Insulin-Like Growth Factor I Gene Expression iii GH3Rat Pituitary Cells: Messenger Ribonucleic Acid Content, Immunocytochemistry, and Secretion*

Abstract

Insulin-like growth factor I (IGF-I) is present in multiple tissues and cell types. Expression of the IGF-I gene was examined in GH3 cells, a rat pituitary tumor cell line secreting GH and PRL. Increasing concentrations of RNA extracts of GH3 cells yielded a linear increase in hybridization intensity with a 32P-labeled mouse IGF-I cDNA probe. Northern analysis of GH3 cells poly(A) RNA revealed IGF-I mRNA transcripts 1.3, 5.3, and 7.7, kilobases in size. Poly(A) RNA extracts of BALBc/3T3 fibroblasts, a cell line dependent on exogenous somatomedins for DNA synthesis, and of JEG-3 cells, a choriocarcinoma cell line, did not hybridize with the IGF-I cDNA probe. GH3 cells showed positive immunoperoxidase staining using a rabbit anti[Thr59]IGF-I antibody which was largely blocked by prior incubation of the antibody with excess IGF-I. Negligible background peroxidase activity was present in cells incubated with a rabbit nonimmune serum and PBS. Furthermore, BALBc/3T3 fibroblasts showed only weak specific staining with the IGF-I antibody. Finally, GH3 cells secreted IGF-I into the culture medium in a time-dependent fashion, while neither 3T3 nor JEG-3 cells produced detectable medium levels of the peptide after 72 h of incubation. As IGF-I is known to inhibit GH production by the pituitary, the data shown suggest that locally produced IGF-I may regulate GH secretion in an autocrine or paracrine fashion.