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Angiogenesis in the developing corpus luteum (CL) is a prerequisite for establishment and maintenance of an early pregnancy. To explore the physiological significance of insulin-like growth factor-binding protein-7 (IGFBP7) in the developing CL, the effects of IGFBP7 on vascular endothelial growth factor (VEGFA)- and luteinizing hormone (LH)-induced in vitro tube formation were tested using isolated luteal microvascular endothelial cells (LECs). Capillary-like tube formation of LECs and their proliferation were stimulated by both VEGFA and LH. IGFBP7 treatment suppressed VEGFA- or LH-induced tube formation. The proliferation and migration of LECs, and phosphorylation of mitogen-activated protein kinase kinase and extracellular signal-regulated kinase 1/2 were inhibited by IGFBP7. Furthermore, IGFBP7 attenuated VEGFA-enhanced cyclooxygenase (COX)-2 mRNA expression and prostaglandin E2 secretion. These findings suggest the possibility that luteal IGFBP7 secretion may suppress the stimulatory effect of VEGFA on angiogenesis in the early CL.

Uterine endometrial glands and their secretory products are critical for the implantation and survival of the peri-implantation embryo, and for the establishment of uterine receptivity. We previously reported that insulin-like growth factor binding protein 7 (IGFBP7) is abundantly expressed in uterine glandular epithelial cells during the secretory phase of the menstrual cycle. In the present study, we used a cultured glandular epithelial cell line of human (EM1) to investigate the significance of IGFBP7 in the function of endometrial glands. EM1 cells formed a mesh-like structure on Matrigel, which was accompanied by elevated levels of intracellular cyclic AMP. However, these morphological changes were blocked by treatment with protein kinase A (PKA) inhibitor (H89). IGFBP7 knockdown using specific short interference RNA (siRNA) inhibited the formation of the mesh-like structure on Matrigel. Cyclic AMP analogs, dibutyryl-cAMP, and N(6)-phenyl-cAMP induced the expression of leukemia inhibitory factor (LIF) which is essential for the onset of implantation. Enhanced LIF expression was suppressed by IGFBP7 siRNA treatment. Western blot analysis revealed that IGFBP7 knockdown results in the aberrant, constitutive expression of the MAPK signaling pathway. These results suggest that IGFBP7 regulates morphological changes of glandular cells by interfering with the normal PKA and MAPK signaling pathways that are associated with the transformation and/or differentiation of endometrial glands.

Insulin-like growth factor binding protein-7 (IGFBP7) and vascular endothelial growth factor (VEGF) are expressed in vascular endothelial cells in several tumor types. In this study, we examined the effect of IGFBP7 on VEGF-induced tube formation in cultured human umbilical vein endothelial cells (HUVECs) and its potential action in the modulation of VEGF signaling in vascular cells. IGFBP7 treatment suppressed VEGF-induced tube formation, proliferation, and the phosphorylation of mitogen-activated protein kinase kinase (MEK) and extracellular signal-regulated kinase (ERK) 1/2 in HUVECs. IGFBP7 attenuated VEGF-enhanced cyclooxygenase (COX)-2 and VEGF mRNA expression, and prostaglandin E(2) secretion. Knocking down endogenous IGFBP7 enhanced COX-2 and VEGF mRNA expression. A significant increase in IGFBP7-induced caspases was not observed in the presence of VEGF. These findings indicate that IGFBP7 can modulate the stimulatory effect of VEGF on angiogenesis by interfering with VEGF expression as well as VEGF signaling and not by inducing apoptosis.

Background: Insulin-like growth factor-binding protein-7 (IGFBP7) is a secretory protein with a molecular mass of approximately 30 kDa. It is abundantly expressed in the uterine endometrium during the secretory phase of the menstrual cycle. Decreased IGFBP7 expression has been observed in some cancers and leiomyomata., Methods: To determine whether serum IGFBP7 levels reflect changes in uterine IGFBP7 expression in humans during the menstrual cycle, and to examine whether serum IGFBP7 levels are altered in patients with various disorders, we developed a novel, dual-antibody sandwich enzyme-linked immunosorbent assay (ELISA). Firstly, concentrations of IGFBP7 released into the medium were determined in cultured endometrial stromal and glandular cells. Blood samples were collected from women who had normal menstrual cycles and who had been diagnosed with endometriosis. Serum from hemodialysis patients and gastrointestinal cancers was also used to determine the IGFBP7 levels., Results: Using this new ELISA, we demonstrated that cultured uterine cells secrete IGFBP7 into the medium. Patients with endometriosis and those with type II diabetes mellitus undergoing hemodialysis had significantly higher serum concentrations of IGFBP7 than the relevant control subjects. There were no differences in serum IGFBP7 levels in women at different stages of the menstrual cycle. Furthermore, serum IGFBP7 levels in patients with colorectal, esophageal, or endometrial cancer were not different than normal healthy subjects., Conclusion: Our observations suggest that IGFBP7 is associated with the pathophysiology of endometriosis and diabetes mellitus, and that serum IGFBP7 levels do not reflect enhanced uterine expression of IGFBP7 mRNA during the menstrual cycle.

Successful implantation and placentation require that trophoblasts adhere to the uterine epithelium and penetrate the decidualized endometrium. However, the biochemical mechanisms of the establishment of pregnancy including these phenomena have not yet to be definitively elucidated. We have found that stathmin, a cytosolic phosphoprotein that regulates microtubule dynamics, and insulin-like growth factor-binding protein (IGFBP)-related protein 1 (IGFBP-rP1, now called IGF-binding protein 7) were highly expressed in the endometrium around the time of implantation and decidualization. In this article, we review our recent findings of the research regarding the functions of these implantation-associated proteins in endocrine physiology and pharmacology. Analysis of the expression of both factors in rodent and human uterus has revealed that both factors are crucial for the process of endometrial stromal cell differentiation.

Uterine decidualization is crucial for successful implantation and the establishment of pregnancy. In the present study, the expression of insulin-like growth factor-binding protein (IGFBP)-related protein 1 (IGFBP-rP1) in the human uterus and endometrial stromal cells (ESCs) and its physiological significance in decidualization were examined. IGFBP-rP1 protein was localized in the glandular epithelium and stromal cells, and blood vessels in the endometrium. Cultured stromal cells expressed IGFBP-rP1 and secreted it into the medium. IGFBP-rP1 was localized mostly in the cytoplasm near the nucleus. Knocking down the endogenous IGFBP-rP1 expression in stromal cells, by a small interfering (si)RNA, diminished the expression of prolactin and IGFBP-1 which serve as decidual markers. These results suggest that IGFBP-rP1 may play a role in decidualization of ESCs.

Insulin-like growth factor (IGF)-binding protein (IGFBP) 7 is a secreted protein that regulates cellular proliferation, adhesion, and angiogenesis, and has low affinity for IGF compared with that of IGFBP1-IGFBP6. We sought to determine whether IGFBP7 is present in follicular fluid and to elucidate whether IGFBP7 participates in the steroidogenesis of rat mature follicles. Follicular fluid and granulosa cells (GCs) were collected from immature rats 2 days after their treatment with equine chorionic gonadotropin (eCG). IGFBP7 protein was detected in the follicular fluid and the conditioned medium of cultured ovarian GCs by immunoblot analysis. When subconfluent GCs were cultured and treated with FSH and activin, coincubation with FSH and activin markedly increased GC expression of Cyp19a1 (aromatase) mRNA and 17beta-estradiol (E(2)) secretion. The addition of recombinant murine IGFBP7 to these cultures decreased in the activin-enhanced, FSH-stimulated Cyp19a1 mRNA levels in the cells and suppressed the 17beta-E(2) levels in the culture medium. Treatment of GCs with Igfbp7-specific small interfering RNA (siRNA), which knocked down Igfbp7 expression, increased the FSH-stimulated levels of Cyp19a1 but not Cyp11a1 expression. Basal and FSH-stimulated 17beta-E(2) secretion into the culture medium was also enhanced by Igfbp7 siRNA. These results suggest that IGFBP7 suppresses estrogen production in GCs. These observations support the notion that this protein, which is secreted into the follicular fluid, may serve as an intraovarian factor that negatively regulates GC differentiation.

IGF binding protein-related protein 1 (IGFBP-rP1) is highly expressed in the rat uterus around the time of implantation. In the present study, we determined the periimplantation localization of IGFBP-rP1 mRNA and assessed the effects of recombinant IGFBP-rP1 on the proliferative and prostacyclin (PGI(2))-producing abilities of cultured endometrial cells early in pregnancy. IGFBP-rP1 mRNA was detected at high levels in endometrial stromal cells close to the smooth muscle of interimplantation sites around the time of implantation but absent from decidual zones surrounding the embryo. Differential uterine IGFBP-rP1 expression was also recognized in the delayed implanting pregnant model, but the level of mRNA decreased as decidual tissues formed in the decidualization model. Recombinant IGFBP-rP1 inhibited the proliferation of endometrial stromal cells in vitro and arrested them in the G(1) phase of the cell cycle. Furthermore, IGFBP-rP1 significantly stimulated PGI(2) synthesis and cyclooxygenase II mRNA expression in myometrial cells, both of which are essential molecules for successful implantation. These data suggest that IGFBP-rP1 is an implantation-associated protein and that it modulates the proliferation of rat uterine cells and their production of PGI(2) during the periimplantation period.