia group?both 24h and 48h?,6.there is little difference between the three groups of oil red o staining.Conclusion:Hypoxia can promote the proliferation and osteogenic differentiation of hUSCs,maintain their multi-directional differentiation potential.