Characterization of the receptor for insulin-like growth factor II in bone cells

We have previously shown that insulin-like growth factor II (IGF-II) is produced by bone cells and that IGF-II stimulates cell proliferation and collagen synthesis in bone cells. We now extend these in vitro findings by demonstrating specific IGF-II binding to bone cells derived from newborn mouse calvaria and embryonic chick calvaria. The kinetics of 125I] IGF-II binding in embryonic chick calvaria cells showed time and temperature dependence. Scatchard analysis of 125I]IGF-II binding to chick calvaria cells showed an apparent Kd of 1.4 x 10(-10) M, with a calculated receptor site concentration of 40,000/cell. The specificity characteristics showed that IGF-II was significantly more potent than IGF-I or insulin in displacing IGF-II tracer. Competition for binding of 125I]IGF-II by unlabeled IGF-II showed a dose-dependent displacement between 0.5 and 25 ng/ml. Fifty percent displacement of 125I]IGF-II binding to chick and mouse calvarial cells was achieved at 1-2 ng/ml; 90% of specific binding of 125I]IGF-II was displaceable in the presence of 125 ng/ml of unlabeled IGF-II. IGF-I showed less than 5% cross reactivity for displacement of 125I]IGF-II binding to chick and mouse bone cells. Type II receptor inhibitory antibodies, R-II-PAB1 inhibited the binding of 125I]IGF-II to mouse bone cells and H-35 rat hepatoma cells (which contain type II but not type I receptors) in a dose-dependent manner. R-II-PAB1 also inhibited basal cell proliferation as well as IGF-II-, IGF-I-, and fibroblast growth factor (FGF)-induced cell proliferation in mouse bone cells. In chick calvaria bone cells and TE89 human osteosarcoma cells, R-II-PABI inhibited neither binding of 125I]IGF-II nor IGF-II-induced cell proliferation. These results together with our findings that IGF-II increased chick bone cell proliferation in the presence of maximal doses of IGF-I suggest that at least part of the mitogenic action of IGF-II is mediated through type II rather than type I receptors in bone cells.