The Role Of Wnt/β3-catenin Signaling Pathway In The Neural-like Differentiation Of Muscle-derived Stem Cells

ObjectiveDetecting the expression of molecules relate to the Wnt/β-cateninsignaling pathway of the muscle-derived stem cells (MDSCs) in the processof Neural-like differentiation.To explore the role of Wnt/β-catenin signalingpathway in the differentiation of MDSCs.MethodsIsolated the musculatures from the hind legs of rats under asepticcondition.Cutting musculatures into pieces.Three kinds of enzyme were usedto digest the Musculature,cells were purified by differential attachmenttechnique.MDSCs were subcultured for5to10generation before follow-upexperiment.Cells were divide into three groups, in the control group cells werecultured with conventional growth medium.In experimental group,valproic acid(VA) was used to induce MDSCs.Inhibitor group was the same as theexperimental group,except the use of Dickkopf-1(DKK-1) and sFRP as theinhibitor of Wnt/β-catenin signaling pathway.Morphology of cells wereobserved under inverted phase contrast microscope after induced for4days.After induced for1and4days,The mRNA expression of Wnt3a wasdetected by RT-PCR and the protein expression of β-catenin and GSK-3βwere detected by Western blot.The expression of neuron specific enolase(NSE) and neurofibril (NF) were analyzed through Immunocytochemistry afterinduced for4days. ResultsMDSCs Isolated from the musculature by enzyme digestion anddifferential attachment technique had a good multiplicationcapacity.Originally,cells were short rod-shaped and small in size,then the cellbody extended gradually.After induced with VA,MDSCs in the experimentalgroup and inhibitor group differentiated into neural-like cells.Multipolar cellswere visible.The number of neuron-like cells in the experimental group wassignificantly higher than that in the inhibitor group.The mRNA expression ofWnt3a in the experimental group and inhibitor group was higher compared withthe control group.The experimental group was higher than that of inhibitorgroup.The protein expression of β-catenin in the experimental group andinhibitor group was higher compared with the control group,simultaneously,theexperimental group was higher than that of inhibitor group.The proteinexpression of GSK-3β in the experimental group and inhibitor group was lowerthan that of control group.Cells with positive staining were visible in bothexperimental group and inhibitor group.The experimental group had a largernumber of positive cells than that of inhibitor group for theimmunocytochemistry staining.ConclusionsWnt/β-catenin signaling pathway is involved in the neural differentiation ofMDSCs.Wnt/β-catenin signaling pathway may play a positive role in the neuraldifferentiation of MDSCs.