Effect of Spatial Arrangement of Substrate Topography on Neuronal Differentiation of Stem Cells

Three-dimensional gel matrices provide specialized microenvironments that mimic native tissues and enable stem cells to grow and differentiate into specific cell types. Here, we show that collagen three-dimensional gel matrices prepared in combination with adhesive proteins, such as fibronectin (FN) and laminin (LN), provide significant cues to the differentiation into neuronal lineage of mesenchymal stem cells (MSCs) derived from rat bone marrow. When cultured within either a three-dimensional collagen gel alone or one containing either FN or LN, and free of nerve growth factor (NGF), the MSCs showed the development of numerous neurite outgrowths. These were, however, not readily observed in two-dimensional culture without the use of NGF. Immunofluorescence staining, western blot and fluorescence-activated cell sorting analyses demonstrated that a large population of cells was positive for NeuN and glial fibrillary acidic protein, which are specific to neuronal cells, when cultured in the three-dimensional collagen gel. The dependence of the neuronal differentiation of MSCs on the adhesive proteins containing three-dimensional gel matrices is considered to be closely related to focal adhesion kinase (FAK) activation through integrin receptor binding, as revealed by an experiment showing no neuronal outgrowth in the FAK-knockdown cells and stimulation of integrin β1 gene. The results provided herein suggest the potential role of three-dimensional collagen-based gel matrices combined with adhesive proteins in the neuronal differentiation of MSCs, even without the use of chemical differentiation factors. Furthermore, these findings suggest that three-dimensional gel matrices might be useful as nerve-regenerative scaffolds.

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Large-scale generation of human iPSC-derived neural stem cells/early neural progenitor cells and their neuronal differentiation

Organogenesis ◽

10.1080/15476278.2015.1011921 ◽

2014 ◽

Vol 10 (4) ◽

pp. 365-377 ◽

Cited By ~ 49

Author(s):

Leonardo D’Aiuto ◽

Yun Zhi ◽

Dhanjit Kumar Das ◽

Madeleine R Wilcox ◽

Jon W Johnson ◽

...

Keyword(s):

Stem Cells ◽

Neural Stem Cells ◽

Progenitor Cells ◽

Neuronal Differentiation ◽

Large Scale ◽

Neural Progenitor Cells ◽

Neural Progenitor ◽

Human Ipsc

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The presence of FGF2 signaling determines whether β-catenin exerts effects on proliferation or neuronal differentiation of neural stem cells

Developmental Biology ◽

10.1016/j.ydbio.2003.12.024 ◽

2004 ◽

Vol 268 (1) ◽

pp. 220-231 ◽

Cited By ~ 145

Author(s):

Nipan Israsena ◽

Min Hu ◽

Weimin Fu ◽

Lixin Kan ◽

John A Kessler

Keyword(s):

Stem Cells ◽

Neural Stem Cells ◽

Neuronal Differentiation

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Icarisid II promotes proliferation and neuronal differentiation of neural stem cells via activating Wnt/β‐catenin signaling pathway

Phytotherapy Research ◽

10.1002/ptr.7022 ◽

2021 ◽

Author(s):

Hong‐He Xiao ◽

Ming‐Bo Zhang ◽

Jun‐Ting Xu ◽

Yan Deng ◽

Ning Li ◽

...

Keyword(s):

Stem Cells ◽

Neural Stem Cells ◽

Signaling Pathway ◽

Neuronal Differentiation

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Dual Inhibition of Activin/Nodal/TGF-βand BMP Signaling Pathways by SB431542 and Dorsomorphin Induces Neuronal Differentiation of Human Adipose Derived Stem Cells

Stem Cells International ◽

10.1155/2016/1035374 ◽

2016 ◽

Vol 2016 ◽

pp. 1-13 ◽

Cited By ~ 13

Author(s):

Vedavathi Madhu ◽

Abhijit S. Dighe ◽

Quanjun Cui ◽

D. Nicole Deal

Keyword(s):

Stem Cells ◽

Nervous System ◽

Neuronal Differentiation ◽

Adult Stem Cells ◽

Embryonic Stem ◽

Bmp Signaling ◽

Specific Marker ◽

Adipose Derived Stem Cells ◽

Neuronal Marker ◽

Dual Inhibition

Damage to the nervous system can cause devastating diseases or musculoskeletal dysfunctions and transplantation of progenitor stem cells can be an excellent treatment option in this regard. Preclinical studies demonstrate that untreated stem cells, unlike stem cells activated to differentiate into neuronal lineage, do not survive in the neuronal tissues. Conventional methods of inducing neuronal differentiation of stem cells are complex and expensive. We therefore sought to determine if a simple, one-step, and cost effective method, previously reported to induce neuronal differentiation of embryonic stem cells and induced-pluripotent stem cells, can be applied to adult stem cells. Indeed, dual inhibition of activin/nodal/TGF-βand BMP pathways using SB431542 and dorsomorphin, respectively, induced neuronal differentiation of human adipose derived stem cells (hADSCs) as evidenced by formation of neurite extensions, protein expression of neuron-specific gamma enolase, and mRNA expression of neuron-specific transcription factors Sox1 and Pax6 and matured neuronal marker NF200. This process correlated with enhanced phosphorylation of p38, Erk1/2, PI3K, and Akt1/3. Additionally,in vitrosubcutaneous implants of SB431542 and dorsomorphin treated hADSCs displayed significantly higher expression of active-axonal-growth-specific marker GAP43. Our data offers novel insights into cell-based therapies for the nervous system repair.

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Hypoxic Culture Promotes Dopaminergic-Neuronal Differentiation of Nasal Olfactory Mucosa Mesenchymal Stem Cells via Upregulation of Hypoxia-Inducible Factor-1α

Cell Transplantation ◽

10.1177/0963689717720291 ◽

2017 ◽

Vol 26 (8) ◽

pp. 1452-1461 ◽

Cited By ~ 8

Author(s):

Yi Zhuo ◽

Lei Wang ◽

Lite Ge ◽

Xuan Li ◽

Da Duan ◽

...

Keyword(s):

Stem Cells ◽

Mesenchymal Stem Cells ◽

Neuronal Differentiation ◽

Hypoxia Inducible Factor ◽

Cell Voltage ◽

Ambient Air ◽

Olfactory Mucosa ◽

Hypoxia Inducible Factor 1Α ◽

Downstream Target

Olfactory mucosa mesenchymal stem cells (OM-MSCs) display significant clonogenic activity and may be easily propagated for Parkinson’s disease therapies. Methods of inducing OM-MSCs to differentiate into dopaminergic (DAergic) neurons using olfactory ensheathing cells (OECs) are thus an attractive topic of research. We designed a hypoxic induction protocol to generate DAergic neurons from OM-MSCs using a physiological oxygen (O2) level of 3% and OEC-conditioned medium (OCM; HI group). The normal induction (NI) group was cultured in O2 at ambient air level (21%). The role of hypoxia-inducible factor-1α (HIF-1α) in the differentiation of OM-MSCs under hypoxia was investigated by treating cells with an HIF-1α inhibitor before induction (HIR group). The proportions of β-tubulin- and tyrosine hydroxylase (TH)-positive cells were significantly increased in the HI group compared with the NI and HIR groups, as shown by immunocytochemistry and Western blotting. Furthermore, the level of dopamine was significantly increased in the HI group. A slow outward potassium current was recorded in differentiated cells after 21 d of induction using whole-cell voltage-clamp tests. A hypoxic environment thus promotes OM-MSCs to differentiate into DAergic neurons by increasing the expression of HIF-1α and by activating downstream target gene TH. This study indicated that OCM under hypoxic conditions could significantly upregulate key transcriptional factors involved in the development of DAergic neurons from OM-MSCs, mediated by HIF-1α. Hypoxia promotes DAergic neuronal differentiation of OM-MSCs, and HIF-1α may play an important role in hypoxia-inducible pathways during DAergic lineage specification and differentiation in vitro.

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