Proteolytic degradation of insulin-like growth factor (IGF)-binding protein-3 by porcine ovarian granulosa cells in culture: regulation by IGF-I

Abstract

Porcine ovarian granulosa cells in culture secrete glycosylated insulin-like growth factor (IGF)-binding protein-3 (IGFBP-3), which inhibits gonadotropin and IGF action in the ovary. Synthesis of IGFBP-3 is stimulated by IGF-I and attenuated by gonadotropin. The purpose of the present study was to determine whether IGFBP-3 levels were also regulated via proteolysis. Exogenously added nonglycosylated recombinant human IGFBP-3 (rhIGFBP-3) was significantly degraded over time by a soluble serine-specific protease, similar to plasmin, in control cultures and those treated with FSH, insulin, or several other classes of hormones. In contrast, degradation was greatly attenuated by the IGFs. Degraded rhIGFBP-3 exhibited much reduced affinity for [125I]IGF-II, suggesting that degradation could make available IGFs for cellular interaction. The mechanism of IGFBP-3 protease inhibition by IGFs is unclear. Mediation by IGF receptors is unlikely, as insulin at a dose that activated both insulin and type I IGF receptors did not alter intrinsic degradation of IGFBP-3 (as does IGF). Additionally, IGF-I attenuation of IGFBP-3 degradation was not inhibited by antagonism of receptor action with a tyrosine kinase inhibitor. Further, IGF-I inhibited degradation in cell-free conditioned medium. Direct stabilization of IGFBP-3 via binding of IGFs was suggested from these results. However, long R3 IGF-I attenuated IGFBP-3 degradation even though it has low affinity for IGFBPs. Inhibition of the protease by IGFs is also possible. We conclude that IGFs inhibit the degradation of exogenous nonglycosylated rhIGFBP-3. If active in vivo, this may serve to increase endogenous IGFBP-3 levels in follicular fluid.

Profiles of Insulin-Like Growth Factor Binding Proteins and the Protease Activity in the Maternal Circulation and Its Local Regulation Between Placenta and Decidua.

“…Recently it was reported that IGFBPs are cleaved by specific proteases [12,13]. Once IGFBPs are cleaved by proteases, their binding affinity for IGF decreases [14], resulting in a change in its effect on IGF [15,16].…”

ADAM 12, a Disintegrin Metalloprotease, Interacts with Insulin-like Growth Factor-binding Protein-3

“…This IGFBP-3 protease activity disappears rapidly after parturition and is thought to arise from the placenta (5,8). IGFBP-3 protease activity has also been identified in a variety of other situations such as in conditioned medium from MCF-7 breast cancer cells (9), skin blister fluid (10), and in ovarian follicular fluid (11). The relationship between each of these protease activities is not clear at this point in time.…”

Proteolysis of Insulin-Like Growth Factor-Binding Protein-3 by Human Skin Keratinocytes in Culture in Comparison to that in Skin Interstitial Fluid: The Role and Regulation of Components of the Plasmin System1

“…IGF-I has been shown to inhibit the plasmin-induced and plasmin-like protease-induced IGFBP-3 proteolysis in human osteosarcoma cells and porcine ovarian granulosa cells, respectively (13,14). In the former, the inhibition was believed to be mediated by depressing the activity of uPA (13).…”