Isolation and characterization of bone marrow-derived equine mesenchymal stem cells. Am J Vet Res

University of Cologne, Köln, North Rhine-Westphalia, Germany
American Journal of Veterinary Research (Impact Factor: 1.34). 11/2007; 68(10):1095-105. DOI: 10.2460/ajvr.68.10.1095
Source: PubMed


To isolate and characterize bone marrow-derived equine mesenchymal stem cells (MSCs) for possible future therapeutic applications in horses.
Equine MSCs were isolated from bone marrow aspirates obtained from the sternum of 30 donor horses.
Cells were cultured in medium (alpha-minimum essential medium) with a fetal calf serum content of 20%. Equine MSC features were analyzed to determine selfrenewing and differentiation capacity. For potential therapeutic applications, the migratory potential of equine MSCs was determined. An adenoviral vector was used to determine the transduction rate of equine MSCs.
Equine MSCs can be culture-expanded. Equine MSCs undergo cryopreservation in liquid nitrogen without altering morphologic characteristics. Furthermore, equine MSCs maintain their ability to proliferate and differentiate after thawing. Immunocytochemically, the expression of the stem cell marker CD90 can be detected on equine MSCs. The multilineage differentiation potential of equine MSCs was revealed by their ability to undergo adipogenic, osteogenic, and chondrogenic differentiation.
Our data indicate that bone marrow-derived stromal cells of horses can be characterized as MSCs. Equine MSCs have a high transduction rate and migratory potential and adapt to scaffold material in culture. As an autologous cell population, equine MSCs can be regarded as a promising cell population for tissue engineering in lesions of the musculoskeletal system in horses.

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    • "Musculoskeletal injuries are amongst the leading causes of early retirement or euthanasia of race, pleasure, and working horses (Arnhold et al. 2007). Bone fractures commonly occur and prove to be very serious injuries, due to the poor regenerative capacity of damaged cartilage, tendons, and ligaments. "
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    ABSTRACT: We investigated the applicability of single layer paper-based scaffolds for the three-dimensional (3D) growth and osteogenic differentiation of equine adipose-derived stem cells (EADSC), with comparison against conventional two-dimensional (2D) culture on polystyrene tissue culture vessels. Viable culture of EADSC was achieved using paper-based scaffolds, with EADSC grown and differentiated in 3D culture retaining high cell viability (>94 %), similarly to EADSC in 2D culture. Osteogenic differentiation of EADSC was significantly enhanced in 3D culture, with Alizarin Red S staining and quantification demonstrating increased mineralisation (p < 0.0001), and an associated increase in expression of the osteogenic-specific markers alkaline phosphatase (p < 0.0001), osteopontin (p < 0.0001), and runx2 (p < 0.01). Furthermore, scanning electron microscopy revealed a spherical morphology of EADSC in 3D culture, compared to a flat morphology of EADSC in 2D culture. Single layer paper-based scaffolds provide an enhanced environment for the in vitro 3D growth and osteogenic differentiation of EADSC, with high cell viability, and a spherical morphology.
    Biotechnology Letters 07/2015; 37(11). DOI:10.1007/s10529-015-1898-x · 1.59 Impact Factor
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    • "In horses, cells from equine bone marrow (eBM) and adipose tissue (eAT) have been isolated, and some researchers demonstrated their multilineage differentiation potential by the ability to undergo adipogenic, osteogenic, and chondrogenic differentiation [11] [13] [16]. However, only few authors determined immunophenotypic characterization of cells from eAT and eBM [17] [18] by flow cytometry, as requested for human MSCs by ISCT [15]. "
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    ABSTRACT: The aim of the present work was to isolate, cultivate, differentiate, and conduct cellular characterization of mesenchymal stem cells (MSCs) derived from equine adipose tissue (eAT) and bone marrow (eBM). Isolated and characterized cells were used in racehorses suffering from a superficial flexor tendon injury. Equine adipose tissue collection was performed at the base of the horse tail, whereas eBM was aspirated from iliac crest. Mononuclear cell fraction was isolated and cultured. In vitro differentiation and molecular characterization at P3 of culture were performed. No statistically significant differences in the number of cell doublings were found among different culture passages (P > .05). Doubling time was greater for eBM than eAT (3.2 ± 1.5 vs. 1.3 ± 0.7; P < .05). Positive von Kossa and Alizarin Red staining confirmed osteogenesis. Alcian Blue and Oil Red O staining illustrated chondrogenesis and adipogenesis, respectively. Isolated cells resulted positive for CD90, CD44, and CD105, whereas negative for hematopoietic markers, CD14, CD45, and CD34. Using isolated cells for injured tendon therapy, no adverse reactions were observed, and all inoculated horses returned to race competitions. In vitro results revealed the immunophenotypic characterization of isolated cells similar to that observed in human MSCs from the same sources; furthermore, in the present study, their clinical use proves the safety of eBM-derived and eAT-derived MSCs and a successful outcome for the treated animals that returned to their previous level of sport activity.
    Journal of Equine Veterinary Science 12/2014; 35(2). DOI:10.1016/j.jevs.2014.12.010 · 0.87 Impact Factor
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    • "Regarding the stem cell reservoirs, the most characterized sources of MSCs are bone marrow (BM) [10-17] and the adipose tissue [12,17]. Also, in 2002, MSCs from BM in the cat were isolated for the first time and these cells appeared to be very similar to those obtained from rodent and human sources [18], but the procedures employed to isolate these tissues are invasive and cells are usually obtained with low efficiency [18-20]. "
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    ABSTRACT: While amniotic mesenchymal cells have been isolated and characterized in different species, amniotic epithelial cells (AECs) have been found only in humans and horses and are recently considered valid candidates in regenerative medicine. The aim of this work is to obtain and characterize, for the first time in the feline species, presumptive stem cells from the epithelial portion of the amnion (AECs) to be used for clinical applications. In our study, we molecularly characterized and induced in vitro differentiation of feline AECs, obtained after enzymatic digestion of amnion. AECs displayed a polygonal morphology and the mean doubling time value was 1.94 ± 0.04 days demonstrating the high proliferating capacity of these cells. By RT-PCR, AECs expressed pluripotent (Oct4, Nanog) and some mesenchymal markers (CD166, CD44) suggesting that an epithelial-mesenchymal transition may occur in these cells that lack the hematopoietic marker CD34. Cells also showed the expression of embryonic marker SSEA-4, but not SSEA-3, as demonstrated by immunocytochemistry and flow cytometry. Moreover, the possibility to use feline AECs in cell therapies resides in their low immunogenicity, due to the absence of MHC-II antigen expression. After induction, AECs differentiated into the mesodermic and ectodermic lineages, demonstrating high plasticity. In conclusion, feline AECs appear to be a readily obtainable, highly proliferative, multipotent and non-immunogenic cell line from a source that may represent a good model system for stem cell biology and be useful in allogenic cell-based therapies in order to treat tissue lesions, especially with loss of substance.
    Stem Cell Research & Therapy 10/2013; 4(5):133. DOI:10.1186/scrt344 · 3.37 Impact Factor
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