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A simple method for the generation of insulin producing cells from bone marrow mesenchymal stem cells

  • Gholamreza Daryabor
  • Esmaeil Hashemi Shiri
  • Eskandar Kamali-SarvestaniEmail author
Article
  • 35 Downloads

Abstract

To produce insulin-producing cells (IPCs) from bone marrow mesenchymal stem cells (BM-MSCs) using a simple and cost effective method. During the initial 7 days of three-dimensional (3D) culture, BM-MSCs were cultured on 1% agar or agarose to form multicellular spheroids. Spheroids and spheroid-derived single cells (SS and SSC, respectively) were cultured in the absence of any proteinaceous growth factor in a simple specific medium for a further 7 d. The insulin content of the differentiated cells was evaluated at the mRNA and protein levels. Furthermore, the expression of pancreatic beta cells-related genes other than INS as well as the in vitro responses of IPCs to different glucose concentrations were investigated. Cellular clusters generated on agar and SS conditions (agar+SS-IPCs) stained better with beta cell specific stains and were more reactive to serum-containing insulin reactive antibodies compared with agarose-SS-IPCs. Gene expression analysis revealed that in comparison to agarose + SS-IPCs, agar+SS-IPCs expressed significantly higher levels of INS-1, INS-2, PDX-1, NKX6.1, and XBP-1. Of interest, agar+SS-IPCs expressed 2215.3 ± 120.8-fold more INS-1 gene compared to BM-MSCs. The expression of β-cell associated genes was also higher in agar+SS-IPCs compared to the agar+SSC-IPCs. Moreover, the expression of INS-1 gene was significantly higher in agar+SS-IPCs compared with agar+SSC-IPCs after culture in media with high concentration of glucose. Compared to the most expensive and time-consuming protocols, 3D culture of MSCs on agar followed by 2D culture of cellular clusters in a minimally supplemented high glucose media produced highly potent IPCs which may pay the way to the treatment of diabetic patients.

Keywords

Insulin-producing cells (IPCs) Mesenchymal stem cells Multicellular spheroids Three-dimensional (3D) culture 

Notes

Acknowledgments

The authors acknowledge staff in the Autoimmune Diseases Research Centre in particular Mrs. Taki for their help to accomplish the goals of the present study. The authors are also grateful to Ms. Yasamin Kamali Sarvestani for thorough English editing and correction of grammatical mistakes.

Funding

This study was financially supported by the Shiraz University of Medical Sciences (Grant Numbers: 94-7616). This paper was extracted from PhD theses of Gholamreza Daryabor for fulfilling his PhD degree.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflicts of interest.

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Copyright information

© The Society for In Vitro Biology 2019

Authors and Affiliations

  • Gholamreza Daryabor
    • 1
    • 2
  • Esmaeil Hashemi Shiri
    • 3
  • Eskandar Kamali-Sarvestani
    • 1
    • 3
    Email author
  1. 1.Department of Immunology, Shiraz Medical SchoolShiraz University of Medical SciencesShirazIran
  2. 2.Student Research CommitteeShiraz University of Medical SciencesShirazIran
  3. 3.Autoimmune Diseases Research CentreShiraz University of Medical SciencesShirazIran

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