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

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 cardi...

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Bibliographic Details
Published in:Animals (Basel) 2022-08, Vol.12 (16), p.2049
Main Authors: Trachsel, Dagmar S., Stage, Hannah J., Rausch, Sebastian, Trappe, Susanne, Söllig, Katharina, Sponder, Gerhard, Merle, Roswitha, Aschenbach, Jörg R., Gehlen, Heidrun
Format: Article
Language:English
Subjects:
Adipose tissue
adipose tissue differentiation
Azacytidine
Body fat
Cardiology
cardiomyocyte-like cells
Cardiomyocytes
CD29 antigen
CD34 antigen
CD44 antigen
CD45 antigen
Cell culture
Cell differentiation
Collagen
Collagenase
Differentiation
Digestion
DNA methylation
Electrophysiology
Explants
Flow cytometry
Heart
horse
Incubation
mesenchymal stem cells
Mesenchyme
Nkx2.5 protein
Pluripotency
preadipocytes
Rabbits
Stem cell transplantation
Stem cells
Tissue culture
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Summary: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.
ISSN:2076-2615
2076-2615
DOI:10.3390/ani12162049