Fetal adnexa are a noncontroversial source of mesenchymal stem cells (MSCs) with high plasticity, proliferation rate and ability to differentiate towards multiple lineages. Mesenchymal SCs have been characterized for both their stemness and their differentiation abilities. Recently, the scientific debate has focused on MSCs selection and on establishing predictable elements to discriminate the cells with most promising potential in regenerative medicine. In this study, we characterized and followed in vitro proliferation and differentiation potency of canine amniotic membrane MSCs (AM-MSCs) and umbilical cord matrix MSCs (UCM-MSCs) isolated from fetuses at early (35-40 days) and late (45-55 days) gestational ages. We found that cells from both fetal gestational ages showed similar features. In all examined cell lines, the morphology of proliferating cells typically appeared fibroblast-like and the population doubling of cells, cultured up to passage 10, significantly increased with passage number. In both cell types, cell viability and chromosomal number and structure were not affected by gestational age. In AM- and UCM-MSCs of both gestational phases, the expression of embryonic (Oct-4) and mesenchymal stemness (CD29, CD44) markers was observed. Hematopoietic and histocompatibility markers were never found in any sample. Cells of the two cell types at P3 showed multipotent abilities and differentiated to neurocytes and osteocytes, as demonstrated by specific stains and molecular analysis. These results indicated that MSCs retrieved from UCM and AM in early and late fetal phase of gestation could be used in regenerative medicine approaches in the dog.
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