Avicenna J Med Biotech arij002 Avicenna Journal of Medical Biotechnology 2008-2835 2008-4625 Avicenna Research Institute ajmb226 Differentiation of Definitive Endoderm from Human Induced Pluripotent Stem Cells on hMSCs Feeder in a Defined Medium JaafarpourZahraDepartment of Clinical Nutrition and Dietetics, School of Nutrition Sciences and Food Technology, Shahid Beheshti University of Medical Sciences, Tehran, IranSoleimaniMasoudHepatitis & AIDS Department, Pasteur Institute of Iran , Tehran, Iran HosseinkhaniSamanReproductive Immunology Research Center, Avicenna Research Institute, ACECR, Tehran, IranKarimiMohammad HosseinDepartment of Botany and Microbiology, College of Science, King Saud University, Riyadh, Kingdom of Saudi ArabiaYaghmaeiMarjanDepartment of Pharmaceutical Biotechnology and Biotechnology Research Center, Faculty of Pharmacy, Tehran University of Medical Sciences, Tehran, IranMobarraNaserMonoclonal Antibody Research Center, Avicenna Research Institute, ACECR, Tehran, IranGeramizadehBitaMonoclonal Antibody Research Center, Avicenna Research Institute, ACECR, Tehran, Iran 8 1 2 8 26 5 2015 17 8 2015

<p>Background:<strong> </strong>The Definitive Endoderm (DE) differentiation using the undefined media and non-human feeders can cause contaminations in the generated cells for therapeutic applications. Therefore, generating safer and more appropriate DE cells is needed. This study compared five different methods to establish an appropriate method for inducing an efficient DE differentiation from Human Induced Pluripotent Stem Cells (hiPSCs) on an appropriate feeder in a more defined medium.<br /> Methods: Human Induced Pluripotent Stem Cells (hiPSCs) were cultured on inactivated feeders. Passaged hiPSCs, without feeder, were incubated for three days with Activin-A and different endodermal differentiation media including 1-FBS, 2-B27, 3-ITS and albumin fraction-V, 4-B27 and ITS and 5-like the third medium. The feeder cells in the first four methods were Mouse Embryonic Fibroblasts (MEFs) and in the fifth method were human adult bone marrow Mesenchymal Stem Cells (hMSCs). DE markers FOXA2, SOX17 and CXCR4 and also pluripotency marker OCT4 were evaluated using qRT-PCR, as well as FOXA2 by the immunocytochemistry.<br /> Results: QRT-PCR analysis showed that after three days, the expression levels of DE and pluripotency markers in the differentiated hiPSCs among all five groups did not have any significant differences. Similarly, the immunocytochemistry analysis demonstrated that the differentiated hiPSCs expressed FOXA2, with no significant differences.<br /> Conclusion: Despite this similarity in the results, the third differentiation medium has more defined and cost effective components. Furthermore, hMSC, a human feeder, is safer than MEF. Therefore, the fifth method is preferable among other DE differentiation methods and can serve as a fundamental method helping the development of regenerative medicine.</p>