Characterization of Equine Adipose Tissue–Derived Progenitor Cells Before and After Cryopreservation

Abstract

In horses, stem cell therapies are a promising tool to the treatment of many injuries, which are common consequences of athletic endeavor, resulting in high morbidity and often compromising the performance. In spite of many advantages, the isolation of stem cells similar to human, from equine adipose tissue, occurred only recently. The aim of this study was to isolate equine adipose tissue-derived progenitor cells (eAT-PC), to characterize their proliferative potential, and to study their differentiation capacity before and after cryopreservation. The cells, isolated from horse adipose tissue, presented similar fibroblast-like cell morphology in vitro. Their proliferation rate was evaluated during 63 days (23 passages) before and after cryopreservation. After the induction of osteogenic differentiation, von Kossa staining and positive immunostaining studies revealed the formation of calcified extracellular matrix confirming the osteogenic potential of these cells. Adipogenic differentiation was induced using two protocols: routine and other one developed by us, while our protocol requires a shorter time (Oil Red O staining revealed significant accumulation of lipid droplets after 7 days). Chondrogenic differentiation was observed after 21 days of induced pellet culture, as evidenced by histological (toluidine blue) and immunohistochemistry studies. Our data demonstrate that eAT-PC can be easily isolated and successfully expanded in vitro while presenting significant proliferating rate. These cells can be maintained undifferentiated in vitro and can efficiently undergo differentiation at least into mesodermal derivates. These eAT-PC properties were preserved even after cryopreservation. Our findings classify eAT-PC as a promising type of progenitor cells that can be applied in different cell therapies in equines.

Isolation and differentiation potential of an equine amnion-derived stromal cell line

“…Cells showed the potential for in vitro differentiation into adipocytes as demonstrated by the presence of red granules identified as lipid droplets in the cytoplasm, similar to those already observed in horse adipose tissue-derived stem cell (Mambelli et al 2009) and stem cells differentiated from umbilical cord blood (Koch et al 2007). The amnion-derived stromal cells also had the potential to be differentiated into This study showed that horse amnion represents a valuable source of stromal cells alternative to bone marrow and fat stem cells.…”

Isolation, expansion and differentiation of cellular progenitors obtained from dental pulp of agouti (Dasyprocta prymnolopha Wagler, 1831)

“…The differentiation media tested (Stem cell Pro ® kit) demonstrated potential to induce adipogenic and osteogenic differentiation of agouti dental pulp stem cells of agoutis. The protocol for media changes followed the criteria previously reported by Mambelli et al (2009). Both the osteogenic and adipogenic differentiations were visualized by morphological changes observed for a phenotype similar to osteoblasts and adipocytes, respectively.…”

Characterization of mesenchymal stem cells derived from equine adipose tissue

“…Several studies reported the characterization of AdMSCs in horses, where the most common processes are osteogenic, adipogenic and chondrogenic (tri-lineage) differentiation of progenitor cells (Vidal et al, 2007(Vidal et al, , 2008Kisiday et al, 2008;Mambelli et al, 2009;Colleoni et al, 2009;Toupadakis et al, 2010;Ahern et al, 2011;Schwarz et al, 2011a,b). Only five recent studies have reported the analyses of surface antigens of AdMSCs by flow cytometry (de Mattos Carvalho et al, 2009;Braun et al, 2010;Pascucci et al, 2011;Raabe et al, 2011;Ranera et al, 2011).…”