ABSTRACT
Objective: To construct adipose-derived stem cells (ADSCs) line that can stably express brain-derived neurotrophic factor (BDNF) and neurotrophin-3(NT-3) genes and elucidate its significance. Methods: ADSCs were obtained by collagenase digestion along with differential adhesion method. After 2 and 4 weeks of osteogenic induction, alkaline phosphatase staining and alizarin red staining were performed. The third generation of ADSCs were transfected with Lenti-BDNF-GFP and Lenti-NT-3-RFP recombinant lentivirus solution. The ADSCs line that stably expressed BDNF and NT-3 genes were obtained by the optimal infection value determined before. Results: The ADSCs isolated and cultured successfully had the potential to differentiate in varous directions. After being induced to osteogenesis, alkaline phosphatase and alizarin red staining both showed positive. The best infection value for Lenti-BDNF-GFP and Lenti-NT-3-RFP recombinant lentivirus transfection was 100 while the infection duration was 72 hours. Expressions of BDNF and NT-3 in co-transfection group remained stable and high at both gene and protein levels. Conclusion: Establishment of ADSCs with stable and over-expressed BDNF and NT-3 genes is of great significance for treatment of spinal cord injury (SCI). It can solve the problem of low amount of neurotrophin secreted and short half-life during the treatment of SCI by ADSCs transplantation, which has great significance for further studies on the repair mechanism of SCI.