Mechanism of regulation of fibroblastic cell replication. IV. An analysis of the serum dependence of cell replication based on Michaelis-Menten kinetics.

Abstract

AbstractThe quantitative dependence of the initiation of DNA synthesis in nonreplicating cultures of human diploid fibroblasts (HDFs) on the concentration of fresh serum present in the stimulating medium has been analyzed. A simple formal analysis of the system suggested that the theory of dissociation reactions should lead to a model analogous to the Michaelis‐Menten formulation. Accumulation of enough data for statistical analysis became possible by innovations in the culture techniques and isotope incorporation methods. The data were shown to conform to this model, and parameters Vm and K′m were calculated, together with appropriate statistical functions, from a weighted linear regression of the double reciprocal transformation of the data. Several variables in the culture system were assessed for their effects on the kinetic parameters. The results were consistent with the hypothesis that Vm is a measure of the maximum population of cells capable of responding to the stimulation, and K′m is an estimate of the interaction between the mitogenic factors in serum and the cell surface.Different strains of HDFs had different parameter values. Of several culture variables tested, the Vm was found to be influenced by the type of serum used, and by overall cell density as well as density variations within the monolayer. K′m showed significant changes only in response to cell density, and the evidence suggests that the affinity of cells for serum factors decreases as the cell density increases in confluence.