In the present study we used 125I-labeled insulin-like growth factor I (125I-IGF-I) to identify and characterize IGF-I receptors in the well-characterized and propagable human esophageal epithelial (HEE) cell line and to characterize their role in cell growth. Binding of 125I-IGF-I was saturable, time and temperature dependent, reversible, and specific for IGF-I and related peptides. Scatchard analysis of binding data demonstrated that HEE cells possess two classes of IGF-I receptors: high affinity [dissociation constant (Kd) = 0.058 nM] and low capacity (13,870 receptors/cell), and low affinity (Kd = 2.2 nM) and high capacity (39,000 receptors/cell). Binding of 125I-IGF-I was inhibited with the following relative potencies (half-maximal inhibition): IGF-I (3.0 pM) > IGF-II (1.2 mM) >> insulin (1.0 microM). Affinity cross-linking of cell membranes using disuccinimidyl suberate as a cross-linking agent under reducing conditions revealed a single polypeptide band (relative mol wt 133,000) representing the alpha-subunit of the IGF-I receptor. IGF-I stimulated [3H]thymidine incorporation and cell proliferation in a dose-dependent manner with detectable effect observed with 0.5 nM IGF-I and maximal effect at 50 nM IGF-I. IGF-I occupation of low-affinity IGF-I receptors appears to mediate cell growth. The present results demonstrate that HEE cells possess two classes of IGF-I receptors: one class has a high affinity and low capacity and the other has a low affinity and high capacity for IGF-I. Occupation of low-affinity IGF-I receptors by IGF-I appears to mediate cell growth.
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