The present investigation describes the peculiar properties of AECs, which have a diffuse expression of adhesion molecules CD29 and CD49 that are likely responsible for the high plating efficiency of these cells and account for the quick establishment of intercellular connections that result in the in vitro formation of three-dimensional spheroids as reported for human amniotic stem cells (Yen et al 2005). Moreover, the cells display a diffuse expression of stemness markers that is consistent with the rapid osteogenic differentiation that the cells undergo when exposed to inducing medium. Osteogenic differentiation can be achieved after prolonged expansion in vitro, and this process can be potentiated by simple molecules, such as 17β-estradiol, as described by others (Olivier et al. 2006; Kawaguchi et al 2005), and that could be used to improve the regenerative potential of the cells. Collectively, the data reported for the sheep model confirm that AECs represent a promising source of stem cells for the development of regenerative strategies. This investigation has demonstrated that after prolonged expansion, both the expression of stemness markers and osteogenic plasticity are reduced, thus suggesting that improved culture conditions are required whenever an extended amplification in vitro is necessary to obtain very large cell numbers.