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Molecular Biology Reports

, Volume 45, Issue 6, pp 2377–2391 | Cite as

Comparison of two different media for maturation rate of neural progenitor cells to neuronal and glial cells emphasizing on expression of neurotrophins and their respective receptors

  • Reihane Ebadi
  • Dor Mohammad Kordi-Tamandani
  • Kamran Ghaedi
  • Mohammad Hossein Nasr-Esfahani
Original Paper
  • 43 Downloads

Abstract

Neural cells derived from embryonic stem cells (ESCs) have potential usefulness for the treatment of neurodegenerative disorders. Modulation of intrinsic growth factors expression such as neurotrophins and their respective receptors by these cells is necessary to obtain functional neural cells for transplantation. In present study, we compared neural differentiation potential of two different media, NB + 5%ES-FBS + N2B27 and Ko-DMEM + 5%ES-FBS for conversion of mESC derived neural progenitors (NPs) into mature neural cells with emphasis on effect of the these two media on neurotrophins and their respective receptors expression. Immunofluorescence staining, RT-qPCR and western blot analysis showed that the expression of neuronal specific markers, MAP2 and Tuj-1, in NB + 5%ES-FBS + N2B27 medium was significantly higher than the other medium. Western blot assay revealed that the expression of BDNF and NGF increased significantly in mature neural cells obtained from NB + 5%ES-FBS + N2B27 medium but decreased in neural cells from Ko-DMEM + 5%ES-FBS medium compared to mESCs. TrkB protein was not detectable in mESCs but its expression increased in neural cells obtained from both media although its expression in NB + 5%ES-FBS + N2B27 medium was significantly higher than the other medium. In contrast to TrkB, p75NTR protein was detectable in mESCs and is remained constant in neural cells cultured in NB + 5%ES-FBS + N2B27 medium but decreased significantly in the other medium. In conclusion, our results indicated that NB + 5%ES-FBS + N2B27 medium promoted neural differentiation process of mESCs and caused enhancement of neurotrophins protein expression in addition to their cognate receptors.

Keywords

Mouse embryonic stem cell Medium Neural differentiation Neurotrophins Neurotrophin receptors 

Abbreviations

ANOVA

Two-way analysis of variance

BDNF

Brain-derived neurotrophic factor

bHLH

Basic helix–loop–helix

BSA

Bovine serum albumin

CNS

Central nervous system

CTCF

Corrected total cell fluorescence

DAPI

4,6-Diamidino-2-phenylindole

DMEM/F12

Dulbecco’s modified Eagle’s medium/Hams F12 medium

EB

Embryoid body

ECL

Enhanced chemiluminescence

ERK

Extracellular signal-regulated kinase

ESC

Embryonic stem cell

ES-FBS

Embryonic stem cell qualified fetal bovine serum

FITC

Fluorescein isothiocyanate

GAPDH

Glyceraldehyde 3-phosphate dehydrogenase

GFAP

Glial fibrillary acidic protein

HRP

Horse radish peroxidase

JNK

Jun N-terminal kinase

Ko-DMEM

Knock-out DMEM

LIF

Leukemia inhibitory factor

MAP2

Microtubule associated protein 2

mESC

Mouse embryonic stem cell

miRNA

MicroRNAs

NB

Neurobasal

NC

Neural cell

NGF

Nerve growth factor

NP

Neural progenitor

NSC

Neural stem cell

NT

Neurotrophin

NT-3

Neurotrophin-3

NT-4

Neurotrophin-4

p75NTR

p75 neurotrophin receptor

PBS

Phosphate buffered saline

PI3

Phosphatidylinositol-3-kinase

PVDF

Polyvinylidene difluoride

RA

Retinoic acid

RT

Room temprature

RT-qPCR

Real-time quantitative polymerase chain reaction

SDS-PAGE

Sodium dodecyl sulfate-polyacrylamide gel electrophoresis

SEM

Standard error of the mean

TRI

Total RNA isolation

Trk

Tropomyosin related kinase

Notes

Acknowledgements

Authors would like to thank University of Sistan and Baluchestan and Royan Institute for the financial support of this project. The authors are thankful to other members of Royan Institute for their excellent technical assistance and advice.

Author contributions

RE experimental design, data collection, data analysis, data interpretation and manuscript writing. DMKT experimental design, financial support, data analysis, data interpretation, and final approval of the manuscript. KG experimental design, data analysis, data interpretation, manuscript writing, and final approval of the manuscript. MHNE experimental design, financial support, data analysis, data interpretation, and final approval of manuscript.

Compliance with ethical standards

Conflict of interest

The authors indicate no potential conflict of interest.

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Copyright information

© Springer Nature B.V. 2018

Authors and Affiliations

  1. 1.Department of BiologyUniversity of Sistan and BaluchestanZahedanIran
  2. 2.Department of Biology, Faculty of SciencesUniversity of IsfahanIsfahanIran
  3. 3.Department of Cellular Biotechnology, Cell Science Research Center, Royan Institute for BiotechnologyACECRIsfahanIran

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