Protoplasmic Astrocytes Enhance the Ability of Neural Stem Cells to Differentiate into Neurons In Vitro

Yuan Liu,Li Wang,Yamin Wu,Zaiyun Long,Lin Zeng,Andreas Androutsellis-Theotokis

doi:10.1371/journal.pone.0038243

Abstract

Protoplasmic astrocytes have been reported to exhibit neuroprotective effects on neurons, but there has been no direct evidence for a functional relationship between protoplasmic astrocytes and neural stem cells (NSCs). In this study, we examined neuronal differentiation of NSCs induced by protoplasmic astrocytes in a co-culture model. Protoplasmic astrocytes were isolated from new-born and NSCs from the E13-15 cortex of rats respectively. The differentiated cells labeled with neuron-specific marker β-tubulin III, were dramatically increased at 7 days in the co-culture condition. Blocking the effects of brain-derived neurotrophic factor (BDNF) with an anti-BDNF antibody reduced the number of neurons differentiated from NSCs when co-cultured with protoplasmic astrocytes. In fact, the content of BDNF in the supernatant obtained from protoplasmic astrocytes and NSCs co-culture media was significantly greater than that from control media conditions. These results indicate that protoplasmic astrocytes promote neuronal differentiation of NSCs, which is driven, at least in part, by BDNF.

Highlights

The discovery that neural stem cells (NSCs) have the ability to regenerate functional neural cells has revolutionized neural tissue engineering and brought hopes to patients with central nervous system (CNS) disorders [1]
After withdraw of basic fibroblast growth factor (bFGF) and epidermal growth factor (EGF), the cells in spheres can differentiate into neurons and astrocytes as indicated with immunofluorescent staining by neuronal-specific marker b-tubulin III and astrocyte-specific marker glial fibrillary acidic protein (GFAP), which indicated that the NSCs have a multidifferentiation potential. (Fig. 1D)
Our results demonstrated that 7 days after co-culturing, more differentiated cells in the co-culture medium expressed the neuronal specific marker b-tubulin III, (75%66.4% vs. 59%65.2% in the NB+B27 group, 43%64.2% in the co-culture+brain-derived neurotrophic factor (BDNF) antibody and 32%64.6% in DMEM+10% FBS group; Fig. 3E), Which indicated that protoplasmic astrocytes and NSCs co-culture medium is more suitable for the neuronal differentiation of NSCs in vitro

Summary

Introduction

The discovery that neural stem cells (NSCs) have the ability to regenerate functional neural cells has revolutionized neural tissue engineering and brought hopes to patients with central nervous system (CNS) disorders [1]. NSCs can be expanded in the presence of mitogenic growth factors. NSCs have the potential to differentiate into neurons, astrocytes and oligodendrocytes in certain culture conditions [2]. Epidermal growth factor (EGF) and basic fibroblast growth factor (bFGF) have been shown to promote the proliferation of NSCs in culture [3,4]. Nerve growth factors such as neurotrophin-3 (NT-3) [5] and ciliary neurotrophic factor [6] have been reported to be key factors in regulating the differentiation of NSCs. Alternatively, signals from cell-cell contact can regulate the multi-potential of NSCs. For example, amniotic stem cells can promote dendrites outgrowth of differentiated neurons arising from NSCs [7]

Methods

Results

Conclusion