International Journal of Stem Cells : eISSN 2005-5447

Fig. 2.

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Fig. 2. Formation of diverse cellular structures from the leaflet during 12 weeks of tissue culture (A~P). Outgrowth of adherent fibroblast-like cells started a few hours after placing the leaflet tissue in the culture; these fibroblast-like cells stabilized the attachment of the leaflet tissue to the culture dish and provided a feeder layer for non-adherent cells (A). Retraction of the confluent feeder layer created a space (B) for formation of multinucleated syncytium-like structures (C) and a niche area (D). By the second week, bubble-like structures of differing sizes and shapes were detected (E). The phenotype of these structures was similar to those of oocytes, 2-cell embryos, 3-cell embryos (F), 4-cell embryos (G), and morulae (H). Formation of large dense colonies and hollow spheres occurred in the confluent cultures between week 3 and 12 (I). The dense colonies were formed either by recruiting the non-adherent cells to the morula-like structures (J) or by the retraction of the mature niche area (K). Interconnecting bundles were generated between adjacent niche syncytia (L, arrow), a process that often was followed by unification of the two structures to form a larger colony. Phalloidin staining (green) and DAPI nuclear counterstaining (blue) showed the structure of the embryonic-like cells and colonies. Oocyte-like cells exhibited a ring-shaped structure with an elongated and peripheral nucleus (M). Multiple nuclei were located on the periphery of 2-cell-embryo-like structure (N), indicating the possibility that other non-adherent cells would be arranged around these structures. Two-dimensional imaging of one blastula-like structure showed a round aggregation of cells with distinct borders between the cells (O). The unification of two adjacent niche areas was promoted by cytoplasmic fusion (P).
International Journal of Stem Cells 2018;11:121-30
© 2018 International Journal of Stem Cells