scholarly journals Neuron-Specific Fluorescence Reporter-Based Live Cell Tracing for Transdifferentiation of Mesenchymal Stem Cells into Neurons by Chemical Compound

2017 ◽  
Vol 2017 ◽  
pp. 1-10 ◽  
Author(s):  
Do Won Hwang ◽  
Hyun Woo Kwon ◽  
Jaeho Jang ◽  
Hee Jung Jung ◽  
Kwang Rok Kim ◽  
...  
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Neuronal Differentiation ◽  
Small Molecule ◽  
Plasmid Vector ◽  
Time Lapse ◽  
Live Cell ◽  
Easy Method ◽  
Specific Fluorescence

Although transdifferentiation of mesenchymal stem cells (MSCs) into neurons increases the possibility of therapeutic use of MSCs for neurodevelopmental disorders, the use of MSCs has the limitation on differentiation efficiency to neuronal lineage and lack of an easy method to monitor the transdifferentiation. In this study, using time-lapse live cell imaging, we assessed the neuronal differentiation of MSCs induced by a small molecule “NHPDQC (N-hydroxy-2-oxo-3-(3-phenylprophyl)-1,2-dihydroquinoxaline-6-carboxamide, C18H17N3O3).” Plasmid vector containing red fluorescence reporter genes under the control of the tubulin α1 (Tα1) promoter (pTα1-DsRed2) traced the neuronal differentiation of MSCs. Two days after NHPDQC treatment, MSCs showed neuron-like phenotype with neurite outgrowth and high expression of neuron-specific markers in more than 95% cells. The fluorescence signals increased in the cytoplasm of pTα1-DsRed2-transfected MSCs after NHPDQC treatment. In vitro monitoring of MSCs along the time courses showed progressive increase of fluorescence till 30 h after treatment, corresponding with the increase in neurite length. We examined an efficient neuronal differentiation of MSCs by NHPDQC alone and monitored the temporal changes of neuronal differentiation by neuron-specific fluorescence reporter along time. This method would help further our understanding of the differentiation of MSCs to produce neurons by simple treatment of small molecule.

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

2017 ◽  
Vol 26 (8) ◽  
pp. 1452-1461 ◽  
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.

scholarly journals Live-cell time-lapse imaging and single-cell tracking of in vitro cultured neural stem cells – Tools for analyzing dynamics of cell cycle, migration, and lineage selection

Methods ◽  
2018 ◽  
Vol 133 ◽  
pp. 81-90 ◽  
Author(s):  
Katja M. Piltti ◽  
Brian J. Cummings ◽  
Krystal Carta ◽  
Ayla Manughian-Peter ◽  
Colleen L. Worne ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Stem Cells ◽  
Neural Stem Cells ◽  
Single Cell ◽  
Cell Tracking ◽  
Time Lapse ◽  
Live Cell ◽  
Time Lapse Imaging

scholarly journals Optimal Pore Size of Honeycomb Polylactic Acid Films for In Vitro Cartilage Formation by Synovial Mesenchymal Stem Cells

2021 ◽  
Vol 2021 ◽  
pp. 1-9
Author(s):  
Misaki Yagi ◽  
Mitsuru Mizuno ◽  
Ryota Fujisawa ◽  
Hisako Katano ◽  
Kentaro Endo ◽  
...  
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Pore Size ◽  
Polylactic Acid ◽  
Time Lapse ◽  
Induction Medium ◽  
High Porosity ◽  
Cartilage Formation ◽  
Selection Of

Background. Tissue engineering of cartilage requires the selection of an appropriate artificial scaffold. Polylactic acid (PLA) honeycomb films are expected to be highly biodegradable and cell adhesive due to their high porosity. The purpose of this study was to determine the optimal pore size of honeycomb PLA films for in vitro cartilage formation using synovial mesenchymal stem cells (MSCs). Methods. Suspensions of human synovial MSCs were plated on PLA films with different pore sizes (no pores, or with 5 μm or 20 μm pores) and then observed by scanning electron microscopy. The numbers of cells remaining in the film and passing through the film were quantified. One day after plating, the medium was switched to chondrogenic induction medium, and the films were time-lapse imaged and observed histologically. Results. The 5 μm pore film showed MSCs with pseudopodia that extended between several pores, while the 20 μm pore film showed MSC bodies submerged into the pores. The number of adhered MSCs was significantly lower for the film without pores, while the number of MSCs that passed through the film was significantly higher for the 20 μm pore film. MSCs that were induced to form cartilage peeled off as a sheet from the poreless film after one day. MSCs formed thicker cartilage at two weeks when growing on the 5 μm pore films than on the 20 μm pore films. Conclusions. Honeycomb PLA films with 5 μm pores were suitable for in vitro cartilage formation by synovial MSCs.

scholarly journals Treatment of Periodontal Ligament Stem Cells with MOR and CBD Promotes Cell Survival and Neuronal Differentiation via the PI3K/Akt/mTOR Pathway

2018 ◽  
Vol 19 (8) ◽  
pp. 2341 ◽  
Author(s):  
Veronica Lanza Cariccio ◽  
Domenico Scionti ◽  
Antonio Raffa ◽  
Renato Iori ◽  
Federica Pollastro ◽  
...  
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Neuronal Differentiation ◽  
Periodontal Ligament ◽  
Combined Treatment ◽  
Mtor Pathway ◽  
Periodontal Ligament Stem Cells ◽  
Differentiation Potential ◽  
Neuronal Markers

Periodontal ligament mesenchymal stem cells (hPDLSCs), as well as all mesenchymal stem cells, show self-renewal, clonogenicity, and multi-tissue differentiation proprieties and can represent a valid support for regenerative medicine. We treated hPDLSCs with a combination of Moringin (MOR) and Cannabidiol (CBD), in order to understand if treatment could improve their survival and their in vitro differentiation capacity. Stem cells survival is fundamental to achieve a successful therapy outcome in the re-implanted tissue of patients. Through NGS transcriptome analysis, we found that combined treatment increased hPDLSCs survival, by inhibition of apoptosis as demonstrated by enhanced expression of anti-apoptotic genes and reduction of pro-apoptotic ones. Moreover, we investigated the possible involvement of PI3K/Akt/mTOR pathway, emphasizing a differential gene expression between treated and untreated cells. Furthermore, hPDLSCs were cultured for 48 h in the presence or absence of CBD and MOR and, after confirming the cellular viability through MTT (3-(4,5-dimethylthiazolyl-2)-2,5-diphenyltetrazoliumbromide) assay, we examined the presence of neuronal markers, through immunofluorescence analysis. We found an increased expression of Nestin and GAP43 (growth associated protein 43) in treated cells. In conclusion, hPDLSCs treated with Moringin and Cannabidiol showed an improved survival capacity and neuronal differentiation potential.

scholarly journals Polydopamine-based biofunctional substrate coating promotes mesenchymal stem cell migration

MRS Advances ◽  
2021 ◽  
Author(s):  
Zijun Deng ◽  
Weiwei Wang ◽  
Xun Xu ◽  
Nan Ma ◽  
Andreas Lendlein
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Cell Migration ◽  
Time Lapse ◽  
Implant Surface ◽  
Tissue Integration ◽  
Coated Substrate ◽  
Fibronectin Deposition

Abstract Rapid migration of mesenchymal stem cells (MSCs) on device surfaces could support in vivo tissue integration and might facilitate in vitro organoid formation. Here, polydopamine (PDA) is explored as a biofunctional coating to effectively promote MSC motility. It is hypothesized that PDA stimulates fibronectin deposition and in this way enhances integrin-mediated migration capability. The random and directional cell migration was investigated by time-lapse microscopy and gap closure assay respectively, and analysed with softwares as computational tools. A higher amount of deposited fibronectin was observed on PDA substrate, compared to the non-coated substrate. The integrin β1 activation and focal adhesion kinase (FAK) phosphorylation at Y397 were enhanced on PDA substrate, but the F-actin cytoskeleton was not altered, suggesting MSC migration on PDA was regulated by integrin initiated FAK signalling. This study strengthens the biofunctionality of PDA coating for regulating stem cells and offering a way of facilitating tissue integration of devices. Graphic abstract Polydopamine-coated substrate induces increased fibronectin deposition of mesenchymal stem cells, and promotes cell migration via integrin-initiated FAK signaling, compared to non-coated polystyrene-based standard tissue culture surface. In this way, multifunctional PDA coating could support in vivo tissue integration on implant surface and promote in vitro organoid formation.

scholarly journals Adipose-Derived Mesenchymal Stem Cells From a Hypoxic Culture Improve Neuronal Differentiation and Nerve Repair

2021 ◽  
Vol 9 ◽  
Author(s):  
Szu-Hsien Wu ◽  
Yu-Ting Liao ◽  
Kuang-Kai Hsueh ◽  
Hui-Kuang Huang ◽  
Tung-Ming Chen ◽  
...  
Keyword(s):  
Stem Cells ◽  
Animal Model ◽  
Mesenchymal Stem Cells ◽  
Sciatic Nerve ◽  
Neuronal Differentiation ◽  
Gastrocnemius Muscle ◽  
Differentiation Potential ◽  
Neuronal Markers

Hypoxic expansion has been demonstrated to enhance in vitro neuronal differentiation of bone-marrow derived mesenchymal stem cells (BMSCs). Whether adipose-derived mesenchymal stem cells (ADSCs) increase their neuronal differentiation potential following hypoxic expansion has been examined in the study. Real-time quantitative reverse transcription-polymerase chain reaction and immunofluorescence staining were employed to detect the expression of neuronal markers and compare the differentiation efficiency of hypoxic and normoxic ADSCs. A sciatic nerve injury animal model was used to analyze the gastrocnemius muscle weights as the outcomes of hypoxic and normoxic ADSC treatments, and sections of the regenerated nerve fibers taken from the conduits were analyzed by histological staining and immunohistochemical staining. Comparisons of the treatment effects of ADSCs and BMSCs following hypoxic expansion were also conducted in vitro and in vivo. Hypoxic expansion prior to the differentiation procedure promoted the expression of the neuronal markers in ADSC differentiated neuron-like cells. Moreover, the conduit connecting the sciatic nerve gap injected with hypoxic ADSCs showed the highest recovery rate of the gastrocnemius muscle weights in the animal model, suggesting a conceivable treatment for hypoxic ADSCs. The percentages of the regenerated myelinated fibers from the hypoxic ADSCs detected by toluidine blue staining and myelin basic protein (MBP) immunostaining were higher than those of the normoxic ones. On the other hand, hypoxic expansion increased the neuronal differentiation potential of ADSCs compared with that of the hypoxic BMSCs in vitro. The outcomes of animals treated with hypoxic ADSCs and hypoxic BMSCs showed similar results, confirming that hypoxic expansion enhances the neuronal differentiation potential of ADSCs in vitro and improves in vivo therapeutic potential.

P3-400: Neuronal differentiation of mesenchymal stem cells from bone marrow of human mastoid process in vitro

2010 ◽  
Vol 6 ◽  
pp. S569-S569
Author(s):  
Min-Kyung Song ◽  
Hyong Ho Cho ◽  
Byung Chae Kim ◽  
Han Seong Jeong ◽  
Su Jeong Jang
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells ◽  
Neuronal Differentiation ◽  
Mastoid Process

scholarly journals The migratory and mitotic behaviour of glioma stem cells in vitro: Optimisation of live-cell time-lapse microscopy

2011 ◽  
Vol 9 (7) ◽  
pp. 514-515
Author(s):  
James Barnett ◽  
Omar Pathmanaban ◽  
Ian Kamaly-Asl ◽  
Brian Bigger
Keyword(s):  
Stem Cells ◽  
Time Lapse ◽  
Live Cell ◽  
Glioma Stem Cells ◽  
Time Lapse Microscopy
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