Human placental trophoblasts secrete a disintegrin metalloproteinase very similar to the insulin-like growth factor binding protein-3 protease in human pregnancy serum

During the course of human pregnancy, there is a marked increase in insulin -like growth factor (IGF) binding protein (IGFBP)-3 protease activity in ma ternal serum that is first evident at 6 weeks of gestation, persists throug h term, and returns to nonpregnancy levels by day 5 postpartum This proteas e activity cleaves IGFBP-3 into smaller fragments that have markedly reduce d affinity fur the IGFs. To date, the precise identity and cellular origin of the pregnancy-associated serum IGFBP-3 protease have not been establishe d. To investigate whether placental and/or decidual tissues, which uniquely develop during pregnancy, may be sources of the pregnancy-associated serum IGFBP protease, we examined the secretion of IGFBP-3 protease in vitro by isolated human cytotrophoblasts or fibroblasts from second trimester placen tae and by in vitro decidualized human endometrial stromal cells. Cytotroph oblasts were either cultured alone, which favors aggregation and fusion, or cocultured with decidualized endometrial stromal cells, which favors diffe rentiation to an invasive phenotype. IGFBP-3 protease activity was detected in trophoblast, but not in placental fibroblast or decidualized endometria l cultures, and was also present in trophoblast-endometrial cocultures. Wes tern ligand blot and Western immunoblot analyses showed that most of the en dogenous IGFBP-3 in trophoblast cultures was in the form of low molecular w eight fragments with reduced IGF binding affinity. The substrate specificit y of the trophoblast-derived protease was identical to that in pregnancy se rum, showing activity against IGFBP-2, -3, and -4, but being inactive again st IGFBF-1. IGFBP-3 protcolysis by both pregnancy serum and trophoblast con ditioned medium showed a major peak of activity at neutral pH. The trophobl ast-derived activity caused time and temperature-dependent proteolysis of I GFBP-3 into fragments of identical size as those produced by pregnancy seru m, and also shared its sensitivity to protease inhibitors: highly sensitive to EDTA and o-phenanthroline, partially sensitive to the serine protease i nhibitors AEBSF and aprotinin, and insensitive to alpha(2)-antiplasmin, and to aspartic and cysteine protease inhibitors. IGFBP-3 proteolysis by both pregnancy serum and trophoblast conditioned medium was also insensitive to tissue inhibitor of metalloproteinase-1, precluding the involvement of the matrix metalloproteinases. In contrast, both the pregnancy serum- and troph oblast-derived proteases were preferentially inhibited by a hydroxamic acid derivative with selective activity against the disintegrin-metalloproteina se tumor necrosis factor-alpha converting enzyme. This study shows that pla cental trophoblasts produce an IGFBP-3 protease with characteristics very s imilar to the activity found in pregnancy serum and indicates these cells a t the maternal-fetal inter-face are a potential source of the pregnancy-ass ociated serum IGFBP-3 protease. The findings further suggest that the main IGFBP-3 protease activity in both pregnancy serum and trophoblast condition ed medium may correspond to a disintegrin-metalloproteinase type enzyme.