Cytodifferentiation of the mouse secondary palate in vitro: morphological, biochemical, and histochemical aspects.

Previous investigations had indicated that the epithelial cells at the medial surface of the palate differentiate prior to fusion. An increase in adhesiveness and signs of partial autolysis were reported to be part of the differentiation. It was not determined, however, whether or not the autolytic processes may result in the total lysis of the cells, even if fusion is prevented. This problem was approached in the present investigation by culturing shelves in isolation, thus preventing fusion. The morphological properties of the presumptive fusing cells were then observed during and after the actual time of fusion. Differentiation was assessed in vivo at equivalent developmental ages. In addition, the role of the lysosomes in the process of fusion was investigated. This was done by measuring the activities of two lysosomal enzymes, glucosaminidase and arylsulfatase, before, during, and after fusion in vivo and at the equivalent times in vitro. Glucosaminidase activity was also localized histochemically. The in vitro and in vivo differentiation were similar. The cells of the mesenchyme produced intramembranous bone and loose mesenchyme. The nasal surface differentiated into a pseudostratified columnar ciliated epithelium, while the oral surface became stratified squamous epithelium. At the medial surface, however, the morphological changes during fusion in vivo were different from those observed at the equivalent time in vitro. Few, if any, of the presumptive fusing cells forming a bi-layered or multilayered epithelium degenerated in the explants, while many cells forming the epithelial seam in vivo underwent complete degeneration. In addition, after fusion, the medial surface of the palatal explants was covered by a stratified squamous and/or pseudostratified columnar ciliated epithelia. In vivo, the mesenchyme merged as a consequence of the total destruction of the seam. The specific activity of glucosaminidase increased progressively, reaching a peak three days after fusion. In all cases no statistically significant differences were found between the in vivo and in vitro values. Arylsulfatase specific activity remained the same both in vivo and in vitro. Histochemically glucosaminidase activity was localized in the cells undergoing degeneration during fusion. It is concluded that fusion is a necessary condition for the completion of the autolysis of the presumptive fusing cells. The cells may alter their developmental fate, forming a stratified squamous and/or pseudostratified columnar epithelia, if fusion is prevented.