Sandeep Kadekar1, Laurent Barbe2, Martin Stoddart3, Oommen P Varghese1, Maria Tenje2, Gemma Mestres2. 1. Department of Chemistry-Ångström Laboratory, Uppsala University, 751 21 Uppsala, Sweden. 2. Department of Materials Science and Engineering, Science for Life Laboratory, Uppsala University, Box 35, 751 03 Uppsala, Sweden. 3. AO Research Institute Davos, 7270 Davos, Switzerland.
Abstract
INTRODUCTION: Human mesenchymal stem cells (hMSCs) have a great clinical potential for tissue regeneration purposes due to its multilineage capability. Previous studies have reported that a single addition of 5-azacytidine (5-AzaC) causes the differentiation of hMSCs towards a myocardial lineage. The aim of this work was to evaluate the effect of 5-AzaC addition frequency on hMSCs priming (i.e., indicating an early genetic differentiation) using two culture environments. METHODS: hMSCs were supplemented with 5-AzaC while cultured in well plates and in microfluidic chips. The impact of 5-AzaC concentration (10 and 20 μM) and addition frequency (once, daily or continuously), as well as of culture period (2 or 5 days) on the genetic upregulation of PPARγ (adipocytes), PAX3 (myoblasts), SOX9 (chondrocytes) and RUNX2 (osteoblasts) was evaluated. RESULTS: Daily delivering 5-AzaC caused a higher upregulation of PPARγ, SOX9 and RUNX2 in comparison to a single dose delivery, both under static well plates and dynamic microfluidic cultures. A particularly high gene expression of PPARγ (tenfold-change) could indicate priming of hMSCs towards adipocytes. CONCLUSIONS: Both macro- and microscale cultures provided results with similar trends, where addition frequency of 5-AzaC was a crucial factor to upregulate several genes. Microfluidics technology was proven to be a suitable platform for the continuous delivery of a drug and could be used for screening purposes in tissue engineering research.
INTRODUCTION: Human mesenchymal stem cells (hMSCs) have a great clinical potential for tissue regeneration purposes due to its multilineage capability. Previous studies have reported that a single addition of 5-azacytidine (5-AzaC) causes the differentiation of hMSCs towards a myocardial lineage. The aim of this work was to evaluate the effect of 5-AzaC addition frequency on hMSCs priming (i.e., indicating an early genetic differentiation) using two culture environments. METHODS: hMSCs were supplemented with 5-AzaC while cultured in well plates and in microfluidic chips. The impact of 5-AzaC concentration (10 and 20 μM) and addition frequency (once, daily or continuously), as well as of culture period (2 or 5 days) on the genetic upregulation of PPARγ (adipocytes), PAX3 (myoblasts), SOX9 (chondrocytes) and RUNX2 (osteoblasts) was evaluated. RESULTS: Daily delivering 5-AzaC caused a higher upregulation of PPARγ, SOX9 and RUNX2 in comparison to a single dose delivery, both under static well plates and dynamic microfluidic cultures. A particularly high gene expression of PPARγ (tenfold-change) could indicate priming of hMSCs towards adipocytes. CONCLUSIONS: Both macro- and microscale cultures provided results with similar trends, where addition frequency of 5-AzaC was a crucial factor to upregulate several genes. Microfluidics technology was proven to be a suitable platform for the continuous delivery of a drug and could be used for screening purposes in tissue engineering research.
Authors: V Labovsky; E L Hofer; L Feldman; V Fernández Vallone; H García Rivello; A Bayes-Genis; A Hernando Insúa; M J Levin; N A Chasseing Journal: Differentiation Date: 2009-11-18 Impact factor: 3.880
Authors: J Zych; M A Stimamiglio; A C Senegaglia; P R S Brofman; B Dallagiovanna; S Goldenberg; A Correa Journal: Braz J Med Biol Res Date: 2013-05 Impact factor: 2.590
Authors: Judit Vágó; Katalin Kiss; Edina Karanyicz; Roland Takács; Csaba Matta; László Ducza; Tibor A Rauch; Róza Zákány Journal: Cells Date: 2021-10-06 Impact factor: 6.600