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In order to study the effects of insulin-like growth factor (IGF-I) and insulin-like growth factor binding protein (IGFBP-1) on human granulosa cell proliferation after in vitro fertilization, cells were obtained after oocyte retrieval and cultured in the presence or absence of graded amounts of recombinant IGF-I, purified IGFBP-1 and [3H]thymidine. Physiological concentrations of IGF-I (2-200 ng/ml) were found to stimulate [3H]thymidine incorporation into the cells in a concentration-dependent manner. Half-maximal stimulation of [3H]thymidine incorporation was obtained with 10 ng/ml exogenous IGF-I, which was chosen for suppression experiments with graded amounts of purified IGFBP-1. Suppression of IGF-stimulated thymidine incorporation was observed when 200 ng/ml or more of IGFBP-1 was added to the culture medium. The same concentration of IGFBP-1 also markedly inhibited binding of [125I]iodotyrosyl IGF-I to the cells. It is concluded that: (i) after a refractory period, granulosa cells from hyperstimulated follicles retained their mitogenic activity; (ii) IGF-I is capable of stimulating DNA amplification in granulosa cells; and (iii) IGFBP-1 inhibits the IGF-I stimulated proliferation in these cells. In view of our previous studies showing that IGFBP-1 is synthesized by the granulosa cells as they luteinize, the present results suggest that IGFBP-1 is one of the endogenous factors locally regulating the growth and differentiation of granulosa cells.

Oral thyroxine produces a rise in serum IGFBP-1 levels without a change in serum IGF-I concentration.

The steroid hormone estrogen profoundly influences growth and differentiation programs in the reproductive tract of cycling and pregnant mamals. It is thought that estrogen exerts its cellular effects by regulating the expression of specific target genes. We utilized a messenger RNA differential display method to identify the genes whose expression is modulated by estrogen in the preimplantation rat uterus. Here we report the cloning of a novel gene (ERG1) that is tightly regulated by estrogen in two key reproductive tissues, the uterus and oviduct. Spatio-temporal analyses reveal that ERG1 mRNA is expressed in a highly stage-specific manner in the uterus and oviduct, and its expression is restricted to the surface epithelium of both of these tissues. Nucleotide sequence analysis of the full-length ERG1 cDNA indicates that it has an open reading frame of 1821 nuceotides encoding a putative protein of 607 amino acids with a single transmembrane domain and a short cytoplasmic tail. The extracellular part of the protein contains several distinct structural motifs. These include a zona pellucida binding domain, which is present in a number of proteins such as the zona pellucida sperm binding proteins, and uromodulin, In addition, there is a repeat of a motif called CUB domain, which exists in a number of genes involved in development and differentiation such as bone morphogenetic protein 1 (BMP1). Although the precise function of ERG1 eludes us presently, its unique pattern of expression in the uterus and oviduct and its regulation by estrogen, a principal reproductive hormone, lead us to speculate that this novel gene plays an important role in events during the reproductive cycle and early pregnancy.The steroid hormone estrogen modulates the structure and function of the female reproductive tissues, such as the uterus and oviduct, by eliciting an array of biochemical responses in these tissues. Estrogen critically influences the transport of the fertilized egg through the oviduct into the uterus (1, 2). Estrogen also promotes the growth, differentiation, and remodeling of the uterus at various physiological states, such as the reproductive cycle and pregnancy (3-6). The cellular actions of this hormone are mediated through its nuclear receptors, which function as ligand-inducible transcription factors (7-10). Previous studies in rodents employing immature and ovariectomized model systems have demonstrated that in the uterus, estrogen modulates the expression of genes that are likely to be involved in the regulation of cell growth and proliferation. These include the genes encoding protooncogenes, such as c-fos and c-myc, growth factors, such as epidermal growth factor and insulin-like growth factor-1, and their receptors (11-21). The identity of the majority of estrogen-regulated genes that mediate the uterotropic hormonal responses to estrogen, however, remains largely unknown. In order to understand the role of estrogen in complex physiological processes, such as the changes in the endometrium during the reprodu...

The growth-regulating actions of IGFs are modulated by their binding proteins (IGFBPs). The serum concentration of IGFBP-1 is down-regulated by insulin, and in-vitro studies have demonstrated that IGFBP-1 secretion from various tissues and cells can be stimulated by theophylline, forskolin, oestrogen and progesterone. We have studied the effects and mechanisms of thyroid hormone action on IGFBP-1 gene expression and secretion by human hepatoma cells in vitro. Tri-iodothyronine dose-dependently enhanced IGFBP-1 secretion in serum-free HepG2 cell cultures after 24-48 h of exposure, as measured by a specific immunofluorometric assay. This was accompanied by an increase (+ 50%) in the amount of IGFBP-1 mRNA, which could be prevented by cycloheximide, a protein synthesis inhibitor. Cycloheximide transiently enhanced (+ 200%) the accumulation of IGFBP-1 mRNA at 3-12 h of incubation, when no effect of tri-iodothyronine was observed. It is concluded that thyroid hormone stimulates IGFBP-1 secretion slowly by enhancing IGFBP-1 gene expression by a protein mediator. The acute stimulation of IGFBP-1 gene transcription by cycloheximide associates this gene with a number of growth-related genes encoding growth- and tumour-associated peptides.

We carried out a study on the effect of T4 withdrawal on serum insulin-like growth factor-binding protein-1 (IGFBP-1) levels in 10 thyroidectomized patients with papillary thyroid cancer receiving T4 replacement therapy. As controls we also measured serum sex hormone-binding globulin (SHBG) levels, known to be regulated by T4. Each patient acted as his/her own control, studied both before and 30 days after T4 withdrawal. On the average, the IGFBP-1 level decreased by 36% from 66 +/- 9 to 44 +/- 8 micrograms/L (mean +/- SE; P = 0.0007) during T4 withdrawal. The SHBG level decreased by 47% from 78 +/- 17 to 46 +/- 13 nmol/L (P = 0.0001). The fasting insulin levels were unaffected by T4 withdrawal. There also was an 18% decline in the serum IGF-I concentration after T4 withdrawal (P = 0.011). It is concluded that in athyreotic patients receiving T4, withdrawal of the drug decreases serum IGFBP-1 and SHBG concentrations and IGF-I levels. In view of the possible role of IGFBP-1 in glucose counterregulation, these results indicate a novel mechanism by which T4 may exert its metabolic action on liver carbohydrate metabolism.

Insulin-like growth factor-I (IGF-I) enhances and epidermal growth factor (EGF) inhibits gonadotrophin-induced aromatization in granulosa cells. Our previous studies have shown that human ovarian granulosa cells synthesize insulin-like growth factor-binding protein-1 (IGFBP-1) which inhibits IGF-stimulated DNA synthesis. The present study addresses the effect of EGF and gonadotrophins in the regulation of IGFBP-1 release by human granulosa cells cultured in serum-free medium. At concentrations of 1-100 micrograms/l EGF was found to stimulate IGFBP-1 secretion. This was not due to cell proliferation, as the viable cell count remained unaffected. Growth hormone and gonadotrophins had no effect on IGFBP-1 secretion when added alone to culture medium. These results suggest that EGF regulates IGFBP-1 secretion in human granulosa-luteal cells.

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