Study On Expression Of Matrix Metalloproteinase And Effect Of Cell Migration On Rat Mesenchymal Stem Cells By Cytokines

Objective: To investigate the influence of matrix metalloproteinase expression and cell migration on rat mesenchymal stem cell by erythropoietin combination with granulocyte colony stimulating factor.Methods: MSC culture was performed with the classical adhering method; characteristic of MSC was identified through multi-differentiation and surface marker assay(CD90,CD29,CD34,CD45,CD44).P3MSCs were treated with different concentrations of EPO ,G-CSF and EPO combination with G-CSF. The expression of MMP and TIMP mRNA was detected by RT-PCR. The ability of cell migration was evaluated by Transwell and Wound healing Assay. Western blot assay was used to detect the changes of ERK1/2 protein.Results:①P3MSC were positive for CD90,CD29 and negative for CD34,CD45,CD44. MSC were capable of osteogenic and adipogenic differentiation.②MSCs were treated with various concentrations of EPO for 12h, the expression of MMP-2 mRNA was increased compared with the control group and this result was highest in 1IU/ml(P<0. 05).The expression of MMP-9, TIMP-1 and TIMP-2 mRNA was no difference significantly.③MSCs were treated with various concentrations of G-CSF for 12h, the expression of MMP-2 mRNA was increased compared with the control group and this result was close to the peak in 1ng/ml(P<0. 05).The expression of MMP-9, TIMP-1 and TIMP-2 mRNA was no difference significantly.④MSCs were treated with various concentrations of EPO(1IU/ml),G-CSF(0.1ng/ml,1ng/ml)and EPO(1IU/ml)+G-CSF(0.1ng/ml,1ng/ml)for 12h, the expression of MMP-2 mRNA was increased. The expression of MMP-2 mRNA was strongest in the EPO(1 IU/ml) combination with G-CSF(0.1ng/ml) treatment group when compared with the control group and single factor group. The expression of MMP-9, TIMP-1 and TIMP-2 mRNA was no difference significantly.⑤MSCs were treated with various concentrations of EPO(1IU/ml),G-CSF(0.1ng/ml,1ng/ml)and EPO(1IU/ml)+G-CSF(0.1ng/ml,1ng/ml)for 12h, the numbers of MSCs invasion was increased. The numbers of cell invasion was maximum in the EPO(1 IU/ml) combination with G-CSF(0.1ng/ml) treatment group when compared with the control group and single factor group.⑥MSCs were treated with various concentrations of EPO(1IU/ml),G-CSF(0.1ng/ml,1ng/ml)and EPO(1IU/ml)+G-CSF(0.1ng/ml,1ng/ml)for 18h, the areas of MSC migration was increased. The areas of cell migration was obvious in the EPO(1 IU/ml) combination with G-CSF(0.1ng/ml) treatment group when compared with the control group and single factor group.⑦MSCs were treated with various concentrations of EPO(1IU/ml),G-CSF(0.1ng/ml,1ng/ml)and EPO(1IU/ml)+G-CSF(0.1ng/ml,1ng/ml)for 3h, ERK1/2 protein was increased. The expression of ERK1/2 protein was strongest in the EPO(1IU/ml) combination with G-CSF(0.1ng/ml) treatment group when compared with the control group and single factor group.Conclusion: EPO combination with G-CSF treatment could promote cell migration by stimulating the expression of MMP-2 on MSC,and probably related to ERK1/2 signal pathway.