Ligustrazine induces rat bone morrow mesenchymal stem cells to differentiate into neuron-like cells: Screening of the optimal inductive concentration / 中国组织工程研究

ABSTRACT

BACKGROUND: There are numerous inducers used in inducing bone marrow mesenchymal stem cells (BMMSCs) differentiate into neuron-like cells, however, due to poisonous, most chemical inducers can not be used in human.OBJECTIVE: To investigate the effect of ligustrazine on differentiation of rat BMMSCs into neuron-like cells in vitro, and to search for the optimal inductive concentration.METHODS: After SD rats were anesthetized, bone marrow was obtained from the femoral and tibial bones, centrifuged, and the supernatant was discarded. The extracted cells were cultured in L-DMEM supplemented with 15% fetal bovine serum. The expression of CD44 and CD45 of the 5~(th) passage of BMMSCs were identified by immunocytochemical technique. Serum-free L-DMEM medium contains 1.00, 1.25 and 1.50 g/L ligustrazine concentrations were used to induce the 5~(th) passage of BMMSCs in vitro. Morphology changes of BMMSCs were observed under an inverted phase microscope. Expression of nestin, neuron-specific enolase and glial fibrillary acidic protein were identified by immunocytochemical technique, and the expression ratio of neuron-like cells' surface antigens induced by different concentrations of ligustrazine were compared.RESULTS AND CONCLUSION: ①Most primarily cultured BMMSCs adhered to the wall at 3 days after culture, which proliferated faster after passaged, and the 5~(th) passage of cells were mostly purified into BMMSCs, spread radially or vortex-likely. ②The 5~(th) passage of BMMSCs was positive expressed (98.02±0.81)% CD44, but negative for CD45. ③Neuron-like cells with prominence and bifurcation could be seen after induction. The immunocytochemical method showed that nestin and neuron-specific enolase in most induced cells were positive expressed, especially received a highest ration of neuron-specific enolase expressing in the induced group with 1.25 g/L concentration of ligustrazine. It revealed that ligustrazine can induce BMMSCs differentiated into neuron-like cells, and 1.25 g/L is the optimal inductive concentration.