Recombinant CXCR4 gene lentivirus transfection of rat bone marrow mesenchymal stem cells in vitro / 中国组织工程研究

ABSTRACT

BACKGROUND:Chemokine receptor 4 (CXCR4) is the main homing regulating factor for mesenchymal stem cells in vivo. How to improve the self-expression of CXCR4 is crucial for regulating the homing ability of stem cells in vivo targeting organs. OBJECTIVE:To construct lentiviral vectors carrying CXCR4 or green fluorescent protein (GFP) gene, and to observe their effects on growth and secrete function of bone marrow mesenchymal stem cells. METHODS:Lentiviral vectors were used to over-express CXCR4 in rat bone marrow mesenchymal stem cells. GFP gene was transfected as a control. Transfection efficiency was analyzed using fluorescence microscopy, RT-PCR and western blot assay for detecting the expression of CXCR4 or GFP in cells at both the mRNA and protein levels. Effects of CXCR4 expression modified on the cellular proliferation of bone marrow mesenchymal stem cells were assayed with flow cytometry. Effect of CXCR4 regulation on secretion function of bone marrow mesenchymal stem cells was determined by protein analysis from cellsupernatant. RESULTS AND CONCLUSION:CXCR4 gene or GFP gene could be transfected into bone marrow mesenchymal stem cells safely and effectively with lentivirus transfection system. Analysis of CXCR4 expression at the mRNA and protein levels confirmed the transfection efficiency. Effects of CXCR4 over-expression on the cellular proliferation of bone marrow mesenchymal stem cells and effects of CXCR4 regulation on secretion of protective factors from bone marrow mesenchymal stem cells were improved. The proliferation ability of bone marrow mesenchymal stem cells with CXCR4 over-expression was improved, which was approved with flow cytometry (P<0.05). And it was helpful for bone marrow mesenchymal stem cells to secrete some protective factors. Many protein and soluble factors were at higher levels, and those were good at cells proliferation and immunoloregulation in bone marrow mesenchymal stem cells (P<0.05). The method of lentiviral transfection is a safe and effective way to modify bone marrow mesenchymal stem cells. We can improve the ability of bone marrow mesenchymal stem cells in the survival and emiocytosis ability through genetic engineering.