Gonadotrophin-releasing hormone (GnRH) agonists and antagonists have been widely used to prevent premature LH surge during ovarian stimulation. However, studies have shown a significantly lower serum oestradiol concentration on the day of human chorionic gonadotrophin administration for cycles using GnRH antagonist. This study compared aromatase gene expression in granulosa lutein cells from 50 women randomly assigned to receive either GnRH agonist (group 1, n=28) or GnRH antagonist (group 2, n=22). The cellular mechanism involved in the observed effects was also investigated. GnRH antagonist treatment significantly affected serum oestradiol concentration (1894+/-138 versus 1074+/-63 pg/ml; P < or = 0.001), follicular-fluid oestradiol concentration in large follicles (18,565+/-2467 versus 10,184+/-1993 pg/ml; P < or = 0.05), aromatase activity (9600+/-1179 versus 5376+/-997 fmol/10(6) cells/h; P < or = 0.05) and mRNA aromatase/mRNA glyceraldehyde 3-phosphate dehydrogenase (15+/-3 versus 6+/-1; P < 0.05). Protein kinase C (PKC) activity in granulosa lutein cells from the GnRH antagonist group was 2.5-fold higher than in the GnRH agonist group. In-vitro experiments showed that selective down-regulation of PKC was only observed in GnRH-desensitized granulosa lutein cells. This report suggests that, in granulosa lutein cells, the modulation of the FSH-induced protein kinase A pathway by PKC was different in agonist versus antagonist cycles.

LH/human CG (LH/hCG) high affinity binding sites were detected in crude membrane preparations of rat uteri. There was little competition for receptor occupancy between hCG and ovine FSH (oFSH) (0.05%) and no competition between hCG and ovine PRL (less than 0.01%). No similar binding sites were detected in crude membrane preparation of heart, kidney, skeletal muscle, liver, and lung tissues. Concentrations of uterine unoccupied binding sites (RLH) were determined for each stage of the 4-day estrous cycle. The RLH were found in all preparations of metestrus uteri (n = 10) but only in some preparations from the other stages of the estrous cycle (1 of 7 on proestrus, 3 of 4 on estrus, 5 of 7 on diestrus). The concentration of uterine RLH varied throughout the estrous cycle with highest values during the metestrus (1.50 +/- 0.15 fmol/mg protein) and lowest values during the proestrus (less than 0.2 fmol/mg protein). The affinity constant for hCG of uterine RLH remained constant during the estrous cycle (about 0.8 x 10(11) M-1) and was nearly identical to that of rat ovarian receptors. On metestrus, RLH concentration appeared to be approximately 35-fold lower in the uterus than in the ovaries when expressed per mg protein (1.50 +/- 0.15 vs. 52.83 +/- 3.61 fmol/mg protein) but only 20 times lower when expressed per organ (2.2 vs. 48.3 fmol/organ). The estrous cycle-related changes of uterine RLH concentration, together with our data establishing an in vitro influence of hCG on progesterone metabolism in rat uterus, suggest that some uterine functions could be directly regulated either by LH from the pituitary or during early pregnancy by an LH-like substance originating from the embryo.

Breast cancer (BC) is the primary cause of cancer-related mortality among women. Patients who express the estrogen receptor (ER), which mediates the tumorigenic effects of estrogens, respond to antihormonal therapy. Loss of ER expression or acquired resistance to E2 is associated with aggressive malignant phenotypes, which lead to relapse. These BC subtypes overexpress syndecan-1 (SDC1), a transmembrane heparan sulfate proteoglycan that mediates angiogenesis as well as the proliferation and invasiveness of cancer cells. We showed here that the activation of ER-alpha (ERα) by estrogens induces downregulation of SDC1 expression in ER(+) MCF7 cells but not in T47D cells. Loss of ERα expression, induced by RNA interference or a selective ER downregulator, led to subsequent SDC1 overexpression. E2-dependent downregulation of SDC1 expression required de novo protein synthesis and was antagonized by treatment with BAY 11-7085, an irreversible inhibitor of IκBα phosphorylation, which inhibits the activation of NFκB. Downregulation of SDC1 expression required ERα and activation of IKK, but was independent to downstream transcriptional regulators of NFκB. BAY 11-7085 prevented E2-mediated phosphorylation of ERα on Ser118, increasing its proteasomal degradation, suggesting that IKK stabilized E2-activated ERα, leading to subsequent downregulation of SDC1 expression. Our results showed that sustained ER signaling inhibits SDC1 expression. Such antagonism elucidates the inverse correlation between SDC1 and ER expression in ER(+) BC as well as the overexpression of SDC1 in hormone receptor-negative BC subtypes with the most aggressive phenotypes. These results identify SDC1 as an attractive therapeutic target for BC as well as for other endocrine-associated cancers.

Major changes in the cAMP-dependent signal transduction pathway triggered by FSH take place during transition of rat Sertoli cells from proliferative to the quiescent/terminally differentiated state. Using Sertoli cell cultures isolated from 10-, 20-, and 30-day-old rats, we recorded a specific increase in PDE4 activity in both the soluble and particulate subcellular fractions of 20-day-old Sertoli cells, which also displayed the highest cAMP response to FSH and the highest FSH-induced increase in PDE4 activity in both subcellular compartments. RT-PCR and immunoblotting experiments showed that almost all the PDE4D isoforms, known as the main cAMP-regulated rolipram-sensitive PDE in Sertoli cells, were expressed throughout the early postpartum period, whereas only the short PDE4D isoforms (PDE4D1 and PDE4D2) were transcriptionally regulated by FSH. Unexpectedly, the immunoblot data also revealed that the soluble PDE4 activities were mainly related to the long PDE4D isoforms and that short PDE4D1 was predominantly particulate. The subcellular distribution and expression of PDE4D proteins were unaffected by the developmental status of the Sertoli cells. Only the expression of short PDE4D1 appeared to be upregulated by FSH and only in 20-day-old Sertoli cells, which suggests phenotype-dependent differential regulation of Pde4d1 mRNA translation. Resensitization of the cAMP response to FSH in 20-day-old Sertoli cells was also associated with the highest FSH-induced transient increase in both soluble and particulate PDE4 activities, which suggests developmental changes in the PKA-mediated upregulation of the catalytic activities of long PDE4D. Such alterations may be involved in the phenotype-dependent alterations in FSH receptor coupling with its associated G proteins in rat Sertoli cells.