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Three-dimensional (3D) culture provides a biomimicry of the naive microenvironment that can support cell proliferation, differentiation, and regeneration. Some growth factors, such as epidermal growth factor (EGF), facilitate normal meiosis during oocyte maturation in vivo. In this study, a scaffold-based 3D coculture system using purified alginate was applied to induce oocyte differentiation from mouse embryonic stem cells (mESCs). mESCs were induced to differentiate into oocyte-like cells using embryoid body protocol in the two-dimensional or 3D microenvironment in vitro. To increase the efficiency of the oocyte-like cell differentiation from mESCs, we employed a coculture system using ovarian granulosa cells in the presence or absence of epidermal growth factor (+EGF or -EGF) for 14 days and then the cells were assessed for germ cell differentiation, meiotic progression, and oocyte maturation markers. The cultures exposed to EGF in the alginate-based 3D microenvironment showed the highest level of premeiotic (Oct4 and Mvh), meiotic (Scp1, Scp3, Stra8, and Rec8), and oocyte maturation (Gdf9, Cx37, and Zp2) marker genes (p < .05) in comparison to other groups. According to the gene-expression patterns, we can conclude that alginate-based 3D coculture system provided a highly efficient protocol for oocyte-like cell differentiation from mESCs. The data showed that this culture system along with EGF improved the rate of in vitro oocyte-like cell differentiation.

Fresh human breast milk consists of a heterogeneous population of cells that may offer a non-invasive source of cells for therapeutic proposes. The aims of this study were to characterize the breast milk-derived cells cultured in vitro. To do this, the cells from human breast milk were cultured and the expression of the CD markers along with the embryonic stem cell markers, endothelial and luminal mammary epithelial cell markers was evaluated by flow cytometry and immunofluorescence. The presence of fetal microchimerism among the isolated cells was also determined by the presence of SRY gene. They were also differentiated into adipocytes and osteoblasts. The results showed that a remarkable number of cells expressed the mesenchymal stem cell (MSC) markers such as CD90, CD44, CD271, and CD146. A subpopulation of the human breast milk-derived cells (HBMDC) also expressed the embryonic stem cell markers, such as TRA 60-1, Oct4, Nanog and Sox2 but not SSEA1 or 4. The frequencies of the cells which expressed the endothelial, hematopoietic cell markers were negligible. SRY gene was not detected in the breast milk isolated cells. A subpopulation of the cells also expressed cytokeratin 18, the marker of luminal mammary epithelial cells. These cells showed the capability to differentiate into adipocytes and osteoblasts. In conclusion, these finding highlighted the presence of cells with various sources in the breast milk. Different stem cells including MSCs or embryonic stem cell-like cell along with the exfoliated cells from luminal epithelial cells were found among the isolated cells. The breast milk-derived stem cells might be considered as a non-invasive source of the stem cells for therapeutic purpose.

Fresh human breast milk consists of a heterogeneous population of cells that may offer a non-invasive source of cells for therapeutic proposes. The aims of this study were to characterize the breast milk-derived cells cultured in vitro. To do this, the cells from human breast milk were cultured and the expression of the CD markers along with the embryonic stem cell markers, endothelial and luminal mammary epithelial cell markers was evaluated by flow cytometry and immunofluorescence. The presence of fetal microchimerism among the isolated cells was also determined by the presence of SRY gene. They were also differentiated into adipocytes and osteoblasts. The results showed that a remarkable number of cells expressed the mesenchymal stem cell (MSC) markers such as CD90, CD44, CD271, and CD146. A subpopulation of the human breast milk-derived cells (HBMDC) also expressed the embryonic stem cell markers, such as TRA 60-1, Oct4, Nanog and Sox2 but not SSEA1 or 4. The frequencies of the cells which expressed the endothelial, hematopoietic cell markers were negligible. SRY gene was not detected in the breast milk isolated cells. A subpopulation of the cells also expressed cytokeratin 18, the marker of luminal mammary epithelial cells. These cells showed the capability to differentiate into adipocytes and osteoblasts. In conclusion, these finding highlighted the presence of cells with various sources in the breast milk. Different stem cells including MSCs or embryonic stem cell-like cell along with the exfoliated cells from luminal epithelial cells were found among the isolated cells. The breast milk-derived stem cells might be considered as a non-invasive source of the stem cells for therapeutic purpose.