Evidence for the presence of insulin binding sites in isolated rat intestinal epithelial cells.
Insulin receptors have been demonstrated in isolated rat intestinal epithelial cells. The specific binding of 125I-insulin was time--and temperature--dependent, the optimal temperature of study being 15 degrees. Dissociation of bound 125I-insulin by an excess of unlabelled hormone was rapid and attained 66 +/- 2% in 2 h. When initiated by dilution, the dissociation attained 35 +/- 4% in 2 h, and 72 +/- 1% in 2 h when 10(-7) mol/l unlabelled insulin was added. The pH optimum for the binding process was between 7.5 and 8, and the binding increased proportionally to cell protein concentration up to 1.5 mg/ml. Under standard conditions (2 h at 15 degrees) the degradation of the labelled hormone in the medium accounted for 20--50% of total tracer, depending on the concentration of cells. At apparent equilibrium (2 h at 15 degrees), unlabelled insulin in the range of 10(-10) to 10(-7) mol/l inhibited competitively the binding of 4.3--7 X 10(-11) mol/l 125I-insulin; fifty per cent inhibition was obtained with 3 X 10(-9) mol/l native insulin. Scatchard analysis, after correction for degradation, gave curvilinear plots, that may be explained by two orders of binding sites, with 2,000 +/- 200 sites/cell of high affinity (Ka = 2.2 +/- 0.2 X 10(9) l/mol) and 39,000 +/- 3,000 sites/cell of low affinity (Ka = 5.6 +/- 1.6 X 10(7) l/mol). The potency of proinsulin to compete with 125I-insulin for the binding site was 3% that of insulin, unrelated peptides were inactive. Such results give a molecular basis to different reports suggesting that the intestine could be a target-tissue for insulin.
D000818:Animals; D001667:Binding, Competitive; D004848:Epithelium / Q000378:metabolism; D005260:Female; D007328:Insulin / Q000378:metabolism*; D007421:Intestine, Small / Q000378:metabolism*; D007700:Kinetics; D008871:Microvilli / Q000378:metabolism; D051381:Rats; D011972:Receptor, Insulin / Q000378:metabolism*
D007328:Insulin; D011972:Receptor, Insulin