Reversible release of chick embryo fibroblast cultures from density dependent inhibition of growth

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Abstract
Initiation of proliferation in density‐inhibited chick embryo fibroblast cultures induced by insulin or trypsin was partially reversed by replacing the medium with supernatants from parallel non‐stimulated cultures. Growth stimulation by neuraminidase, pokeweed mitogen, bacterial lipo polysaccharide or purified tuberculin was less, or not at all, affected by this procedure. Medium change per se caused some proliferation in non‐stimulated cultures. Increased rate of sugar uptake in insulin‐stimulated cultures returned to the level of that in non‐stimulated cultures within a few hours after medium change. This reversion took place apparently irrespective of the phase of the cell cycle. Replacing the medium with supernatants from non‐stimulated cultures induced a rapid decline in subsequent thymidine incorporation during the first S‐phase, and completely abolished the second peak of DNA synthesis. The fraction of cells irreversibly committed to mitosis increased when the time after stimulation increased. Less than three hours' incubation with insulin or trypsin was needed to initiate proliferation of a significant fraction of the cell population. It is concluded that reversion of the initiated cycle of a given cell is no more possible after the cell has entered the S‐phase.