Abstract:

BACKGROUND: Before transplantation, the ability of stem cells was obviously increased by the transfection of target gene. The key of gene therapy is that target gene was transferred effectively and was expressed appropriately in stem cells. The choice of carrier system will affect the treatment effectiveness. OBJECTIVE: To observe the transfection efficiency of lentiviral vector transfecting the human adipose-derived stem cells (hASCs) and to detect the vitality of hASCs transfected.

METHODS: The recombinant lentiviral supernatant was obtained by standard lipofectamine reagent. The hASCs were transfected by lentiviral vector containing enhanced-green fluorescent protein (EGFP) with different multiplicity of infection (MOI=1, 5, 10, 15, 20). Transfection efficiency was assessed by flow cytometer for EGFP expression under fluorescence microscope. The multi-directional differentiation of hASCs after transfection was evaluated by alizarin red staining and oil red O staining. The proliferation of hASCs after transfection was detected by MTT assay. RESULTS AND CONCLUSION: Transfection efficiency was 60% at 3 day and 90%-95% at 5-7 day after infection under MOI=20. The alizarin red staining and oil red O staining showed hASCs after transfected hASCs were induced to osteogenic and adipogenic lineages, which suggested that viral infection could not affect multi-directional differentiation of hASCs. The proliferation of transfected hASCs had no marked change. This showed that hASCs transfected by lentiviral vectors carrying EGFP can effectively infect hASCs.