ACTH and MSH Production by a Single Cloned Mouse Pituitary Tumor Cell Line

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Abstract
The production of ACTH and MSH was studied in the cloned, functional, murine pituitary tumor cell line, AtT–20. Extracts of normal whole LAFj mouse pituitaries, tumors induced by injecting AtT–20 cells into recipient LAFX mice, cultured AtT–20 cells, and the medium in which the cells were cultured were subjected to molecular sieve chromatography on Sephadex G—SO fine resin. The original extracts and the Sephadex fractions were assayed for bioactive ACTH and MSH. Each of the extracts contained both ACTH and MSH. The ACTH existed as two distinct moieties, one of which eluted earlier than the ACTH standard. The MSH eluted in later fractions and thus was a separate molecular entity from the ACTH's. The extracts and fractions were subjected to radioimmunoassays for ACTH, a—MSH, and P—MSH. Both ACTH moieties were immunologically indistinguishable from highly purified human ACTH standard. The MSH in normal mouse pituitary and AtT–20 tumor was immunologically indistinguishable from α—MSH standard, but less than one—fourth of the MSH biologic activity could be accounted for by immunoreactive α—MSH. The MSH in the cultured AtT—20 cells and culture medium, which eluted earlier from the Sephadex column, failed to react in the α—MSH radioimmunoassay. The MSH in each extract reacted with antibodies in the β—MSH radioimmunoassay, generating competition curves parallel to each other but less steep than β—MSH standard. Thus, the MSH was immunologically similar, but not identical, to any known β—MSH. The normal mouse pituitary and the AtT–20 tumor cell synthesize two ACTH's, the larger of which may be a precursor molecule, and a smaller MSH molecule which has immunochemical similarities to both α—MSH and β—MSH. (Endocrinology92: 385, 1973)

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