Study On In Vitro Differentiation Of Human Adenoid Mesenchymal Stem Cells Into Olfactory Sensory Neurons

Objective: 1.To isolation and culture the mesenchymal stem cells(MSCs)derived from human adenoid tissues in vitro.The aim was to establish and obtain a new source of MSCs for scientific research and potential clinical applications.2.To investigate the differentiation of human adenoid MSCs into olfactory sensory neurons induced in vitro.Methods: 1.The adenoid tissues were obtained from the children with adenoid hypertrophy who underwent adenoidectomy,digested and isolated by trypsin,and cultured with adhesion method.The primary MSCs are purified by continuous direct adherent subculture.MSCs were further identified by detecting cell surface antigens(CD45,CD73,CD90)of the P5 generation by flow cytometry,and differentiation into osteoblasts and adipocytes induced by osteogenic and adipocytes induction solution,respectively.2.Adenoid MSCs were induced by RA,SHH,b FGF,RA+SHH,RA+b FGF,SHH+b FGF and RA+SHH+b FGF.The morphology of induced MSCs were observed under inverted microscope.The expression of β-tubulin 3 which is the specific marker of sensory neuron,and the expression of GAP43 and OMP which are the specific markers of olfactory sensory neuron,were detected by mmunofluorescence antibody assay.The expression intensity of the markers of olfactory sensory neuron in group cells were compared.Results: 1.The positive rates of CD73,CD90 and CD45 of the P5 human adenoid MSCs were 99.3%,97.7% and 0.61%respectively.Calcium nodules were observed by alizarin red staining after osteogenic induction,and lipid droplets were observed by oil red 0staining after lipid induction.2.The cells in every induction groups presented neuron-like appearance.Immunofluorescence showed thatβ-tubulin 3 was positively expressed in every group cells,and GAP43 positive cells in b FGF+SHH group and RA+SHH+b FGF group,no OMP positive cells in every group.The intensity of GAP43 on the cells of RA+SHH+b FGF group was higher than that of b FGF+SHH group.Conclusions:1.MSCs can be obtained from human adenoid tissues by isolation and culture.2.Adenoid MSCs can be induced to differentiate into neuron-like cells.3.human adenoid MSCs can differentiate into immature olfactory sensory neurons while induced by RA+SHH+ b FGF in vitro.5 figures,5 table,56 references...