Transfection of ECSOD to Rhesus bone marrow mesenchymal stem cells in vitro

Abstract

This study was aimed to investigate the biological effects of Rhesus bone marrow mesenchymal stem cells (R-BMMSC) transfected by adenovirus bearing extracellular superoxide dismutase gene (AD-ECSOD). Using density gradient centrifugation and adherent culture way, the R-BMMSC transfected by AD-ECSOD and reporter gene EGFP were isolated, cultured and purified; the transfection efficiency was detected by fluorescence microscopy and flow cytometry; the ECSOD protein expression in cell culture supernatant were detected by ELISA; the surface antigens on R-BMMSC (CD34, CD29, CD45, CD90, HLA-DR) were detected by flow cytometry; and differentiation capability of transfected R-BMMSC were detected by oil red O and alizarin staining; the proliferation capability of R-BMMSC was assay by MTT method. The results showed that the transfection efficiency of AD-ECSOD (MOI 500, 1 000, 1 500 and 2 000) for R-BMMSC was > 95%. At 24 h after transfection, the ECSOD protein could be detected in cell culture supernatant, and its level was significantly higher than that of control group (P < 0.01). At 48 h after transfection, the expression level of ECSOD protein on MOI 1 500 and 2 000 was the highest. The proliferative capability, surface antigen expression and multi directive differentiation ability of transfected R-BMMSC were similar to non-transfected R-BMMSC. It is concluded that the AD-ECSOD can effectively transfect the R-BMMSC without influences on its biological features.