Comparison of equine bone marrow-, umbilical cord matrix and amniotic fluid-derived progenitor cells

Abstract

The aim of the study was to compare in vitro the stemness features of horse progenitor cells derived from bone marrow (BM-MSCs), amniotic fluid (AF-MSCs) and umbilical cord matrix (EUC-MSCs). It has been suggested that there may be a stem cell population within both umbilical cord matrix and amniotic fluid. However, little knowledge exists about the characteristics of these progenitor cells within these sources in the equine species. This study wanted to investigate an alternative and non-invasive stem cell source for the equine tissue engineering and to learn more about the properties of these cells for future cell banking. Bone marrow, umbilical cord and amniotic fluid samples were harvested from different horses. Cells were analyzed for proliferation, immunocytochemical, stem cell gene expression and multilineage plasticity. BM- and AF-MSCs took similar time to reach confluence and showed comparable plating efficiency. All cell lines expressed identical stem cell markers and capability to differentiate towards osteogenic lineage. Almost all cell lines differentiated into the adipogenic lineage as demonstrated by cytochemical staining, even if no adipose gene expression was detectable for AF-MSCs. AF- and EUC-MSCs showed a limited chondrogenic differentiation compared with BM-MSCs as demonstrated by histological and biochemical analyses. These findings suggest that AF-MSCs appeared to be a readily obtainable and highly proliferative cell line from an uninvasive source that may represent a good model system for stem cell biology. More studies are needed to investigate their multilineage potential. EUC-MSCs need to be further investigated regarding their particular behavior in vitro represented by spheroid formation.

Mesenchymal stem cells from amnion and amniotic fluid in the bovine

“…This was evident during the primary culture (at P0), when fibroblastic-like or flatter and circular cells were recognized. By the first passage, however, a selection of spindle-shaped cells occurred, according to what has been previously observed in equine species by our group (Lovati et al 2011) or other researchers (Iacono et al 2012a). Amniotic epithelial cells displayed the typical polygonal morphology reported for human (Miki et al 2005(Miki et al , 2007a and equine species (Lange-Consiglio et al 2012).…”

“…Our findings suggest, for the first time, that bovine amnion and AF represent alternative sources of progenitor cells to employ in cell-based therapies, in a similar manner to several other species (Chen et al 2011, Lovati et al 2011, Iacono et al 2012a, 2012b, Lange-Consiglio et al 2012. Further studies, including pre-clinical and a deeper evaluation of immunological properties, are needed for the in vivo applications in order to better understand their role in cellular therapy.…”

“…Samples were transported to the laboratory at 4 8C and processed within 8 h. Two AFs were collected into sterile syringes during the full-term delivery of different cattle as previously reported (Lovati et al 2011). When it was possible, the AF was aspirated into a sterile syringe directly from the vulva protruded amniotic sac before its spontaneous rupture.…”

“…Isolation of MSCs from AF was performed following the procedures previously described (Lovati et al 2011), consisting of centrifuging the samples at 400 g for 10 min, removing the supernatant and collecting the pellet formed at the bottom of the tube. Before seeding the cells, the pellet was washed three times in PBS.…”

“…As such, gestational tissues including umbilical cord blood (Kern et al 2006, Koch et al 2007, Guest 2008, Reed & Johnson 2008, Bartholomew et al 2009, Schuh et al 2009, Seo et al 2009, Raoufi et al 2011, umbilical cord tissues (Mitchell et al 2003, Carlin et al 2006, Hoynowski et al 2007, Cremonesi et al 2008, Passeri et al 2009, Lovati et al 2011, Iacono et al 2012a, amniotic tissues (Filioli Uranio et al 2011, Iacono et al 2012a, 2012b, Lange-Consiglio et al 2012, and amniotic fluid (AF) (Chen et al 2011, Dev et al 2011, Lovati et al 2011, Iacono et al 2012a) have been recently suggested, also in veterinary medicine, as appealing candidates for the derivation of MSCs to use in cell biotechnology. The human lesson teaches that presumptive adult stem cells derived from gestational tissues retain highest proliferation capacity, longest telomere length, broadest differentiation, and extensive proliferative potential when compared with cells obtained from adult tissues (Kogler et al 2004, Kern et al 2006.…”