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In vitro differentiation of adipose-tissue-derived mesenchymal stem cells into neural retinal cells through expression of human PAX6 (5a) gene

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Abstract

The neural retina is subjected to various degenerative conditions. Regenerative stem-cell-based therapy holds great promise for treating severe retinal degeneration diseases, although many drawbacks remain to be overcome. One important problem is to gain authentically differentiated cells for replacement. Paired box 6 protein (5a) (PAX6 (5a)) is a highly conserved master control gene that has an essential role in the development of the vertebrate visual system. Human adipose-tissue-derived stem cell (hADSC) isolation was performed by using fat tissues and was confirmed by the differentiation potential of the cells into adipocytes and osteocytes and by their surface marker profile. The coding region of the human PAX6 (5a) gene isoform was cloned and lentiviral particles were propagated in HEK293T. The differentiation of hADSCs into retinal cells was characterized by morphological characteristics, quantitative real-time reverse transcription plus the polymerase chain reaction (qPCR) and immunocytochemistry (ICC) for some retinal cell-specific and retinal pigmented epithelial (RPE) cell-specific markers. hADSCs were successfully isolated. Flow cytometric analysis of surface markers indicated the high purity (∼97 %) of isolated hADSCs. After 30 h of post-transduction, cells gradually showed the characteristic morphology of neuronal cells and small axon-like processes emerged. qPCR and ICC confirmed the differentiation of some neural retinal cells and RPE cells. Thus, PAX6 (5a) transcription factor expression, together with medium supplemented with fibronectin, is able to induce the differentiation of hADSCs into retinal progenitors, RPE cells and photoreceptors.

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Acknowledgments

Our sincere thanks to Ali Fallah and Shamila Darvishalipoori for their contribution to the completion of this study.

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Authors and Affiliations

  1. Department of Biology, Faculty of Science, Ferdowsi University of Mashhad, Mashhad, Iran

    Habib Rezanejad, Farhang Haddad & Maryam M. Matin

  2. National Institute of Genetic Engineering and Biotechnology, Tehran, Iran

    Zahra-Soheila Soheili

  3. Iranian Blood Transfusion Organization Research Center, Tehran, Iran

    Shahram Samiei

  4. Tehran University of Medical Sciences, Tehran, Iran

    Ali Manafi

  5. Ophthalmic Research Center, Shahid Beheshti University of Medical Sciences, Tehran, Iran

    Hamid Ahmadieh

Authors
  1. Habib Rezanejad
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  2. Zahra-Soheila Soheili
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  3. Farhang Haddad
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  4. Maryam M. Matin
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  5. Shahram Samiei
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  6. Ali Manafi
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  7. Hamid Ahmadieh
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Corresponding author

Correspondence to Habib Rezanejad.

Additional information

This study was supported by the Iranian Council of Stem Cell Technology and the National Institute of Genetic Engineering and Biotechnology (NIGEB) via grant no. 410 and by the Department of Research and Technology of the Ferdowsi University of Mashhad via grant no. 3/20368.

Z.-S.S. and F.H. contributed equally to this work.

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Rezanejad, H., Soheili, ZS., Haddad, F. et al. In vitro differentiation of adipose-tissue-derived mesenchymal stem cells into neural retinal cells through expression of human PAX6 (5a) gene. Cell Tissue Res 356, 65–75 (2014). https://doi.org/10.1007/s00441-014-1795-y

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  • DOI: https://doi.org/10.1007/s00441-014-1795-y

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