The Effect of Pancreas Islet-Releasing Factors on the Direction of Embryonic Stem Cells Towards Pdx1 Expressing Cells
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Diabetes mellitus, which is the result of autoimmune destruction of the insulin-producing β cells, occurs by loss of insulin-secreting capacity. The insufficient source of insulin-producing cells (IPCs) is the major obstacle for using transplantation as diabetes treatment method. The present study suggests a method to form islet-like clusters of IPCs derived from mouse embryonic stem cells (mESCs). This protocol consists of several steps. Before starting this protocol, embryoid bodies (EBs) should be cultured in suspension in conditioned medium of isolated mouse pancreatic islet in combination with activing A to be induced. Then differentiated mESCs were replaced with dishes supplemented with basic fibroblast growth factor (bFGF). Next, bFGF was withdrawn, and cyclopamine and noggin were added. Then the cells were treated with B27, nicotinamide, and islet-conditioned medium for maturation. mESCs, as the control group, were cultured without any treatment. An enhanced expression of pancreatic-specific genes was detected by qRT-PCR and immunofluorescence in the differentiated mESCs. The differentiated mESCsco express other markers of pancreatic islet cells as well as insulin. This method exhibited higher insulin generation and further improvement in IPCs protocol that may result in an unlimited source of ES cells suitable for transplantation. The results indicated that conditioned medium, just as critical components of the stem cell niche associated with other factors, had high potential to differentiate mESCs into IPCs.
KeywordsCo-culture Diabetes Differentiation ES cells Insulin-secreting cells Pancreatic development
Compliance with Ethical Standards
Conflict of Interest
The authors declare that they have no conflicts of interest.
- 7.D’Amour, K. A., Bang, A. G., Eliazer, S., Kelly, O. G., Agulnick, A. D., Smart, N. G., Moorman, M. A., Kroon, E., Carpenter, M. K., & Baetge, E. E. (2006). Production of pancreatic hormone-expressing endocrine cells from human embryonic stem cells. Nature Biotechnology, 24(11), 1392–1401.CrossRefGoogle Scholar
- 10.Ebrahimi-Barough, S., Hoveizi, E., Norouzi Javidan, A., & Ai, J. (2015). Investigating the neuroglial differentiation effect of neuroblastoma conditioned medium in human endometrial stem cells cultured on 3D nanofibrous scaffold. Journal of Biomedical Materials Research. Part A., 103(8), 2621–2627.CrossRefGoogle Scholar
- 16.Hoveizi, E., Nabiuni, M., Parivar, K., Ai, J. and Massumi, M. (2013) Definitive endoderm differentiation of human-induced pluripotent stem cells using signaling molecules and IDE1 in three-dimensional polymer scaffold. Journal of biomedical materials research. Part A, 102, 4027–36.Google Scholar
- 19.Kroon, E., Martinson, L. A., Kadoya, K., Bang, A. G., Kelly, O. G., Eliazer, S., Young, H., Richardson, M., Smart, N. G., Cunningham, J., Agulnick, A. D., D’Amour, K. A., Carpenter, M. K., & Baetge, E. E. (2008). Pancreatic endoderm derived from human embryonic stem cells generates glucose-responsive insulin-secreting cells in vivo. Nature Biotechnology, 26(4), 443–452.CrossRefGoogle Scholar
- 21.Lee, S. H., Park, M. H., Park, S. J., Kim, J., Kim, Y. T., Oh, M. C., Jeong, Y., Kim, M., Han, J. S., & Jeon, Y. J. (2012). Bioactive compounds extracted from Ecklonia cava by using enzymatic hydrolysis protects high glucose-induced damage in INS-1 pancreatic beta-cells. Applied Biochemistry and Biotechnology, 167(7), 1973–1985.CrossRefGoogle Scholar
- 23.Maehr, R., Chen, S., Snitow, M., Ludwig, T., Yagasaki, L., Goland, R., Leibel, R. L., & Melton, D. A. (2009). Generation of pluripotent stem cells from patients with type 1 diabetes. Proceedings of the National Academy of Sciences of the United States of America, 106(37), 15768–15773.CrossRefGoogle Scholar
- 24.Massumi, M., Hoveizi, E., Baktash, P., Hooti, A., Ghazizadeh, L., Nadri, S., Pourasgari, F., Hajarizadeh, A., Soleimani, M., Nabiuni, M., & Khorramizadeh, M. R. (2014). Efficient programming of human eye conjunctiva-derived induced pluripotent stem (ECiPS) cells into definitive endoderm-like cells. Experimental Cell Research, 322(1), 51–61.CrossRefGoogle Scholar
- 25.McLean, A. B., D’Amour, K. A., Jones, K. L., Krishnamoorthy, M., Kulik, M. J., Reynolds, D. M., Sheppard, A. M., Liu, H., Xu, Y., Baetge, E. E., & Dalton, S. (2007). Activin a efficiently specifies definitive endoderm from human embryonic stem cells only when phosphatidylinositol 3-kinase signaling is suppressed. Stem Cells, 25(1), 29–38.CrossRefGoogle Scholar
- 27.Ni, Z., Zhang, Y., Wang, H., Wei, Y., Ma, B., Hao, J., Tu, P., Duan, H., Li, X., Jiang, P., Ma, X., Wang, B., Wu, R., Zhu, J., & Li, M. (2016). Construction of a fusion peptide 5rolGLP-HV and analysis of its therapeutic effect on type 2 diabetes mellitus and thrombosis in mice. Applied Biochemistry and Biotechnology, 179(1), 59–74.CrossRefGoogle Scholar
- 34.Yang, J., Wu, C., Stefanescu, I. and Horowitz, A. (2017) Analysis of Retinoic Acid-induced Neural Differentiation of Mouse Embryonic Stem Cells in Two and Three-dimensional Embryoid Bodies. Journal of visualized experiments, https://doi.org/10.3791/55621.
- 36.Zhu, R., Liu, H., Liu, C., Wang, L., Ma, R., Chen, B., Li, L., Niu, J., Fu, M., Zhang, D. and Gao, S. (2017) Cinnamaldehyde in diabetes: A review of pharmacology, pharmacokinetics and safety. Pharmacological research, 122, 78–89.Google Scholar