Author/Authors :
Mobarra, Naser tehran university of medical sciences tums - School of Medicine, Students’Scientific Research Center - Department of Clinical Biochemistry, تهران, ايران , Soleimani, Masoud tarbiat modares university - School of Medical Sciences - Department of Hematology, تهران, ايران , Kouhkan, Fatemeh Stem Cell Technology Research Center - Department of Molecular Biology and Genetic Engineering, ايران , Hesari, Zahra tehran university of medical sciences tums - Faculty of Pharmacy - Department of Pharmaceutics, تهران, ايران , Lahmy, Reyhaneh tarbiat modares university - Faculty of Biological Sciences - Department of Genetics, تهران, ايران , Mossahebi-Mohammadi, Majid tarbiat modares university - School of Medical Sciences - Department of Hematology, تهران, ايران , Arefian, Ehsan Stem Cell Technology Research Center - Department of Molecular Biology and Genetic Engineering, ايران , Arefian, Ehsan university of tehran - School of Biology, College of Science - Microbial Biotechnology Laboratory, Department of Microbiology, تهران, ايران , Jaafarpour, Zahra islamic azad university - Department of Biology, ايران , Nasiri, Hajar tehran university of medical sciences tums - Hematology-Oncology and Stem cell Transplantation Research Center, Shariati Hospital, تهران, ايران , Pakzad, Reza tehran university of medical sciences tums - School of Public Health - Departments of Epidemiology and Biostatistics, تهران, ايران , Tavakoli, Rezvan Stem Cell Technology Research Center - Department of Molecular Biology and Genetic Engineering, ايران , Pasalar, Parvin tehran university of medical sciences tums - Metabolic disorder Research center, Endocrinology and Metabolism, Molecular sciences institute, تهران, ايران
Abstract :
Background: The use of stem cells is considered as an appropriate source in cell therapy and tissue engineering. Differentiation of human induced Pluripotent Stem Cells (hiPSCs) to Hepatocyte-like Cells (HLCs) on mouse embryonic fibroblasts (MEFs) feeders is confronted with several problems that hinder the clinical applications of these differentiated cells for the treatment of liver injuries. Safe appropriate cells for stem cell-based therapies could create new hopes for liver diseases. This work focused on the determination of a capacity/efficiency for the differentiation of the hiPSCs into Hepatocyte-like Cells on a novel human adult bone marrow mesenchymal stem cells (hMSCs) feeder. Materials and Methods: Undifferentiated human iPSCs were cultured on mitotically inactivated human adult bone marrow mesenchymal stem cells. A three-step differentiation process has been performed in presence of activin A which added for 3 days to induce a definitive endoderm formation. In the second step, medium was exchanged for six days. Subsequently, cells were treated with oncostatin M plus dexamethasone for 9 days to generate hepatic cells. Endodermic and liver-specific genes were assessed via quantitative reverse transcription-polymerase chain reaction and RT-PCR, moreover, immunocytochemical staining for liver proteins including albumin and alpha-fetoprotein. In addition, functional tests for glycogen storage, oil red examination, urea production and alpha-fetoprotein synthesis, as well as, cells differentiated with a hepatocyte-like morphology was also performed. Results: Our results show that inactivated human adult bone marrow mesenchymal stem cell feeders could support the efficient differentiation of hiPSCs into HLCs. This process induced differentiation of iPSCs into definitive endocrine cells that expressed sox17, foxa2 and expression of the specific genes profiles in hepatic-like cells. In addition, immunocytochemical analysis confirmed albumin and alpha-fetoprotein protein expression, as well as, the hiPSCs-derived Hepatocyte-like Cells on human feeder exhibited a typical morphology. Conclusions: we suggested a successful and efficient culture for differentiation and maturation of hepatocytes on an alternative human feeders; this is an important step to generate safe and functional hepatocytes that is vital for regenerative medicine and transplantation on the cell-based therapies.
