Organization of the membrane skeleton in spreading mouse blastomeres. I. Morphological analysis.

Differentiation in the mouse embryo begins at the 8-cell stage when the blastomeres spread against each other in a process called compaction. The spreading behavior of blastomeres on lectin-coated coverslips mimics that of blastomeres in the embryo, and we have utilized this model system to obtain an en face view of the membrane skeleton in the spreading blastomeres. Embryos were cultured on the coverslips for periods ranging from 20 sec to 6 hr, and the cells were disrupted to expose the cytoplasmic face of the adherent membranes and their associated filaments. The "membrane lawn" preparations were fixed, critical point dried, rotary shadowed, and the replicas examined by transmission electron microscopy. Using this technique we found that the plasmalemma of rounded blastomeres is associated with a lacy 3-dimensional filamentous meshwork that is transformed into a thin mat of densely woven filaments when the cells flatten. The overall organization of the membrane skeleton is similar in flattening 2- and 8-cell embryos, but there are significant differences in the time required for spreading to take place, in the means whereby the membrane skeletons are reorganized, and in the extent of maximal flattening. The significance of these observations for the compaction process is discussed.

Epidermal growth factor or transforming growth factor alpha is required for kidney tubulogenesis in matrigel cultures in serum-free medium.

The ability of matrigel, a reconstituted basement membrane gel, to induce the differentiation of baby mouse kidney cells has been examined in a hormonally defined serum-free medium. Primary cultures of baby mouse kidney cells were observed to form tubules over a time interval of 1-2 weeks in matrigel. Electron microscopic studies showed that tubules with lumens were present, and the tubule morphology was similar to that of the collecting duct. When using matrigel from which the growth factors had been removed, tubule formation no longer occurred, unless the medium was further supplemented with epidermal growth factor (10 ng/ml). Transforming growth factor alpha stimulated tubule formation as effectively as epidermal growth factor, whereas transforming growth factor beta had an inhibitory effect on tubule formation. These data suggest that both an extracellular matrix and specific growth factors may regulate kidney differentiation during development.