Comparison of Sources and Methods for the Isolation of Equine Adipose Tissue-Derived Stromal/Stem Cells and Preliminary Results on Their Reaction to Incubation with 5-Azacytidine
Dagmar S. Trachsel,
Hannah J. Stage,
Sebastian Rausch,
Susanne Trappe,
Katharina Söllig,
Gerhard Sponder,
Roswitha Merle,
Jörg R. Aschenbach,
Heidrun Gehlen
Affiliations
Dagmar S. Trachsel
Equine Clinic: Surgery and Radiology, Department of Veterinary Medicine, Freie Universität Berlin, Oertzenweg 19b, 14163 Berlin, Germany
Hannah J. Stage
Equine Clinic: Surgery and Radiology, Department of Veterinary Medicine, Freie Universität Berlin, Oertzenweg 19b, 14163 Berlin, Germany
Sebastian Rausch
Institute of Immunology, Department of Veterinary Medicine, Freie Universität Berlin, Robert-von-Ostertag-Str. 7, 14163 Berlin, Germany
Susanne Trappe
Institute of Veterinary Physiology, Department of Veterinary Medicine, Freie Universität Berlin, Oertzenweg 19b, 14163 Berlin, Germany
Katharina Söllig
Institute of Veterinary Physiology, Department of Veterinary Medicine, Freie Universität Berlin, Oertzenweg 19b, 14163 Berlin, Germany
Gerhard Sponder
Institute of Veterinary Physiology, Department of Veterinary Medicine, Freie Universität Berlin, Oertzenweg 19b, 14163 Berlin, Germany
Roswitha Merle
Institute for Veterinary Epidemiology and Biostatistics, Department of Veterinary Medicine, Freie Universität Berlin, Königsweg 67, 14163 Berlin, Germany
Jörg R. Aschenbach
Institute of Veterinary Physiology, Department of Veterinary Medicine, Freie Universität Berlin, Oertzenweg 19b, 14163 Berlin, Germany
Heidrun Gehlen
Equine Clinic: Surgery and Radiology, Department of Veterinary Medicine, Freie Universität Berlin, Oertzenweg 19b, 14163 Berlin, Germany
Physiological particularities of the equine heart justify the development of an in vitro model suitable for investigations of the species-specific equine cardiac electrophysiology. Adipose tissue-derived stromal/stem cells (ASCs) could be a promising starting point from which to develop such a cardiomyocyte (CM)-like cell model. Therefore, we compared abdominal, retrobulbar, and subcutaneous adipose tissue as sources for the isolation of ASCs applying two isolation methods: the collagenase digestion and direct explant culture. Abdominal adipose tissue was most suitable for the isolation of ASCs and both isolation methods resulted in comparable yields of CD45-/CD34-negative cells expressing the mesenchymal stem cell markers CD29, CD44, and CD90, as well as pluripotency markers, as determined by flow cytometry and real-time quantitative PCR. However, exposure of equine ASCs to 5-azacytidine (5-AZA), reportedly inducing CM differentiation from rats, rabbits, and human ASCs, was not successful in our study. More precisely, neither the early differentiation markers GATA4 and NKX2-5, nor the late CM differentiation markers TNNI3, MYH6, and MYH7 were upregulated in equine ASCs exposed to 10 µM 5-AZA for 48 h. Hence, further work focusing on the optimal conditions for CM differentiation of equine stem cells derived from adipose tissue, as well as possibly from other origins, are needed.
