Forskolin-containing conditioned medium induces neural differentiation of adipose-derived stem cells:An in vitro experimental study
Objective:To explore the neural differentiation of adipose-derived stem cells(ASCs)induced by neural-induced conditional medium in vitro,and to study the optimal concentration of forskolin and the effect of optimal concentration on neurogenic differentiation of ASCs in vitro.Methods:ASCs were isolated by enzyme digestion method.The phenotype and differentiation ability of stem cells are identified by flow cytometry and cell staining.Nerve induction differentiation was performed in vitro by neural-induced conditional medium containing different concentrations of forskolin,and the optimal concentration was screened by immunofluorescence staining.ASCs were induced by neural-induced conditional medium containing the optimal concentration of forskolin and the morphological changes of cells were observed by microscope before induction,6 h after induction and 24 h after induction.Effects of induction on rat ASCs were analyzed by real-time quantitative polymerase chain reaction(RT-qPCR)and immunofluorescence.Results:The optimal concentration of forskolin in neural-induced conditional medium was 10 µmol/L,and the cells induced by neural-induced conditional medium containing 10 µmol/L forskolin could rapidly induce ASCs to differentiate in the direction of nerve formation.After induction,the cell morphology was similar to nerve cells.Results of RT-qPCR and immunofluorescence indicated that the expressions of anti-class Ⅲβ-tubulin(Tuj1),neurofilament-M(NF-M),nestin and S100 β at the mRNA level and protein level were increased after induction(P<0.05).Conclusion:Nerve differentiation of ASCs can be induced by conditional medium containing 10 µmol/L forskolin,and the induced cells are expected to become seed cells of facial nerve conduits.
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