Differentiation of adipose-derived stem cells into neuron-like cells with different methods

Abstract

Objective To evaluate the differentiation of adipose stem cells(ADSCs)into neuron-like cells(NLCs) by comparing chemical and growth factor induction methods, and deeply explore the induction mechanism and the problems. Methods ADSCs were isolated from the groin and retroperitoneal fat of SD rats and cultured. The ADSCs of 3rd generation were used for identification of phenotypes(CD29, CD34, CD45, CD73, CD90, CD105) by flow cytometry. The experiment was divided into chemical induction group [DMEM/F12+ 5 mmol/L β-mercaptoethanol(β-BME)+2% dimethylsurfoxide(DMSO)+200μmol/L butyrate hydroxyanisole(BHA)] and growth factor induction group[first step: DMEM/F12+20μg/L epidermal growth factor(EGF)+20μg/L basic fibroblast growth factor(bFGF)+2% B27; second step: DMEM/F12+ 10μg/L brain derived neurotrophic factor(BDNF)+10μg/L glial cell linederived neurotrophic factor(GDNF)+1μmol/L retinoic acid(RA)]. Differentiated ADSCs were evaluated for Nestin, NeuN, microtubule- associated protein(MAP)- 2 and glial fibrillary acidic protein(GFAP)expression by immunocytochemistry. The data were statistically analyzed. Results Flow cytometric analysis demonstrated that ADSCs at 3rd generation were positive for CD29(99.38%), CD90(96.21%), CD73(99.80%) and CD105 (97.97%), but negative for CD34(0.57%) and CD45(0.67%). In chemical induction group, Nestin and MAP-2 were highly expressed at first day, but the nerve-like morphology reversed at the fifth day and some cells begin to die at the seventh day. In growth factor induction group, Nestin expression was detected on the third day, and MAP-2, NeuN and GFAP expression was gradually increased afer one week. The stable NLCs were eventually induced. Conclusion Comparative analysis showed that ADSCs differentiate into neuron-like cells more quickly by chemical induction, but their nature is not stable. Induction by growth factors spends a relatively long time, but the induced NLCs maintain longer and more stable. Key words: Adipose-derived stem cells; Induce; Differentiation; Neuron-like cells