Differentiation of marrow derived mesenchymal stem cells into neurons by noggin gene transfected with recombinant adenovirus vector / 解放军医学杂志

ABSTRACT

Objective To explore the appropriate method of isolating,purifying and culturing the rat bone marrow mesenchymal stem cells(MSCs) in vitro,and observe the differentiation of MSCs into neuron-like cells induced by noggin gene transfected with adenovirus vector.Methods MSCs obtained from bone marrow of SD rats were isolated,cultivated and amplified by Percoll density gradient centrifugation with adherent method,the the third passage of purified MSCs was then induced to differentiate into neurocytes by using the reconstructed noggin adenovirus vector(pAdEasy-1-GFP-noggin) and control vector(pAdEasy-1-GFP),respectively.After cultivation,the differentiated cells were identified by using immunocytochemical method with neurone-specific enolase(NSE),neurofilament 200(NF-200),neuronal nuclei(NeuN) and glial fibrillary acidic protein(GFAP),and the inductivity in the respective groups were analyzed.The Nissl bodies in the induced cells were displayed by thionine-eosin staining.Results The primarily cultured MSCs were spindle in shape,adhered 24 hours after cultivation,and then grew into small clones.Forty-eight hours after transfection by noggin recombinant adenovirus vector,the MSCs started to change their shape as observed under inverted microscope,and several axon-or dendrite-like processes with branches stretched out from the cell body.The induced cells derived from bone marrow MSCs specifically expressed NSE,NF and NeuN,but not GFAP by immunocytochemistry.A lot of Nissl bodies could be seen in the cell body of induced cells shown by Nissl staining.Conclusions Highly purified MSCs can be obtained by Percoll density gradient centrifugation combined with adherent method.The bone marrow derived MSCs transfected with noggin gene can differentiate into neuron-like cells in vitro.