Department of Veterinary Medical Sciences, University of Bologna, via Tolara di Sopra 50, 40064 Ozzano Emilia, Bologna, Italy.
Reproduction. 2012 Apr;143(4):455-68. doi: 10.1530/REP-10-0408. Epub 2012 Jan 24.
Mesenchymal stem cells (MSCs) have been derived from multiple sources of the horse including umbilical cord blood (UCB) and amnion. This work aimed to identify and characterize stem cells from equine amniotic fluid (AF), CB and Wharton's Jelly (WJ). Samples were obtained from 13 mares at labour. AF and CB cells were isolated by centrifugation, while WJ was prepared by incubating with an enzymatic solution for 2 h. All cell lines were cultured in DMEM/TCM199 plus fetal bovine serum. Fibroblast-like cells were observed in 7/10 (70%) AF, 6/8 (75%) CB and 8/12 (66.7%) WJ samples. Statistically significant differences were found between cell-doubling times (DTs): cells isolated from WJ expanded more rapidly (2.0±0.6 days) than those isolated from CB (2.6±1.3 days) and AF (2.3±1.0 days) (P<0.05). Positive von Kossa and Alizarin Red S staining confirmed osteogenesis. Alcian Blue staining of matrix glycosaminoglycans illustrated chondrogenesis and positive Oil Red O lipid droplets staining suggested adipogenesis. All cell lines isolated were positive for CD90, CD44, CD105; and negative for CD34, CD14 and CD45. These findings suggest that equine MSCs from AF, UCB and WJ appeared to be a readily obtainable and highly proliferative cell lines from a uninvasive source that may represent a good model system for stem cell biology and cellular therapy applications in horses. However, to assess their use as an allogenic cell source, further studies are needed for evaluating the expression of markers related to cell immunogenicity.
间充质干细胞（MSCs）已从马的多个来源中分离得到，包括脐带血（UCB）和羊膜。本研究旨在从马的羊水（AF）、CB 和 Wharton 胶中鉴定和表征干细胞。从 13 匹分娩的母马中获得样本。通过离心分离 AF 和 CB 细胞，而 WJ 则通过用酶溶液孵育 2 小时来制备。所有细胞系均在 DMEM/TCM199 加胎牛血清中培养。在 7/10（70%）的 AF、6/8（75%）的 CB 和 8/12（66.7%）的 WJ 样本中观察到成纤维细胞样细胞。细胞倍增时间（DT）存在统计学差异：从 WJ 分离的细胞比从 CB（2.6±1.3 天）和 AF（2.3±1.0 天）分离的细胞增殖更快（2.0±0.6 天）（P<0.05）。成骨诱导的 von Kossa 和茜素红 S 染色阳性，基质糖胺聚糖的阿尔辛蓝染色表明软骨形成，油红 O 脂质滴染色阳性提示脂肪生成。分离得到的所有细胞系均表达 CD90、CD44、CD105，不表达 CD34、CD14 和 CD45。这些发现表明，来自 AF、UCB 和 WJ 的马 MSC 似乎是一种从非侵入性来源获得的易于获得且高增殖的细胞系，可能代表马干细胞生物学和细胞治疗应用的良好模型系统。然而，为了评估它们作为同种异体细胞来源的用途，需要进一步研究来评估与细胞免疫原性相关的标志物的表达。
