The luteal phase of nonsupplemented cycles after ovarian superovulation with human menopausal gonadotropin and the gonadotropin-releasing hormone antagonist Cetrorelix

OBJECTIVE: To analyze the luteal phase of six patients undergoing controlled ovarian hyperstimulation (COH) with hMG and a new GnRH antagonist, Cetrorelix, without receiving luteal phase supplementation. DESIGN: Phase II study involving the first six patients who did not receive luteal phase support. SETTING: Tertiary referral center. PATIENT(S): Six healthy women undergoing COH for assisted reproductive techniques. INTERVENTION(S): Oocyte retrieval was performed 36 hours after hCG administration, followed by embryo transfer 2 days later. No luteal phase supplementation was given. MAIN OUTCOME MEASURE(S): Serum E2, progesterone, LH, and FSH concentrations were measured. RESULT(S): The length of the luteal phase was < or =12 days in three of the six patients. One of the patients in whom the luteal phase was >12 days had a low serum progesterone concentration (2.9 ng/mL) on day 10. Serum LH concentrations decreased after the preovulatory hCG injection in all patients. However, a progressive increase in LH was observed after day 7, reaching normal values. CONCLUSION(S): Corpus luteum function seems to be impaired in cycles that are stimulated with hMG and the GnRH antagonist Cetrorelix.     

Studies on the mechanism of embryo implantation

Implantation is a complex process accomplished by synchronization and interactions between embryo and endometrium by local exchange of signals including a number of cytokines and growth factors and direct cell-cell and cell-matrix contact. However, the research in early events of human implantation is still in its infancy. This presentation comprises the results of our attempts to investigate the mechanisms of human implantation process in its early stage by cell-biological method, including establishment of experimental implantation model in vitro. 1. Human trophoblast of early stage of gestation showed active cell locomotion, active endocytosis, and invasion of endometrial cell monolayer in mixed cultures. Trophoblast invasion was later arrested by transformed endometrial cells similar to decidual cells in vivo. These results appeared to indicate the interactions between trophoblast and endometrial cells in implantation. 2. Coculture system of rabbit preimplantation blastocyst and endometrial epithelium reformed from isolated endometrial epithelial cells on basement membrane matrix (Matrigel) simulated the in vivo rabbit implantation processes. This coculture system may provide a useful experimental implantation model. 3. A human trophoblast cell line was established from chorionic tissues of normal early pregnancy. These cells were cytotrophoblast-like morphology and endocrine functions. They formed the villous structures similar to those in vivo in culture on Matrigel and invasion of Matrigel was observed. These indicated the extracellular matrix may affect the morphology and function of invading trophoblast in implantation site. 4. Human endometrial epithelial single cells were cultured on Matrigel. Reconstruction of gland followed by epithelium formation quite similar to in vivo structures by migration and proliferation of isolated cells was demonstrated. Height of gland was promoted by estrogen and initiation of epithelization was upregulated by platelet-derived growth factors. This system revealed the extracellular matrix regulated morphogenesis of endometrial epithelium in vivo and is an essential substrate in experimental implantation model of endometrial epithelium. 5. Parallel cultures of endometrial epithelial cells on Matrigel were carried out with the IVF. ET patients to evaluate the endometrial morphology at time of ET. Endometrial cultures were initiated in previous cycles on Matrigel and the sera of patients were added to her own cultures from 1st day of IVF treatment cycle. Evaluation of reformed epithelium revealed the apparently unsuitable morphology for implantation in group of patients who eventually failed in pregnancy. This system may provide a useful measures in evaluation of endometrial receptivity and modality of treatment for endometrial aberrations. 6. Cyclic changes of extracellular matrix components in endometrium were investigated. Collagen I, III, IV, V were immunohistochemically estimated. Relative levels of all types of collagen except for collagen V declined at early secretory phase. In rodents, not only collagen but also laminin and fibronectin levels declined at early secretory phase. These changes may facilitate trophoblast invasion of endometrium. Collagen V distributed in myometrial surface was found to consist of subunit (alpha 1)2 alpha 2 and trophoblast growth was inhibited on substrate of alpha 1 subunit. Collagen V in myometrial surface may have a role in blocking trophoblast invasion. 7. HGF (hepatocyte growth factor) mRNA was demonstrated to be expressed during menstruation and secretory phase in endometrium distinctly and its receptor in endometrial epithelial cells and decidual cells. Positive correlation between plasma HGF levels and ultrasonographic thickness of endometrium was observed at late secretory phase. Recombinant HGF promoted proliferation of endometrial epithelial cells and decidual cells and upregulated initiation of endometrial epithelization of Matrigel.     

Pulsatile release of LH and secretion of ovarian steroids in sheep during the luteal phase of the estrous cycle

The concentration of LH and progesterone in jugular venous plasma and the secretion of steroids by the ovary were measured every 10 minutes for 2 hours on days 12, 14, and 16 of the estrous cycle in 5 ewes with utero-ovarian autotransplants. A pulse of LH occurred about once every 2 hours, when the concentration rose from a basal level of 0.57 +/- 0.08 ng/ml to a peak of 2.97 +/- 0.57 ng/ml. Within 5 minutes of the pulse of LH, the secretion of estradiol (an exclusive product of the follicle) rose rapidly from a basal level of 0.75 +/- 0.12 ng/min to reach a peak value of 2.16 +/- 0.33 ng/min in about 30 minutes. In contrast, the secretion of progesterone from the corpus luteum, and the concentration of progesterone in the peripheral plasma changed very little following the pulse of LH. The secretion of androstenedione, which arises from the follicle and corpus luteum, increased from 3.03 +/-0.75 ng/min to 7.85 +/- 1.78 ng/min by 30 minutes after the pulse of LH. These findings indicate that the follicle, and possibly the stroma, respond rapidly to episodic fluctuations in the concentration of LH and are probably involved in the negative feedback loop between the ovary and the hypothalamic pituitary system. The fluctuations in the secretion of progesterone from the corpus luteum, on the other hand, are unrelated to pulses of LH.     

The ovarian hyperstimulation syndrome in extracorporeal oocyte fertilization and the transfer of cleavage-stage embryos

Major clinical characteristics of ovarian hyperstimulation syndrome are described and criteria of predicting the course of this disease and measures to prevent it presented. Statistical data on the presence of this syndrome in various forms during various schemes of superovulation stimulation are offered. Optimal schemes for the treatment of the condition are suggested.     

Effect of methoxychlor on implantation and embryo development in the mouse

Methoxychlor (MXC) has been shown to have adverse effects on reproductive functions. However, it has not been fully determined whether the effects of MXC on reproduction are due to its estrogenic or antiestrogenic effects. Therefore, to further elucidate the estrogenic action of this pesticide in the mouse, we studied the effect of MXC on implantation and embryo development. MXC was found to initiate implantation in most delayed implanting mice at 400 microg/g body weight. However, at the higher dose of 800 microg/g body weight, MXC initiated implantation in only 50% of animals and the number of embryos implanting was significantly decreased (P < 0.05). It was determined that MXC inhibited implantation in intact pregnant mice only when given on Day 1 or Day 2 at 800 microg/g but not at lower doses or later in the preimplantation period. Embryonic development and transport were delayed on Days 3 and 4 in these animals. Finally, reciprocal embryo transfers with donor embryos recovered from MXC-treated animals (800 microg/g body weight on Day 1) transferred to untreated recipients resulted in no implantation compared to 79% implantation when donor embryos were treated with vehicle. These data indicate that MXC acts as an estrogen agonist at the level of the uterus and oviduct but as an antiestrogen in the ovary. In addition, MXC appears to alter normal preimplantation embryonic development. These results suggest the need for further studies to assess the mechanism of action of MXC in preimplantation embryos.     

Haematocrit, leukocyte and platelet counts and the severity of the ovarian hyperstimulation syndrome

Previous studies have shown that severe ovarian hyperstimulation syndrome (OHSS) is secondary to circulatory dysfunction due to the simultaneous occurrence of increased vascular permeability and marked arteriolar vasodilation which lead to an intense homeostatic stimulation of the renin-aldosterone and sympathetic nervous systems and antidiuretic hormone (ADH). In the present report, we have investigated the correlation between changes in haematocrit concentration, and white blood cell (WBC) and platelet counts and the severity of OHSS, as assessed by these markers of effective intra-arterial blood volume, in a series of 50 patients. In comparison with recovery values (4-5 weeks after hospital discharge), OHSS patients showed arterial hypotension, tachycardia, oliguria, very high plasma concentrations of renin, aldosterone, norepinephrine and ADH, and increased mean haematocrit values and WBC and platelet counts. The haematocrit concentration values were directly related to the plasma concentrations of vasoactive substances (plasma renin activity, aldosterone, norepinephrine and ADH) during OHSS (P < 0.001). In contrast, no correlation was evident between WBC or platelet counts and neurohormonal measurements during the syndrome. It is concluded that haematocrit, but not WBC or platelet counts, can act as a biological marker of the severity of OHSS as indicated by plasma measurement of volume-dependent endogenous vasoactive substances.     

First established pregnancy after controlled ovarian hyperstimulation with recombinant follicle stimulating hormone and the gonadotrophin-releasing hormone antagonist ganirelix (Org 37462)

This case report describes the first established pregnancy after the use of gonadotrophin-releasing hormone (GnRH) antagonist, ganirelix (Org 37462; Organon), to prevent a premature luteinizing hormone surge during ovarian hyperstimulation with recombinant human follicle stimulating hormone (rhFSH). The pregnancy progressed normally and ended with the birth of a healthy boy and a girl after an elective Caesarean section at gestational age of 37 weeks. This case illustrates, for the first time, the use of a GnRH antagonist in combination with a pure FSH preparation for ovarian stimulation.     

The role of vascular endothelial growth factor and interleukins in the pathogenesis of severe ovarian hyperstimulation syndrome

Recent studies have suggested that the proliferation of malignant gliomas may result from activation of protein kinase C (PKC)-mediated pathways; conversely, inhibition of PKC may provide a strategy for blocking tumor growth. In the current studies, we examined the effect of a novel PKC inhibitor, calphostin C, which is a selective, highly potent, photo-activatable inhibitor of the PKC regulatory domain, on the proliferation and viability of three established and three low-passage malignant glioma cell lines, four low-passage low-grade glioma cell lines, and in adult human and neonatal rat non-neoplastic astrocyte cell lines in vitro. Under light-treated conditions, calphostin C consistently inhibited cell proliferation in each of the tumor cell lines and in the neonatal rat astrocyte cell line with a 50% effective concentration of 30 to 50 ng/ml (40 to 60 nm), which was comparable to the previously reported median inhibitory concentration (IC50) for PKC inhibition by calphostin C. Complete elimination of proliferation was achieved at concentrations of 50 to 100 ng/ml (60 to 125 nM). Cell viability decreased sharply with calphostin C concentrations of 100 to 300 ng/ml (125 to 380 nM). In contrast, under light-shielded conditions, calphostin C had a comparatively modest effect on cell proliferation and viability, with a median effective concentration of approximately 300 ng/ml. No significant inhibition of proliferation was noted in the non-neoplastic adult astrocyte cell line under either light-treated or light-shielded conditions. These findings provide further evidence that PKC may play an essential role in mediating the proliferation of both benign and malignant glioma cells in vitro and may also contribute to the proliferation of non-neoplastic immature astrocytes. Light-sensitive inhibition of proliferation and viability by agents such as calphostin C may provide a novel strategy for applying photodynamic therapy to the treatment of neoplastic glial cells.     

Effect of corticotropin-releasing factor on the pituitary-ovary axis in human luteal phase

In order to test the possible negative effect of corticotropin-releasing factor (CRF) on the pituitary-gonadal axis, two gonadotropin-releasing hormone (GnRH) tests, on two consecutive days, were performed in five normally cycling women in the midluteal phase of the menstrual cycle. The first GnRH test was performed after 2 h during which time basal blood samples were collected every 10 min. On the second day, the GnRH test was repeated after a 2-h CRF infusion which was continued throughout the GnRH test. Our results demonstrate that while CRF increases cortisol levels it affects neither the basal nor the GnRH-stimulated gonadotropin levels. Similarly, both basal and luteinizing hormone (LH)-stimulated progesterone concentrations were not modified by the CRF infusion thereby also excluding a possible direct effect of the peptide on corpus luteum steroidogenesis.     

Contribution of Doppler color ultrasonography to the screening and surveillance of ovarian hyperstimulation syndrome

Color and power Doppler producing ovarian vascular mapping allows to realize a new functional approach of ovarian hyperstimulation syndrome (OHS); local vascular factors (ovarian angiogenesis, increase in cappillary permeability) released by hCG injection (LH effect) have in fact an important part in this syndrome. Vaginal Doppler sonography allows: -before induction, to predict potential risk of hyperstimulation: polycystic ovaries, spontaneous stromal hypervascularization; -in monitoring of ovulation stimulation, to reduce risk of multiple pregnancy and to detect early hyperstimulation, decision of hCG injection depends on number of follicles able to be fertilized (> 15 mm) and luteinized (> 10 mm); -to assess severity of OHS and search complications: ascites, hydrothorax, ovarian torsion and deep venous thrombosis. So OHS prevention imposes a monitoring by Doppler sonography and estradiol assay of all cycles induced by gonadotropins.     

In controlled ovarian hyperstimulation, steroid production, oocyte retrieval, and pregnancy rate correlate with gene expression of vascular endothelial growth factor

PURPOSE: Whether the gene expression of vascular endothelial growth factor (VEGF) in human granulosa cells is a predictor of fertilization was evaluated in patients participating in an in vitro fertilization program. METHODS: Fifty patients with normal ovaries who were participating in an in vitro fertilization program at the University of Milan, San Raffaele Scientific Institute, were included in the study. We correlated E2 and P serum levels on the day of oocyte collection, the number of follicles, oocytes collected, and fertilized, and pregnancies with mRNA for VEGF of luteinizing granulosa cells obtained at the time of oocyte retrieval. RESULTS: Comparing E2 and P serum levels, the number of follicles, oocytes collected and fertilized, and pregnancies with gene expression for VEGF, we found a positive correlation. E2 and P serum levels were higher in patients with increased VEGF (P < 0.01). Furthermore, there were more follicles, oocytes collected and fertilized, and pregnancies in patients with maximum expression of VEGF, and the difference was statistically significant (P < 0.05). CONCLUSIONS: Our results suggest that VEGF may be important for vascular development during follicular growth and luteal differentiation, oocyte maturation, and fertilization.     

Placental protein 14 in endometrium during menstrual cycle and effect of early luteal phase mifepristone administration on its expression in implantation stage endometrium in the rhesus monkey

Placental protein 14 (PP14) is a glycoprotein which is secreted by secretory phase endometrium and decidua in women. Despite the suggestion that PP14 is involved in the process of endometrial maturation for blastocyst implantation, our understanding in this regard is poor. In the present study, the concentrations and distribution patterns of immunodetectable PP14 in the endometrium during proliferative and secretory phases of normal ovulatory menstrual cycles, as well as in implantation stage endometrium in naturally mated ovulatory cycles with or without early luteal phase mifepristone treatment, were investigated using the rhesus monkey as a primate model. Immunopositive PP14 was observed mainly in epithelial cells of glands and it was detected in one major immunopositive band at Mr 28 kDa in tissue homogenate and spent medium. The area of immunopositive precipitation of PP14 in glands was minimal in follicular phase endometrium, and was higher (P < 0.01) in early, mid- and late luteal phase endometrium compared with that in pre- and periovulatory phases of the cycle, but there was no change in its area profile in the glandular compartment throughout the luteal phase. Immunopositivity for PP14 in luminal contents of gland displayed an increasing profile from early to late secretory phases. Thus, the concentrations and the distribution of immunodetectable PP14 in luteal phase endometrium of the rhesus monkey showed marked similarity with those of human endometrium during the natural menstrual cycle. Although there was no marked change in the band characterstics for the protein in implantation stage endometrium following early luteal phase mifepristone treatment, it was markedly decreased (P < 0.01) in tissue homogenate and in vitro spent medium along with a lesser (P < 0.02) degree of immunoprecipitation in the glands in implantation stage samples of mifepristone treatment group compared with that in control group samples. Thus, the contragestional effect of early luteal phase mifepristone treatment appears to be associated with a decrease in the concentration of immunodetectable PP14 in implantation stage endometrial glands and its secretion in the rhesus monkey. It remains to be seen whether this decline is caused from direct antiprogesterone action on endometrial glands during progesterone dominance, or secondarily from associated retarded development of endometrium.     

Effect of estradiol-17 beta on basal and hCG stimulated progesterone secretion by porcine luteal cells isolated in various stages of the luteal phase

T cells of mice display V beta-specific reactivity for a spectrum of mouse mammary tumor virus (Mtv) antigens; confrontation with these antigens during ontogeny causes substantial "holes" in the T cell repertoire. Since endogenous Mtv antigens are rare in other species, the question arises whether V beta-specific recognition of Mtv antigens is unique to mice. To examine this question, rat T cells were allowed to differentiate from stem cells in severe combined immunodeficiency (SCID) mice. These rat-->mouse xenochimeras were prepared under a variety of conditions. The results show that rat T cells are strongly reactive to mouse Mtv antigens, both in terms of tolerogenicity and immunogenicity. In fact, the V beta specificity of rat and mouse T cells for Mtv antigens is almost indistinguishable.     

Leptin concentrations in the follicular phase of spontaneous cycles and cycles superovulated with follicle stimulating hormone

It has been reported that oestradiol may play a role in the production of leptin from adipocytes. To investigate this relationship further, nine normally ovulating women were studied during two menstrual cycles, i.e. an untreated spontaneous cycle and a cycle treated with follicle stimulating hormone (FSH) from cycle day 2 until the day of human chorionic gonadotrophin (HCG) injection. Serum leptin values on cycle day 2 did not differ significantly between the spontaneous and the FSH cycles. In the spontaneous cycles, leptin values declined gradually and significantly up to day 7 and then increased progressively up to the day of luteinizing hormone (LH) surge onset, at which point they achieved the highest values. In the FSH cycles, serum leptin values increased gradually and significantly up to day 6, remaining stable thereafter, and were in the midfollicular phase significantly higher than in the spontaneous cycles. Significant positive correlations were found between mean values of leptin and mean values of oestradiol during the second half of the follicular phase in the spontaneous cycles and during the first half in the FSH cycles. A significant negative correlation was found between these two parameters in the spontaneous cycles during the first half of the follicular phase. Serum leptin levels were significantly higher in the midluteal than in the follicular phase in both cycles. These results demonstrate for the first time significant changes in leptin values during the follicular phase of the human menstrual cycle and a significant increase during superovulation induction with FSH. It is suggested that oestradiol may be involved in the regulation of leptin production in women.     

Effects of profound suppression of luteinizing hormone during ovarian stimulation on follicular activity, oocyte and embryo function in cycles stimulated with purified follicle stimulating hormone

The effects of profound suppression of circulating luteinizing hormone (LH) during the follicular phase of in-vitro fertilization cycles were explored in normal women during treatment with a gonadotrophin-releasing hormone analogue and exogenous purified follicle stimulating hormone. Ovarian responses to treatment and the capacity of supernumerary embryos to undergo blastocyst formation were examined in groups of patients defined by the concentration of plasma LH in the mid-follicular phase. Concentrations < or = 0.5 IU/I diagnosed the group with profoundly suppressed LH (相似文献   

Daily measurements and in-vitro effects of human chorionic gonadotrophin in the early luteal phase

The purpose of the present study was to analyse daily measurements of human chorionic gonadotrophin (HCG) in in-vitro fertilization (IVF) cycles and to reproduce the effects of HCG in vitro using human granulosa-luteinized cells from the same patients. The study population consisted of nine women undergoing IVF because of tubal infertility in whom blood was drawn every 24 h from the day of the ovulatory dose of HCG (10,000 IU) until 6 days after ovum pick-up. Granulosa-luteal cells from the follicular aspirates were collected and cultured in vitro up to 6 days in the presence of increasing concentrations (0, 0.01, 0.1, 1.0 and 100.0 IU/ml) of HCG. Serum progesterone and HCG in vivo as well as progesterone accumulation in vitro on days 2, 4 and 6, were the main outcome measures. Maximum HCG concentrations (0.25 IU/ml) were reached the day before ovum pick-up, and continuously decreased until day 6 after ovum retrieval. HCG did not stimulate progesterone production in vitro at any dose tested until day 6 after ovum pick-up. Then, 0.01 IU/ml resulted significantly (P < 0.05) stimulatory compared to controls, while 1.0 IU/ml was inhibitory (P < 0.05). It is concluded that HCG supplementation in an IVF cycle is unnecessary until day 6 after ovum pick-up. On day 6, progesterone production is stimulated with very low concentrations of HCG.