AJP - Endocrinology and Metabolism, Vol 243, Issue 2 132-E139, Copyright © 1982 by American Physiological Society
Binding, degradation, and bioactivity of insulin in primary cultures of rat hepatocytes
P. T. Varandani, R. M. Darrow, M. A. Nafz and G. L. Norris
Hepatocytes from fasted rats, previously maintained as a monolayer in a
serum-free glucagon-containing culture medium are demonstrated to provide a
useful model system for the study of receptor-mediated mechanisms of
insulin action. The cultured liver cells show glucagon- and
insulin-responsive biological effects. These cultures show the long-term
effects of insulin on the syntheses of protein ([3H]leucine incorporation),
glycogen, and lipids (conversion of [3H]glucose) in a dose-dependent manner
in the physiological range of insulin concentrations. The order of the
effects of different analogues of insulin on each of the bioactivities
studied at 37 degrees C is the same as their order to compete at 25 degrees
C for binding to insulin-specific receptors. The characterization of
radioactive products of 125I-insulin using gel filtration and anti-insulin
A chain antibody has shown a transient accumulation of insulin A chain,
indicating that the sequential pathway of insulin degradation is operative
in the anchored, cultured hepatocytes. Whereas the synthesis of proteins
proceeds linearly after insulin inoculation, the syntheses of glycogen and
lipids first occur after a lag period of about 10 and 12 h, respectively.
None of the three 125I-labeled fragments or products of 125I-insulin
released in the culture mediums showed any biological activity
(glycogenesis) in cultured hepatocytes although a high-molecular-weight
125I-product isolated from cells could not be tested because of its
insolubility. The possibility that the insulin effects might have been
mediated via a nonradioactive fragment of insulin or another chemical agent
remains open.
