PLURIPOTENT AND DEVELOPMENTALLY RESTRICTED NEURAL-CREST-DERIVED CELLS IN POSTERIOR VISCERAL ARCHES

The early migratory cells of the posterior rhombencephalic neural crest consist of a heterogeneous population of pluripotent and developmentally restricted neural crest cells (Ito and Sieber-Blum, Dev. Biol. 148, 95-106, 1991). To determine if changes in developmental capacities occur during migration, the developmental potentials of posterior visceral arch mesenchymal cells were investigated by in vitro clonal analysis. Most of these cells consisted of the post-migratory cells of the posterior rhombencephalic neural crest. Four morphologically distinct types of clones were observed, and the cells within these clones expressed characteristic phenotypes as shown by their binding of antibodies against cell type-specific markers: (1) “DP” clones consisted of densely packed polygonal cells, with flattened large cells located predominantly at the periphery of these clones. Immunocytochemical analyses showed that DP clones contained smooth muscle cells, connective tissue cells, chondrocytes, and serotonin (5-HT)-positive neurons, and over 90% of the cells per clone were HNK-1 positive. This suggests that DP clone-forming cells are pluripotent neural-crest-derived cells with the capacity to develop into ectomesenchymal derivatives and serotonergic neurons. (2) “DS” clones consisted of densely packed spindle-shaped cells. These clones included smooth muscle cells, connective tissue cells, and chondrocytes. By contrast, neuronal phenotypes were not present. An average of 0.4% of the cells per clone were HNK-1 positive. DS clones appear to be formed by neural-crest-derived cells that are partially restricted, expressing ectomesenchymal phenotypes only. (3) “DP” clones consisted of densely packed small cells and flattened large cells. Although no HNK-1-positive cells were found in DF clones, these clones contained connective tissue cells and/or smooth muscle cells. DF clones appear to be derived from bipotent cells with the ability to differentiate into connective tissue cells and smooth muscle cells, or cells committed to the connective tissue cell lineage. (4) “LF” clones consisted of loosely arranged, flattened large cells. These clones did not contain HNK-1-positive cells. The clones consisted entirely of smooth muscle cells. Therefore, LF clones are most likely formed by precursors that are committed to the smooth muscle cell lineage. These results indicate the presence of pluripotent neural-crest-derived cells, cells with a restricted developmental potential, and apparently committed cells in the posterior visceral arch. Pluripotent cells can generate up to four neuronal and non-neuronal phenotypes. Other cells are restricted to ectomesenchymal cell types, and the proportion of these cells in the posterior visceral arch changes with progressing embryonic development. The present data in combination with our previous study (Ito and Sieber-Blum, Dev. Biol. 148, 95-106, 1991) suggest a characteristic pattern of lineage segregation during posterior rhombencephalic neural crest development, indicating that specification of some cell types can occur at the anal sites of the neural crest cell localization. © 1993 by Academic Press, Inc.