Regulation of immunoreactive insulin-like growth factor binding protein-6 in normal and transformed human fibroblasts.
Insulin-like growth factor-binding proteins (IGFBPs) have been shown to both potentiate and inhibit IGF bioactivity in vitro; thus, changes to the type or amount of IGFBPs present in the cellular environment will ultimately affect insulin-like growth factor action. In this study, we have investigated the production of immunoreactive IGFBP-6 by normal human fibroblasts (NHF) and an SV-40-transformed human fibroblast line (AG2804). When analyzed by SDS-polyacrylamide gel electrophoresis and immunoblotting, IGFBP-6 appeared as a doublet of 32-34-kDa in conditioned medium of both cell lines, with the lower molecular mass band predominating in the NHF cell line. Measured by a specific radioimmunoassay, serum-free NHF, and AG2804 cultures secreted IGFBP-6 at 1.44 +/- 0.09 and 1.23 +/- 0.08 ng/10(4) cells (mean +/- S.E.), respectively. Despite a relatively weak IGFBP-6 signal by ligand blot compared with IGFBP-3, the two proteins were secreted in similar molar concentrations by NHF. Retinoic acid increased IGFBP-6 by 3-fold in NHF and AG2804-conditioned media, maximal at approximately 100 nM retinoic acid. In contrast, IGFBP-6 production was inhibited by transforming growth factor-beta 1 and agents that increase intracellular cAMP concentrations, including dibutyryl cAMP, forskolin, isobutylmethylxanthine, and cholera toxin. This study indicates that IGFBP-6 has a pattern of regulation unique among the IGFBPs, supporting the concept of specific roles for each binding protein in regulating cell growth and metabolism.
Martin
J L
JL
Department of Medicine, University of Sydney, NSW, Australia.
Coverley
J A
JA
Baxter
R C
RC
D015056:1-Methyl-3-isobutylxanthine / Q000494:pharmacology; D003994:Bucladesine / Q000494:pharmacology; D002352:Carrier Proteins / Q000096:biosynthesis / Q000302:isolation & purification / Q000378:metabolism*; D002460:Cell Line; D002461:Cell Line, Transformed; D002471:Cell Transformation, Neoplastic; D002772:Cholera Toxin / Q000494:pharmacology; D005576:Colforsin / Q000494:pharmacology; D017077:Culture Media, Conditioned; D003513:Cycloheximide / Q000494:pharmacology; D004591:Electrophoresis, Polyacrylamide Gel; D005347:Fibroblasts / Q000187:drug effects / Q000378:metabolism; D006801:Humans; D018976:Insulin-Like Growth Factor Binding Protein 6; D007335:Insulin-Like Growth Factor II / Q000302:isolation & purification / Q000378:metabolism*; D007700:Kinetics; D008297:Male; D008970:Molecular Weight; D011994:Recombinant Proteins / Q000302:isolation & purification / Q000378:metabolism; D013539:Simian virus 40 / Q000235:genetics*; D012867:Skin / Q000378:metabolism; D016212:Transforming Growth Factor beta / Q000494:pharmacology; D014212:Tretinoin / Q000494:pharmacology; D010566:Virulence Factors, Bordetella / Q000494:pharmacology
D002352:Carrier Proteins; D017077:Culture Media, Conditioned; D018976:Insulin-Like Growth Factor Binding Protein 6; D011994:Recombinant Proteins; D016212:Transforming Growth Factor beta; D010566:Virulence Factors, Bordetella; D005576:Colforsin; D014212:Tretinoin; D003994:Bucladesine; D007335:Insulin-Like Growth Factor II; D002772:Cholera Toxin; D003513:Cycloheximide; D015056:1-Methyl-3-isobutylxanthine