Resveratrol Induced Differentiation Of Rat Bone Marrow Stromal Cells Into Neuronal-like Cells And The Study Of Its Mechanism

Objective: To study the differentiation of rat bone marrow stromalcells(MSCs) into neuronal-like cells induced by resveratrol in vitro and itsmechanism.Methods: Rat MSCs from the whole bone marrow were cultured byadherent method in vitro and induced to differentiate into neuronal-likecells with serum-free DMEM/F12containing resveratrol. Cell morphologywas observed with invert microscope. Scanning electron microscopedetected primary cilia at cell surface. The indirect immunofluorescenceand Western blotting detected the protein expressions of nestin and neuronspecific enolase(NSE), microtubule-associated protein2(MAP-2), glialfibrillary acidic protein(GFAP), acetylated α-tubulin(Ac-Tu), Patched1(Ptc1), smoothened(Smo) and Gli1.Results:1. Almost all MSCs and the control group cells were positivefor NSE by immunofluorescence. The differentiated cells took onneuronal-like morphology with the cell body shrinkage, extend longprocesses after induced by resveratrol, were positive for NSE and MAP-2 expression and were negative for GFAP expression byimmunofluorescence. The expression of nestin gradually weakened andthe expression of NSE and MAP-2gradually strengthened with theprolongation of induction and were negative for GFAP expression byWestern blotting.2. There was no expression of primary cilia in normal culturedMSCs. After pre-induction or24h starvation, MSCs expressed primarycilia, Ptc1, Smo, and Gli1. Ptc1is situated in primary cilia. Smo and Gli1are situated in the cytosol. After resveratrol induced, Smo entered into theprimary cilia from cytoplasm and Gli1translocated to the nucleus fromthe cytoplasm. Meanwhile, the expressions of Smo and Gli1proteingradually strengthened with the prolongation of induction by Westernblotting assay. In normal cultivation, resveratrol can not promote thetranslocation of Smo and the differentiation of MSCs into neuronal-likecells. When MSCs expressed primary cilia, resveratrol and Smoagonist(SAG) can make Smo to translocate to primary cilia from thecytosol. At the same time, MSCs were induced to differentiate intoneuronal-like cells. Moreover, expressions of Smo and Gli1wereenhanced. In contrast, cyclopamine can prevent these processes.Conclusion：1. The result showed that resveratrol can induce thedifferentiation of MSCs into neuronal-like cells in vitro2. MSCs express primary cilia and have Shh signaling. Primary cilia mediated Shh signaling regulate the neuronal-like cell differentiationof MSCs.