Regulation of insulin-like growth factor binding protein 4 by a specific insulin-like growth factor binding protein 4 proteinase in normal human osteoblast-like cells: implications in bone cell physiology.
Insulin-like growth factor binding protein 4 (IGFBP-4) is secreted by normal human osteoblast-like cells (hOB) and is a potent inhibitor of insulin-like growth factor (IGF) action in vitro. In previous studies, IGF treatment of hOB in culture led to markedly reduced medium levels of IGFBP-4 as detected by western ligand blotting. In the present study, incubation of hOB-conditioned medium (hOB-CM) with IGF under cell-free conditions resulted in a similar loss of IGFBP-4. Both IGF-I and IGF-II were capable of inducing a decrease in IGFBP-4; however, IGF-II was more effective. When the six characterized IGFBP were added to hOB-CM, only IGFBP-4 disappeared in response to IGF-II addition. This IGF-regulated loss of IGFBP-4 was inhibited by metalloproteinase inhibitors and appeared to be due to a proteinase that cleaved IGFBP-4 in 18 and 14 kD fragments identified by western immunoblotting. Conditioned media from eight of eight different donor hOB lines tested exhibited IGFBP-4 proteinase activity. To assess the biologic consequences of IGF-II-induced IGFBP-4 proteolysis, we treated hOB with IGF-II for 5 h, which decreased medium IGF-BP-4 by 70%, and then measured IGF-I and insulin stimulation of [3H]thymidine incorporation. IGF-II itself was not mitogenic and had no effect on insulin-stimulated [3H]thymidine incorporation. However, pretreatment of cultured hOB with IGF-II enhanced IGF-I-stimulated [3H]thymidine incorporation threefold.(ABSTRACT TRUNCATED AT 250 WORDS)
Durham
S K
SK
Endocrine Research Unit, Mayo Clinic, Rochester, Minnesota.
Kiefer
M C
MC
Riggs
B L
BL
Conover
C A
CA
D000328:Adult; D015153:Blotting, Western; D002352:Carrier Proteins / Q000378:metabolism*; D002478:Cells, Cultured; D017077:Culture Media, Conditioned; D004591:Electrophoresis, Polyacrylamide Gel; D005260:Female; D013006:Growth Hormone / Q000494:pharmacology; D006801:Humans; D007328:Insulin / Q000494:pharmacology; D018974:Insulin-Like Growth Factor Binding Protein 4; D007334:Insulin-Like Growth Factor I / Q000494:pharmacology; D007335:Insulin-Like Growth Factor II / Q000494:pharmacology; D008024:Ligands; D008297:Male; D008666:Metalloendopeptidases / Q000378:metabolism*; D010006:Osteoblasts / Q000187:drug effects / Q000378:metabolism*; D011266:Pregnancy-Associated Plasma Protein-A; D011994:Recombinant Proteins / Q000494:pharmacology; D013002:Somatomedins / Q000378:metabolism
D002352:Carrier Proteins; D017077:Culture Media, Conditioned; D007328:Insulin; D018974:Insulin-Like Growth Factor Binding Protein 4; D008024:Ligands; D011994:Recombinant Proteins; D013002:Somatomedins; D007334:Insulin-Like Growth Factor I; D007335:Insulin-Like Growth Factor II; D013006:Growth Hormone; D008666:Metalloendopeptidases; D011266:Pregnancy-Associated Plasma Protein-A