Density‐associated loss of functional receptors for somatomedin‐C/insulinlike growth factor I (SM‐C/IGF‐I) on cultured human fibroblast monolayers

Abstract

The mitogenic activity of somatomedin-C/insulinlike growth factor-I (SM-C/IGF-I) appears to be greatly influenced by cell culture conditions, especially the presence of other growth factors and nutrients in the culture medium. To investigate the effect of cell density on SM-C/IGF-I activity, we have evaluated SM-C/IGF-I binding and stimulation of DNA synthesis and cell replication as a function of cell density in cultured human fibroblast monolayers. At fibroblast concentrations of 2.7 × 10⁵ and 1.48 × 10⁶ cells per 60-mm dish, specific binding of [¹²⁵I]SM-C/IGF-I per 10⁶ cells was 170% higher in sparse than dense monolayers (9.3% vs. 3.4%). Increased binding in sparse monolayers was attributable to approximately twice as many receptors in sparse as in dense cells (31,000 vs. 16,000 sites per cell), as well as to a modest increase in the affinity constant. Similarly, half-maximal stimulation of [methyl-³H]thymidine incorporation was achieved at SM-C/IGF-I concentrations of 2.5 ng/ml in sparse cells but required 20 ng/ml in dense cells. Although this required only 45% occupancy of membrane receptors on sparse cells, and almost 80% occupancy on dense cells, the total number of occupied receptors was similar in both sparse and dense cells (approximately 13,000 receptors/cell for half-maximal stimulation). The presence of increased numbers of “functional receptors” on sparse fibroblasts thus results in enhanced sensitivity to SM-C/IGF-I stimulation of DNA synthesis and cell replication. Progressive decreases in the number of functional receptors, secondary to cell crowding, may contribute to density-dependent inhibition of fibroblast growth.