Human hepatoma cells synthesize and secrete insulin-like growth factor Ia prohormone under growth hormone control.
Nucleotide sequencing of cDNAs encoding human insulin-like growth factor I (IGF-I) predicts the existence of two different prohormone forms of IGF-I. The E peptide regions extend the carboxy-terminus of the 70 amino acid core IGF-I molecule (BCAD domains) by either an additional 35 (IGF-Ia) or 77 (IGF-Ib) amino acids. Employing antiserum directed against a peptide sequence unique to the E peptide region of IGF-Ia prohormone, we have identified EIa immunoreactive material (IR-EIa) in the conditioned medium of a human hepatoma cell line, HepG2. Human growth hormone (GH) had dose-dependent stimulatory effects on IR-EIa secretion; incubation of HepG2 cells with GH at maximal concentrations (1-5 micrograms/ml) increased specific IR-EIa in 24 h conditioned medium 3-fold. The addition of human placental lactogen, insulin, IGF-I, dexamethasone, beta-estradiol, or progesterone had no significant effect. Acid chromatography of HepG2 cell conditioned medium revealed a single elution peak of IR-EIa corresponding to M(r) = 12,000-20,000. There was no immunologically detectable 7500 M(r) IGF-I peptide in acid-chromatographed conditioned medium under either basal or stimulated conditions. Biosynthetic labelling of HepG2 cell products with [35S]Trans label and immunoprecipitation with antisera specific to the E or to the AD regions of the IGF-Ia molecule indicated a single species of approx. 14,000 M(r). These data indicate that the E peptide region of IGF-Ia is translated and released as part of the larger molecule in cultured HepG2 cells, and that the levels of this prohormone are regulated by GH.
Conover
C A
CA
Endocrine Research Unit, Mayo Clinic, Rochester, MN 55905.
Baker
B K
BK
Bale
L K
LK
Clarkson
J T
JT
Liu
F
F
Hintz
R L
RL
D006528:Carcinoma, Hepatocellular / Q000378:metabolism*; D002460:Cell Line; D017077:Culture Media, Conditioned; D003907:Dexamethasone / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004958:Estradiol / Q000494:pharmacology; D013006:Growth Hormone / Q000494:pharmacology*; D007328:Insulin / Q000494:pharmacology; D007334:Insulin-Like Growth Factor I / Q000096:biosynthesis* / Q000302:isolation & purification / Q000494:pharmacology; D008113:Liver Neoplasms / Q000378:metabolism*; D008970:Molecular Weight; D010446:Peptide Fragments / Q000096:biosynthesis* / Q000302:isolation & purification; D010928:Placental Lactogen / Q000494:pharmacology; D011374:Progesterone / Q000494:pharmacology; D011863:Radioimmunoassay; D011994:Recombinant Proteins / Q000494:pharmacology; D014407:Tumor Cells, Cultured
D017077:Culture Media, Conditioned; D007328:Insulin; D010446:Peptide Fragments; D011994:Recombinant Proteins; C053766:insulin-like growth factor 1A prohormone (91-103); D011374:Progesterone; D004958:Estradiol; D007334:Insulin-Like Growth Factor I; D003907:Dexamethasone; D013006:Growth Hormone; D010928:Placental Lactogen