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Promotion of Mesenchymal Stem Cell Migration and Proliferation by Low
Dose H2O2 Mediated through Activation of SDF-1/CXCR4 Axis
Chen Minjia1, Zhang Ya1, An Tianchen1, Qiu Wei1, Chen Kejin2, Du Juan1, Sun Jianhui1, Wen Dalin1, Jiang Jianxin1, Huang Hong1*
1State Key Laboratory of Trauma, Burn and Combined Injury, Institute of Surgery Research, Daping Hospital, Third Military Medical University, Chongqing 400042, China; 2Chongqing Three Gorges Medical College, Chongqing 400042, China
Abstract: This work was aim to investigate the effect of low dose of H2O2 on the migration and proliferation of bone marrow mesenchymal stem cells (BMSCs)， and its mechanism in BMSCs. BMSCs isolated from mice by adherence of time interval difference were treated with different low dose of H2O2 for 24 h. The surface markers of BMSCs， such as CD34， CD45， CD29， CD44 and CXCR4， were identified by flow cytometry analysis. The experimental groups were divided by random digits method. The effect of H2O2 on the cell migration ability was detected by Transwell migration assay and scraping. The effect of H2O2 on cell proliferation was measured by 3-(4，5-dimethylthiazol-2-yl)-2，5-diphenyltetrazolium bromide (MTT) assay. The rate of BMSCs apoptosis was detected by flow cytometry analysis. The levels of SDF-1 and its receptor CXCR4， aging-related proteins and key proteins in the PI3K/Akt/mTOR pathway were detected by Western blot. The results from flow cytometry analysis showed that BMSCs highly expressed CD29 and CD44， very lowly expressed CD34 and CD45 on their cell surfaces. Low dose of H2O2 (25 and 50 μmol/L) effectively promoted BMSCs proliferation (P<0.05). The apoptosis rates of BMSCs in 25， 50 or 100 μmol/L H2O2 groups were no significant differences compared with the control. However， the rates of BMSCs apoptosis in 150 or 200 μmol/L H2O2 groups were significantly increased compared with the control (P<0.01). The results of Transwell migration assay and scraping showed that 50 μmol/L H2O2 significantly augmented the migration ability of BMSCs (P<0.05)， which could be inhibited by CXCR4 antibody. In addition， we found that 25 or 50 μmol/L H2O2 downregulated the levels of aging-related protein p16 (P<0.05， P<0.01)， inversely， upregulated the levels of pro-cell cycle protein CyclinD1 (P<0.05， P<0.01). Simultaneously， 25-100 μmol/L H2O2 induced an obvious increase in the levels of SDF-1 and its receptor CXCR4， and its downstream key proteins phosphorylated， such as PI3K， AKT and mTOR. Our data suggested that low dose H2O2 could effectively promote BMSCs proliferation and migration， by which activated the SDF-1/CXCR4 signal and its downstream the PI3K/ Akt/mTOR pathway.
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