Stemness Signature of Equine Marrow-derived Mesenchymal Stem Cells

BACKGROUND: Application of competent cells such as mesenchymal stem cells (MSCs) for treatment of musculoskeletal disorders in equine athletes is increasingly needed. Moreover, similarities of horse and human in size, load and types of joint injuries, make horse as a good model for MSCs therapy studies. This study was designed to isolate and characterize stemness signature of equine bone marrow-derived mesenchymal stem cells (BM-MSCs). METHODS: BM of three mares was aspirated and the mononuclear cells (MNCs) were isolated using density gradient. The primary MNCs were cultured and analyzed after tree passages (P3) for growth characteristics, differentiation potentials, and the expression of genes including CD29, CD34, CD44, CD90, CD105, MHC-I, MHC-II and pluripotency related genes (Nanog, Oct-4, Sox-2, SSEA-1, -3, -4) using RT-PCR or immunocytochemistry techniques. RESULTS: The isolated cells in P3 were adherent and fibroblast-like in shape with doubling times of 78.15 h. Their clonogenic capacity was 8.67±4% and they were able to differentiate to osteogenic, adipogenic and chondrogenic lineages. Cells showed expression of CD29, CD44, CD90, MHC-I and Sox-2 while no expression for CD34, MHC-II, CD105, and pluripotency stemness markers was detected. CONCLUSIONS: In conclusion, data showed that isolated cells have the basic and minimal criteria for MSCs, however, expressing only one pluripotency gene (sox-2).

Equine adipose-derived mesenchymal stem cells: phenotype and growth characteristics, gene expression profile and differentiation potentials

Because of the therapeutic application of stem cells [SCs], isolation and characterization of different types of SCs, especially mesenchymal stem cells [MSCs], have gained considerable attention in recent studies. Adipose tissue is an abundant and accessible source of MSCs which can be used for tissue engineering and in particular for treatment of musculoskeletal disorders. This study was aimed to isolate and culture equine adipose-derived MSCs [AT-MSCs] from little amounts of fat tissue samples and determine some of their biological characteristics. In this descriptive study, only 3-5 grams of fat tissue were collected from three crossbred mares. Immediately, cells were isolated by mechanical means and enzymatic digestion and were cultured in optimized conditions until passage 3 [P3]. The cells at P3 were evaluated for proliferative capacities, expression of specific markers, and osteogenic, chondrogenic and adipogenic differentiation potentials. Results showed that the isolated cells were plastic adherent with a fibroblast-like phenotype. AT-MSCs exhibited expression of mesenchymal cluster of differentiation [CD] markers [CD29, CD44 and CD90] and not major histocompatibility complex II [MHC-II] and CD34 [hematopoietic marker]. Cellular differentiation assays demonstrated the chondrogenic, adipogenic and osteogenic potential of the isolated cells. Taken together, our findings reveal that equine MSCs can be obtained easily from little amounts of fat tissue which can be used in the future for regenerative purposes in veterinary medicine