DNA synthesis and cell generation cycle during determination and differentiation of the metanephric mesenchyme

Abstract

Isolated metanephrogenic mesenchyme can be induced for epithelial differentiation and tubule formation by an embryonic spinal cord that is cultivated in a transfilter position to the target cells. Under such conditions, determination of the cells starts at 12 hr and is completed around 24 hr. To correlate this event to DNA synthesis, thymidine incorporation was followed in such cultures from 2 to 96 hr after setting up the transfilter explants. Controls were cultured under conditions which did not result in induction. Incorporation into both induced and noninduced mesenchymes dropped rapidly during the first 12 hr when the tissues were transferred to the in vitro conditions. During the subsequent critical 12- to 24-hr period, mesenchymes continuously exposed to an inductor showed a marked increase in their incorporation of thymidine. If induction was interrupted during the first 10 hr there was no stimulation of incorporation when measured at 24 hr. Similarly, isolated mesenchymes and those exposed to a tissue devoid of inducing capacity did not show stimulation, and their incorporation remained at a low level when measured at 24 hr. Parallel DNA measurements were made by flow cytometry of individual nuclei from mesenchymes exposed to the inductor from 0 to 46 hr. No major changes in the distribution of the various phases of the cell generation cycle were detected nor could any synchronization be shown. Hence the changes in the incorporation rate of thymidine seem to be due to variations in the total length of the cell cycle. We conclude that induction of differentiation of the metanephric mesenchyme is accompanied by a contact-mediated stimulation of DNA synthesis. The stimulation may represent a permissive effect on a predetermined cell population in the metanephrogenic mesenchyme.