Growth-promoting Effect, Biological Activity, and Binding of Insulin in Human Intestinal Cancer Cells in Culture¹

The biological action and binding of insulin were tested in two human intestinal cancer cell lines originating from the duodenum (HUTU 80) and the colon (HT 29).

After serum deprivation for 24 hr, insulin stimulated cell division and the incorporation of labeled precursors into RNA, protein, and DNA for both cell lines. The action on the RNA and protein was rapid and significantly different (1.5 to 2 times that of control) 1 hr after adding insulin. These effects were dose dependent, present at physiological concentration in vivo (10^-10 m), and independent of the transport of precursors. For thymidine incorporation, the stimulation was delayed up to 8 hr and culminated with cell division 20 hr later.

As previously shown for HT 29, HUTU 80 cells exhibited insulin-specific binding sites. Binding of ¹²⁵I-insulin was saturable; reversible; and time, temperature, and pH dependent. Scatchard analysis of the binding data of the two cell lines gave curvilinear plots. Assuming the presence of two independent binding sites, the high-affinity constants were 6 to 8 × 10⁸ m^-1, and the number of high-affinity receptors was similar and accounted for 2000 to 3000 receptors/cell.

For both cell lines, the effect of insulin on protein and RNA synthesis was significantly different from control at 1 hr when binding reached a maximum at 37°. The biological action of insulin on growth and macromolecular synthesis was dose dependent and maximum at about 10^-8 m insulin, which corresponds to 70% displacement of ¹²⁵I-insulin binding. Furthermore, the binding and the biological action of proinsulin were about 2% that of native insulin in the two cell lines studied.

These results show that insulin acts as a growth factor for these two cell lines and that these effects are probably mediated by the interaction of insulin with specific receptors.