Characterization of neurotensin production by a line of rat medullary thyroid carcinoma cells.

Abstract

The rMTC 6-23 cell line was derived from a calcitonin-producing rat medullary thyroid carcinoma. These cells also synthesize and secrete neurotensin (NT), a tridecapeptide originally isolated from the hypothalamus and not previously associated with C [calcitonin] cells. Immunoreactive NT (iNT) was measured with 2 antisera: HC-8, which recognizes the COOH-terminal of 8 amino acids, and TG-1, which binds the NH2-terminal region of NT. By use of these antisera, 2 M acetic acid extracts of rMTC 6-23 cells were found to contain 0.7-5.0 pmol of iNT/mg of cell protein. Successive fractionation of iNT from cell extracts by gel filtration, ion-exchange chromatography and reverse-phase high-pressure liquid chromatography showed at each step a peak of iNT that was indistinguishable from synthetic NT in its chromatographic behavior. Biologic activity of the iNT was confirmed by demonstrating the characteristic fall in arterial blood pressure in the rat after i.v. injection of material purified from rMTC 6-23 cells. Calcium (0.5-4.0 mM) stimulated release of iNT in a dose-dependent manner; the effect was maximal at 3-4 mM Ca. K+ (50 mM) stimulated release of iNT that was maximal in the presence of 1.0-1.5 mM Ca. Synthesis and secretion of iNT by these cells were shown during a 16-day growth experiment. There may be an association between neurotensin and calcitonin.