In vitro differentiation of human adipose-derived adult stromal cells into neuron-like cells in hippocampal astrocyte conditioned medium

Xinchun Ye; Hongjun He; Feng Yang; Kepeng Zhao; Jun Yao; Bin Liu

摘要：BACKGROUND: At present, researches on differentiating from human adipose-derived adult stromal cells (hADASC) to neuron-like cells are focus on inducing by artificial-synthetic compound solution;however,hippocampal astrocyte conditioned medium(HCAM)can induce in vitro differentiation from hADASC to neuron-like cells is still unclear.OBJECTIVE:To observe whether HCAM can induce in vitro differentiation from hADASC to neuron-like cells.DESIGN:Randomized control study.SETTING:Department of Neurology,Taixing People's Hospital;Central Laboratory,North China Coal Medical College.MATERIALS:Donor of adipose tissue was donated by female volunteers suffering from caesarean section in the department of obstetrics ＆ gynecology in our hospital and aged 20-35 years. Adipose tissue was collected from subcutaneous tissue of abdomen during the operation.In addition.8 male newborn Wistar rats within 24 hours with average body mass of 20 g were provided by Animal Institute of Chinese Academy of Medical Sciences.Rabbit-anti-human Nestin polyclonal antibody.Rabbit-anti-human glial fibriliary acidic protein (GFAP)polyclonal antibody, rabbit-anti-human neuro-specific enolase polyclonal antibody and mouse-anti-human microtubal associated protein 2(MAP-2)polyclonal antibody were provided by Wuhan Boster Company.METHODS:The experiment was carried out in the Central Laboratory of North China Coal Medical College from October 2004 to June 2005.hADASC was cultured with HCAM and its growth and morphological changes were observed under inverted phase contrast microscope.Immunocytochemistry.immunofluorescence and Western blotting were used to evaluate the expressions of Nestin,which was a specific sign of nerve precursor,neuro-specific enolase and MAP-2,which was a specific sign of nerve cell,and GFAP,which was a specific sign of neuroglial cells.MAIN OUTCOME MEASURES:Nestin,which was a specific sign of nerve precursor,neuro-specific enolase and MAP-2,which was a specific sign of nerve cell,and GFAP,which was a specific sign of neuroglial cells.RESULTS:On the 3rd day of culture.partial hADASC started deformation from slender shuttle-shape cells to neuron-like cells. It suggested that cells stretched out apophysis. Which were mainly double-pole or multiple-pole cells.Five days later,immunohistochemical detection suggested that expression of Nestin(10.5±0.037)was found out in cells;meanwhile,expressions of GFAP(38.4±0.052)and neuro-specific enolase(NSE)(15.7±0.023)were also found out in cells;however,expression of MAP-2 was not observed.Western blot indicated that,5 days after effect of HCAM,Nestin was found out in hADASC;meanwhile,expressions of GFAP and neuro-specific enolase were also found out:however,expression of MAP-2 was not observed.CONCLUSION:HCAM can induce the differentiation from hADASC to neuron-like cells in vitro.