Ruffling and locomotion: role in cell resistance to growth factor-induced proliferation

It has long been known that the growth rate of cells in vitro can be retarded by providing substrates of restricted area. Such experiments were performed with adhesive islets, made by depositing metals onto agarose layers through templates of various sizes. Since normal cells are unable to adhere to agarose, they become confined to the metallic surface. Using such haptotactic islets, we have studied the role of membrane ruffling and cell locomotion in the resistance of AG1523 human fibroblasts to growth factor-induced mitogenesis. Cells plated on small substrates, i.e., 2,150 microns 2 in area, initially showed vigorous ruffling, which was suppressed by 8 h after plating but had resumed again by 12 h. In contrast, cells on larger-size islets showed a rapid decline and stabilization of ruffling activity. When the growth rate was measured for single cells cultured on haptotactic islets, it was found to increase linearly from areas of 4,280 microns 2 up to 425,000 microns 2. Since the area needed to saturate the growth response was approximately 50-fold larger than the area occupied by a single cell, the growth inhibition was attributed in part to an interference with locomotion. The implication that locomotion provided positive input into growth control mechanisms was subjected to a direct test by evaluating the effect of nine polypeptide growth factors on the motility of serum-starved cells. All except TGF-beta 1 stimulated movement. Finally, the mitogenic effect of growth factors was measured by [3H]thymidine incorporation and found to be proportional to motile activities, as quantitatively assayed. We conclude that locomotion suppression is a factor in AG1523 cell resistance to growth factor-induced mitogenesis.