298 Differentiation of Canine Amniotic Fluid Mesenchymal Stem Cells Into Neuronal Precursors

Abstract

The amniotic fluids contain mesenchymal stem cells and can be readily available for tissue engineering. Recently, regenerative treatments such as tissue engineering, cell therapy, and transplantation have shown potential in clinical trials of degenerative diseases. Physiologically, disease presentation and clinical responses in the dog are much more similar to that in the human compared with other traditional mammalian models. In addition, several researchers have demonstrated Canis familiaris is a suitable model for human diseases. The aim of the present study was to investigate whether canine amniotic fluid (cAF)-derived mesenchymal stem cells (MSC) can differentiate into neural precursor cells in vitro by neural induction reagent. The conditions of differentiation of MSC into neural cells were DMEM and N2-supplement, dibutyryl cyclic adenosine monophosphate, and butylated hydroxyanisole. During neural precursor differentiation, cAF-MSC can progressively acquire neuron-like morphology. Expressions of neuron cell-specific markers were examined before and after in vitro induction of differentiation. Changes in mRNA levels of specific genes were quantified by RT-PCR. The mRNA levels of NEFL (730%), GFAP (350%), β-tubuline 3 (2900%), and NSE (960%) were significantly increased after induction. The value of change in mRNA levels before and after induction was evaluated with the Image J program. In addition, the nestin, β-tubuline 3, and tyrosine hydroxylase protein expressions were confirmed by immunocytochemistry assay following the induction of differentiation, compared with the noninduction. In conclusion, this study demonstrated that cAF-MSC have great potential for neural precursor differentiation in vitro. Therefore, amniotic fluid may be a suitable alternative source of stem cells and can be applied to cell therapy in neurodegeneration diseases including Parkinson’s disease, Alzheimer’s disease, and Huntington’s disease.