Effat Alizadeh1, Nosratollah Zarghami1,2, Mohamadreza Baghaban Eslaminejad3, Abolfazl Akbarzadeh4, Abolfazl Barzegar5, Seyed Abolghasem Mohammadi6. 1. a Department of Medical Biotechnology , Faculty of Advanced Medical Sciences, Tabriz University of Medical Sciences , Tabriz , Iran. 2. b The Umbilical Cord Stem Cell Research Center (UCSRC), Tabriz University of Medical Sciences , Tabriz , Iran. 3. c Department of Stem Cells and Developmental Biology at Cell Sciences Research Center , Royan Institute for Stem Cell Biology and Technology, ACECR , Tehran , Iran. 4. d Department of Medical Nanotechnology , Faculty of Advanced Medical Sciences, Tabriz University of Medical Sciences , Tabriz , Iran. 5. e Research Institute for Fundamental Sciences (RIFS), University of Tabriz , Tabriz , Iran. 6. f Department of Agronomy and Plant Breeding , Faculty of Agriculture, University of Tabriz , Tabriz , Iran.
Abstract
INTRODUCTION: Adipose tissue-derived mesenchymal stem cells (AT-MSCs) are suitable choices in autologous stem cell treatment of liver-associated diseases due to their hepatic differentiation potential. Dimethyl sulfoxide (DMSO) is an amphipathic molecule with potential of delivering both lipophilic and hydrophilic agents into cells, also a common cryoprotectant for freezing of the cells. DMSO was used in some protocols for induction of AT-MSCs towards hepatocyte like cells. However, the effect of DMSO on hepatogenic differentiation of AT-MSCs were not surveyed, previously. In the present study, we aimed at evaluation of the effect of DMSO on differentiation of AT-MSCs into hepatic lineage. METHODS: We isolated mesenchymal stem cells (MSCs) from adipose tissue, and then verifies multi-potency and surface markers of AT-MSCs . Isolated AT-MSCs randomly dispensed in four groups including Group 1: HGF treated, 2: HGF+ DMSO treated, 3: HGF+ DMSO+ OSM treated, and group control for a period of 3 weeks in the expansion medium without serum; EGF and bFGF were also included in the first days of inductions. The morphologic changes during induction period was observed with microscopy. The secretion of albumin (ALB) of the differentiating MSCs was investigated using ELISA, and urea production was evaluated using colorimetric assay. The qRT-PCR was performed for quantitation of hepatocyte marker genes including AFP, ALB, CK18, HNF4a, and HNF6. The glycogen storage of differentiated cells was visualized by periodic-acid Schiff‘s staining. RESULTS: The results demonstrate that DMSO speeds up hepatic differentiation of AT-MSCs characterized by rapid changes in morphology; higher expression of hepatic marker gene (ALB) in both mRNA and protein level (P < 0.05); also increased transcriptional levels of other liver genes including CK18, HNF4a, and HNF6 (P < 0.01); and moreover, greater percentage of glycogen storage(p < 0.05) in DMSO-treated groups. CONCLUSION: DMSO catalyzes hepatic differentiation; therefore, using DMSO for acceleration of the hepatogenic protocols of AT-MSCs appears advantageous.
INTRODUCTION: Adipose tissue-derived mesenchymal stem cells (AT-MSCs) are suitable choices in autologous stem cell treatment of liver-associated diseases due to their hepatic differentiation potential. Dimethyl sulfoxide (DMSO) is an amphipathic molecule with potential of delivering both lipophilic and hydrophilic agents into cells, also a common cryoprotectant for freezing of the cells. DMSO was used in some protocols for induction of AT-MSCs towards hepatocyte like cells. However, the effect of DMSO on hepatogenic differentiation of AT-MSCs were not surveyed, previously. In the present study, we aimed at evaluation of the effect of DMSO on differentiation of AT-MSCs into hepatic lineage. METHODS: We isolated mesenchymal stem cells (MSCs) from adipose tissue, and then verifies multi-potency and surface markers of AT-MSCs . Isolated AT-MSCs randomly dispensed in four groups including Group 1: HGF treated, 2: HGF&plus; DMSO treated, 3: HGF&plus; DMSO&plus; OSM treated, and group control for a period of 3 weeks in the expansion medium without serum; EGF and bFGF were also included in the first days of inductions. The morphologic changes during induction period was observed with microscopy. The secretion of albumin (ALB) of the differentiating MSCs was investigated using ELISA, and urea production was evaluated using colorimetric assay. The qRT-PCR was performed for quantitation of hepatocyte marker genes including AFP, ALB, CK18, HNF4a, and HNF6. The glycogen storage of differentiated cells was visualized by periodic-acid Schiff&lsquo;s staining. RESULTS: The results demonstrate that DMSO speeds up hepatic differentiation of AT-MSCs characterized by rapid changes in morphology; higher expression of hepatic marker gene (ALB) in both mRNA and protein level (P &lt; 0.05); also increased transcriptional levels of other liver genes including CK18, HNF4a, and HNF6 (P &lt; 0.01); and moreover, greater percentage of glycogen storage(p &lt; 0.05) in DMSO-treated groups. CONCLUSION:DMSO catalyzes hepatic differentiation; therefore, using DMSO for acceleration of the hepatogenic protocols of AT-MSCs appears advantageous.