Terminal differentiation of palatal medial edge epithelial cells in vitro is not necessarily dependent on palatal shelf contact and midline epithelial seam formation

Abstract

During fusion of the mammalian secondary palate, it has been suggested that palatal medial edge epithelial (MEE) cells disappear by means of apoptosis, epithelial-mesenchymal transformation (EMT) and epithelial cell migration. However, it is widely believed that MEE cells never differentiate unless palatal shelves make contact and the midline epithelial seam is formed. In order to clarify the potential of MEE cells to differentiate, we cultured single (unpaired) palatal shelves of ICR mouse fetuses by using suspension and static culture methods with two kinds of gasmixtures. We thereby found that MEE cells can disappear throughout the medial edge even without contact and adhesion to the opposing MEE in suspension culture with 95% O 2 /5% CO 2 . Careful examination of MEE cell behavior in the culture revealed that apoptosis, EMT, and epithelial cell migration all occurred at various stages of MEE cell disappearance, including the transient formation and disappearance of epithelial triangles and islets. In contrast, MEE cells showed poor differentiation in static culture in a CO 2 incubator. Furthermore, mouse and human amniotic fluids were found to prevent MEE cell differentiation in the cultured single palatal shelf, although paired palatal shelves fused successfully even in the presence of amniotic fluid. We therefore conclude that terminal differentiation of MEE cells is not necessarily dependent on palatal shelf contact and midline epithelial seam formation, but such MEE cell differentiation appears to be prevented in utero by amniotic fluid unless palatal shelves make close contact and the midline epithelial seam is formed.

The mesenchymal cell, its role in the embryo, and the remarkable signaling mechanisms that create it

“…12A-C); the midline seam basal epithelial cells do not stain for apoptosis (Nawshad et al, 2004) in vivo. In vitro, inappropriate culture conditions are responsible for much of the reported basal cell death (Takigawa and Shiota, 2004). Palates express Snail (Martinez- Alvarez et al, 2004), which promotes cell survival, as well as EMT (Vega et al, 2004).…”

Amniotic fluid induces rapid epithelialization in the experimentally ruptured fetal mouse palate - implications for fetal wound healing

“…Recently, we found that MEE cells can disappear from the medial edge independently of palatal shelf contact and midline seam formation under the amniotic fluid-free condition and that amniotic fluid prevents the disappearance of MEE cells unless palatal shelves make contact and the midline epithelial seam is formed (Takigawa and Shiota, 2004). insight into the unique biological actions of amniotic fluid in the repair of the developing fetal palate when it is torn apart after palatal fusion.…”

“…It has been claimed that MEE cells never disappear unless palatal shelves make close contact with each other and a midline epithelial seam is formed (Griffith and Hay, 1992), although the actual fate of MEE cells is still controversial (Fitchett and Hay, 1989;Carette and Ferguson, 1992;Cuervo and Covarrubias, 2004;Takigawa and Shiota, 2004;Vaziri Sani et al, 2005;Dudas et al, 2006: Xu et al, 2006Jin and Ding, 2006). Recently, we found that MEE cells can disappear from the medial edge independently of palatal shelf contact and midline seam formation under the amniotic fluid-free condition and that amniotic fluid prevents the disappearance of MEE cells unless palatal shelves make contact and the midline epithelial seam is formed (Takigawa and Shiota, 2004).…”

“…Amniotic fluid forms the major environment of mammalian fetuses grown in utero and has been suggested to play important roles in palatal fusion (Pourtois, 1968;Goss, 1976;Nanda et al, 1977;Takigawa and Shiota, 2004) as well as in development of other fetal tissues, such as the lung (Sundell et al, 1980;Yoshimura et al, 2000) and the gastro-intestinal tract (Mulvihill et al, 1985(Mulvihill et al, , 1986Kelly et al, 1997). In addition, amniotic fluid has been suggested to play beneficial roles in fetal wound healing (Mast et al, 1992;Adzick and Lorenz, 1994;Özgenel et al, 2001;Levinson et al, 2001), although the mechanism is poorly understood.…”

“…For this purpose, we artificially separated the fused palate of mouse fetuses into halves along the midline and cultivated them in the presence or absence of mouse or human amniotic fluid in suspension culture (Shiota et al, 1990b;Takigawa and Shiota, 2004). Using this experimental model, we investigated the biological effects of amniotic fluid on the injured fetal palate.…”