BRANCHING MORPHOGENESIS OF MOUSE EMBRYONIC SUBMANDIBULAR EPITHELIA CULTURED UNDER 3 DIFFERENT CONDITIONS

To investigate how the mesenchyme interacts with the epithelium, we em ployed three different culture systems: System A, in which intact subm andibular gland rudiments at the mid 13-day stage were cultured on Mil lipore filters; System B, in which the 13-day epithelium and mesenchym e were separated once with dispase, recombined again, and cultured on the filter; System C, in which the separated 13-day epithelium was clo tted with Matrigel and cultured with the mesenchyme across the filter or in the presence of EGF instead of the mesenchyme. In Systems A and B, 13-day epithelia expanded and produced similar lobules with narrow clefts and stalk. When the 13-day epithelium was cultured in System C under the influence of the mesenchyme, it formed rather oval lobules w ith stalk that were superficially similar to those in System A, but na rrow clefts, as seen in the intact early 13-day grand, were rarely fou nd in System C. Furthermore no long stalk formation was observed when EGF was introduced in place of the mesenchyme. A bacterial collagenase from Clostridium histolyticum gave a considerable inhibition of branc hing of the 13-day epithelium in Systems A and B, but no significant i nhibition was observed in System C when the mesenchyme or EGF was empl oyed as the source of diffusible factor(s). In contrast, although the 13-day epithelium was significantly resistant to the action of heparit inase I from Flavobacterium heparinum in Systems A and B, the enzyme a lmost completely inhibited the expansion and branching of the epitheli um in System C. Judging from these observations, we conclude that the mechanisms of lobular formation in Systems A and B are not the same as those in System C, where the epithelium is clotted with basement memb rane matrix components during tissue culture.