Targeted
tumorigenesis, using the
POMC gene promoter ligated to the simian virus 40
large T antigen, generated transgenic mice with massive
tumors of the intermediate lobe (IL) of the pituitary. Inoculation of nude mice with the IL
tumor cells resulted in very large secondary
tumors. As the IL from several species produces a potent PRL-releasing factor (PRF), it was of interest to determine whether IL
tumors from these mice also contain PRF. The objectives were to 1) measure serum PRL levels in mice with IL
tumors, 2) determine whether these
tumors contain PRF and examine its chromatographic properties, and 3) analyze whether this PRF is related to
POMC, its derivatives, or other PRL
secretagogues. Serum PRL levels were 5- to 6-fold higher in transgenic than in control mice. Primary and secondary IL
tumors were
acid extracted and successively fractionated using
Sephadex G-100 gel filtration and reverse phase and gel permeation HPLC. PRF activity was determined using short term incubation of
tissue extracts or column fractions with GH3 cells. Crude
tumor extracts exhibited a strong and dose-dependent PRF activity. Upon chromatography, the PRF activity from either primary or secondary
tumors resolved into two classes of compounds: a big PRF with an estimated mol wt of 70-80 kilodaltons and two small, very hydrophobic
peptides. The elution profiles of the three PRFs differed from those of
beta-endorphin,
alpha MSH,
beta MSH,
ACTH, TRH,
oxytocin,
angiotensin II,
vasoactive intestinal polypeptide, or
corticotropin-like intermediate
peptide. In summary, we have identified an animal model with IL
tumors that has
hyperprolactinemia and overproduces PRF. Two classes of PRFs, big and small, were resolved which differ from
POMC derivatives and known regulators of PRL release. These data suggest that PRF is produced by melanotrophs, but is not a product of the
POMC gene. The IL
tumors should provide an excellent source for the purification and structural elucidation of PRFs.