The 25-kilodalton insulin-like growth factor (IGF)-binding protein inhibits both basal and IGF-I-mediated growth of chick embryo pelvic cartilage in vitro.
Insulin-like growth factor (IGF)-I stimulates the growth of many tissues, including growth plate cartilage. However, the role of IGF-binding proteins in the growth process is controversial. We purified a 25-kDa IGF-binding protein (BP-25) from amniotic fluid. We tested the effect of this BP-25 preparation on both basal and IGF-I-stimulated growth of chick embryo pelvic cartilages maintained in serum-free organ culture. Cartilage wet weight was 4.1 +/- 0.3 mg/cartilage initially; after 3 days, BP-25 inhibited both basal and IGF-I-stimulated growth. Control cartilages weighed 7.4 +/- 0.7 mg/cartilage, while those incubated with 100 nM BP-25 weighed 5.8 +/- 0.5 mg/cartilage (P less than 0.001 vs. control); BP-25 concentrations as low as 0.2 nM significantly inhibited basal cartilage growth. Cartilages incubated with 1.25 nM IGF-I weighed 10.4 +/- 0.8 mg/cartilage (P less than 0.001 vs. control), while those incubated with both 100 nM BP-25 and 1.25 nM IGF-I weighed 8.1 +/- 0.5 mg/cartilage (P less than 0.001 vs. cartilage incubated with IGF-I alone); BP-25 concentrations as low as 0.4 nM significantly inhibited IGF-I-stimulated cartilage growth. BP-25 also inhibited basal and IGF-I-stimulated increases in cartilage dry weight, [3H]thymidine incorporation into DNA, and 35SO4 incorporation into proteoglycan. A second BP-25 preparation, which in the presence of 1% platelet-poor plasma acts synergistically with IGF-I to stimulate DNA synthesis and cell replication of fibroblasts and smooth muscle cells in tissue culture, inhibited IGF-I-stimulated cartilage growth to the same degree as did our BP-25 preparation. In separate experiments, proteins present in serum-free medium conditioned for 3 days by chick cartilages were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, transferred to nitrocellulose, and incubated with [125I]IGF-I. This medium was found to contain two IGF-binding proteins; one appeared to be the chick equivalent of BP-25, while the other had a molecular mass similar to that of a poorly characterized human 34-kDa IGF-binding protein. We conclude that purified BP-25 inhibits the growth of chick embryo pelvic cartilage in our serum-free organ culture system. Since conditioned medium from these cartilages contains both IGF-I-like peptides and IGF-binding proteins such as BP-25, we suggest that the IGF-binding proteins present may act to down-regulate the growth-promoting effects of the local IGF peptides.
Department of Medicine, Duke University Medical Center, Durham, North Carolina 27710.
W M
WM
Burch
J
J
Correa
J E
JE
Shively
D R
DR
Powell
D000818:Animals; D002352:Carrier Proteins / Q000494:pharmacology*; D002356:Cartilage / Q000187:drug effects / Q000254:growth & development*; D002453:Cell Cycle / Q000187:drug effects; D002642:Chick Embryo; D004247:DNA / Q000096:biosynthesis; D004305:Dose-Response Relationship, Drug; D015536:Down-Regulation; D018969:Insulin-Like Growth Factor Binding Proteins; D007334:Insulin-Like Growth Factor I / Q000037:antagonists & inhibitors / Q000494:pharmacology*; D009924:Organ Culture Techniques; D009929:Organ Size / Q000187:drug effects; D010384:Pelvic Bones / Q000187:drug effects / Q000254:growth & development*; D010455:Peptides / Q000494:pharmacology; D013002:Somatomedins / Q000494:pharmacology*; D013936:Thymidine / Q000378:metabolism
D002352:Carrier Proteins; D018969:Insulin-Like Growth Factor Binding Proteins; D010455:Peptides; D013002:Somatomedins; D007334:Insulin-Like Growth Factor I; D004247:DNA; D013936:Thymidine