An analysis of the aggregation and morphogenesis of area opaca endoderm cells from the primitive-streak chick embryo

The aggregative behaviour and subsequent morphogenesis of extra-embryonic endoderm cells from primitive-streak chick embryos have been investigated. A relatively pure population of area opaca endoderm cells was obtained by differential dissociation, which involves partial separation of epiblast and endoderm cell clumps by sieving through Nitex mesh. For aggregation studies cells were cultured in rotating flasks in Leibovitz (L-15) medium, in saline or in saline supplemented with glucose (1 mg/ml). Aggregation was monitored using the Coulter Counter. In these three media aggregation is rapid; by 10 min an average of 61% of the population had aggregated, to reach a plateau at 30 min when an average percent adhesion value of 83% was obtained. The aggregates in L-15 medium were large and compact. After several days in culture, they cavitated and formed smooth hollow vesicles with thin walls composed of one or a few cell layers. Aggregates formed in PCS were smaller and looser in appearance; the addition of glucose resulted in a certain degree of compaction. Some morphogenesis occurred under these conditions with the aggregates developing numerous irregular cavities. These experiments suggest that some of the factors that affect cell adhesion in early embryonic cells can be studied in vitro. The results also indicate that the ability to cavitate is an intrinsic property of the endoderm cells of the area opaca since this occurs in the absence of epiblast or mesoderm.