Hum. Reprod. Advance Access originally published online on November 16, 2006
Human Reproduction 2007 22(3):676-687; doi:10.1093/humrep/del445
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Endocrine profile in serum and follicular fluid differs after ovarian stimulation with HP-hMG or recombinant FSH in IVF patients
1 Follicle Biology Laboratory and Center for Reproductive Medicine of the Academisch Ziekenhuis Vrije Universiteit Brussel, Brussels, Belgium 2 Rigshospitalet, Fertility Clinic, Copenhagen, Denmark 3 Center for Reproductive Medicine of the Academisch Ziekenhuis Vrije Universiteit Brussel, Brussels, Belgium and 4 Ferring Pharmaceuticals A/S, Obstetrics & Gynaecology, Clinical Research & Development, Copenhagen, Denmark
5 To whom correspondence should be addressed at: Follicle Biology Laboratory and Center for Reproductive Medicine of the Vrije Universiteit Brussel, Laarbeeklaan 101, 1090 Brussels, Belgium. E-mail: johan.smitz{at}az.vub.ac.be
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Abstract |
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BACKGROUND: Highly purified menotrophin (HP-hMG) has been associated with fewer oocytes retrieved and a higher proportion of top-quality embryos compared with recombinant FSH (rFSH). METHODS: A randomized, assessor-blind, multinational trial in 731 women undergoing IVF after stimulation with HP-hMG (MENOPUR) (n = 363) or rFSH (GONAL-F) (n = 368) following a long GnRH agonist protocol was conducted. Blood was collected before, during and after stimulation. Fluid was collected from follicles
17 mm. RESULTS: Serum androstenedione, total testosterone and free androgen index (FAI) were higher (P < 0.001) with HP-hMG than with rFSH after starting stimulation. At the end of stimulation, serum estradiol was higher (P = 0.031) with HP-hMG, whereas progesterone was higher (P < 0.001) with rFSH, even after adjusting for ovarian response. Serum LH was not different between treatments. Mean mid- and end-follicular hCG levels in the HP-hMG group were 2.5 and 2.9 IU/l, respectively. Follicular fluid levels of FSH, LH, hCG, androstenedione, testosterone, FAI and estradiol and ratios of estradiol : androstenedione, estradiol : total testosterone and estradiol : progesterone were higher (P < 0.001) with HP-hMG, whereas progesterone was higher (P < 0.001) with rFSH. CONCLUSION: Major differences in serum and follicular fluid endocrine profile exist after stimulation with HP-hMG or rFSH. Exogenous LH activity induces a differential endocrine environment influencing oocyte quantity and quality, which may be of relevance for clinical outcome.
Key words: endocrine/follicular fluid/IVF/highly purified menotrophin/recombinant FSH
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Introduction |
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In the natural cycle, follicle growth is driven by a delicate interplay of FSH and LH that affects theca and granulosa cells, leading to the selection of a single dominant follicle through a series of feedback mechanisms (van Santbrink et al., 1995
; Sullivan et al., 1999). FSH drives the development of the granulosa cell compartment and is essential for follicle survival and differentiation. Effects of FSH are amplified via several paracrine loops including the products of aromatization that depend on the provision of androgens by the theca cells (Hillier et al., 1994). One of the pivotal cellular actions of FSH is the induction of LH receptors on granulosa cells that will enable the granulosa cells to respond to both gonadotrophins (Zeleznik and Hillier, 1984). Through the LH receptor expression on the granulosa cells, the maturing follicle continues to develop irrespective of the physiological decline in FSH concentration that exists in the pre-ovulatory days (Zeleznik and Hillier, 1984). The concepts that threshold levels for FSH and LH are needed to achieve folliculogenesis and steroidogenesis and the effects by which inappropriately timed and/or abundant secretion of LH lead follicles into atresia (McNatty et al., 1975; Hillier, 1994) have now been clinically documented. Results from various studies have indicated that both the induction of aromatase and the presence/amount of LH and/or hCG are the principal components of the process of follicle selection (The European Recombinant Human LH Study Group, 1998; Filicori et al., 1999, 2001, 2002; Sullivan et al., 1999; Loumaye et al., 2003; Platteau et al., 2006).
Within the field of controlled ovarian stimulation for IVF or ICSI, where high doses of gonadotrophins are administered, the role of LH activity is still heavily debated (Collins, 2003). Among the many reasons for the disagreement on the importance of LH activity are the variability of the endocrine background of the patient, the different definitions of ‘LH depletion’ related to the functional sensitivity and heterogeneity of the routine diagnostic tests for LH, the differential effects of various GnRH agonist protocols and the differences in biochemical composition of gonadotrophin preparations. Measuring the bioactivity of LH by using an immunoassay is inherently approximative (Niccoli et al., 1996). The value of a serum LH concentration depends on both the specificity and the sensitivity characteristics of the immunoassay used (Costagliola et al., 1994). It is also difficult to uniformly quantify the exposure to LH activity brought by the different gonadotrophin preparations, as there are different types of exogenous compounds with LH activity, i.e. LH or hCG. A recent study addressed the differences in composition in glycoproteins between the different types of menotrophins on the market (Wolfenson et al., 2005), which in a large part could be attributable to hCG molecules. The diverse kind of menotrophin preparations used in clinical evaluations combined with the plethora of immunoassays with different threshold figures used to quantify the degree of LH suppression leads to the large heterogeneity in published figures on the prevalence and degree of LH suppression in GnRH agonist-desensitized patients, ranging from 12 to 70% (Fleming et al., 2000; Westergaard et al., 2000; Esposito et al., 2001; Humaidan et al., 2002).
A primary distinction is that the clinical situation associated with exogenous LH/hCG supplementation during stimulation may represent a very different scenario than a pathophysiological condition in patients with elevated endogenous LH levels after pituitary down-regulation. Several earlier studies have found a negative impact of a high endogenous LH on oocyte or embryo quality and clinical outcome (Stanger and Yovich, 1985; Howles et al., 1986; Homburg et al., 1988; Watson et al., 1993), but the mechanisms behind this are still enigmatic. Also intriguing are the clinical reports addressing the effects of an exogenous contribution of LH/hCG on the follicular response and more particularly the quality of the oocyte cohort, its developmental competence and implantation potential (Filicori et al., 1999; Humaidan et al., 2004; Platteau et al., 2004; Humaidan, 2006). More insight into this important question can be acquired methodologically by using a source of exogenous LH activity supplementation (i.e. hCG) different from the source of endogenous LH activity (i.e. LH).
As part of a prospective study comparing highly purified menotrophin (HP-hMG) and recombinant FSH (rFSH) with ongoing pregnancy rate as a primary end-point (Nyboe Andersen et al., 2006), the evaluation of the endocrine profile in serum and follicular fluid was performed to support the concept that LH activity from the start of stimulation would induce a higher serum androgen load, impacting follicle selection, and would change the ratio of intrafollicular steroid balance (estradiol : progesterone, estradiol : androstenedione and estradiol : total testosterone), impacting embryo quality. It is suggested that the different ratio of estrogens to androgens attributed to the presence/absence of LH activity could be responsible for the difference between treatments in follicular dynamics during the recruitment and selection phase. Exposure to LH activity is expected to increase androgen load, and because androgens have a stimulatory role in intracellular regulation of granulosa cell function via effects on FSH receptor expression, on insulin-like growth factor-1 (IGF-1) and on the aromatase gene (Luo and Wiltbank, 2006), a differential status on markers of granulosa differentiation would be expected in women exposed to rFSH or HP-hMG. Exploration of intrafollicular levels and ratios of major steroids, and levels of IGF-1 and insulin and of markers of granulosa differentiation would provide an additional insight into the impact of exogenous LH activity. Furthermore, it would be interesting to study the concept that exogenous LH activity supplementation during stimulation does not lead to early luteinization, as may be detected by investigating progesterone, vascular endothelial growth factor (VEGF), inhibin A and inhibin A : inhibin B.
This prospective study included only patients eligible for IVF and not patients requiring ICSI. This was based on methodological considerations (Arce et al., 2005) and a previous report suggesting a differential treatment outcome between the gonadotrophin preparations tested in the present study when used in IVF cycles (Platteau et al., 2004). The clinical outcome of this study and the pharmacodynamic differences in follicular development, embryo quality and endometrial echogenic profile have been the object of a separate publication (Nyboe Andersen et al., 2006).
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Study population
Women with major indications for IVF such as tubal infertility or unexplained infertility including endometriosis stage I/II and partners with mild semen abnormalities not requiring ICSI were recruited to the study. Patients were 21–37 years of age with regular menstrual cycles of 21–35 days, presumed to be ovulatory. They had been infertile for at least 1 year, except for those with proven bilateral tubal infertility. They had a uterus consistent with expected normal function, presence of both ovaries and without evidence of abnormality and normal adnexa. The early follicular phase serum levels of FSH were within normal limits (1–12 IU/l). The body mass index before inclusion in the study was in the range 18–29 kg/m2. Patients with polycystic ovary syndrome, endometriosis stage III/IV or partners with severe male factors requiring ICSI were not included in the study. Likewise, poor responders (those with more than three previously consecutive unsuccessful IVF cycles or previous cycles with >20 days of gonadotrophin stimulation, or cancellation due to limited follicular response, or <4 follicles of
15 mm) and patients with a previous IVF cycle with unsuccessful fertilization were excluded from participation. A detailed description of the study population and all inclusion and exclusion criteria are provided elsewhere along with the clinical outcome of the study (Nyboe Andersen et al., 2006).
Study design
This was a randomized, open-label, assessor-blind, parallel-group, multicentre, multinational study comparing HP-hMG (MENOPUR; Ferring Pharmaceuticals A/S, Copenhagen, Denmark) and rFSH (follitropin alfa, GONAL-F; Serono, Geneva, Switzerland). A total of 37 fertility clinics in 10 countries randomized patients to the study. The randomization of patients to treatment was stratified by age (<35 years and 35–37 years) in each centre. All investigators, embryologists, laboratory personnel and sponsor staff were blinded to treatment allocation throughout the study. Patients underwent controlled ovarian hyperstimulation following down-regulation with a GnRH agonist in a long protocol for women undergoing IVF. All patients in all centres and countries received identical type and dose of concomitant fertility treatments, i.e. GnRH agonist for down-regulation, hCG for triggering final maturation and progesterone for luteal support. Pituitary down-regulation using triptorelin acetate, 0.1 mg/day s.c. (DECAPEPTYL; Ferring Pharmaceuticals A/S), was initiated 5–7 days before the estimated start of next menses and continued until the end of gonadotrophin administration. Gonadotrophin administration was initiated when down-regulation was confirmed using transvaginal ultrasound showing no ovarian cysts, a shedded endometrium with a thickness of <5 mm or serum estradiol <50 pg/ml (0.184 nmol/l). The starting dose of HP-hMG or rFSH was 225 IU for the first 5 days, followed by individual adjustments according to the patient’s follicular response. The dose could be changed by 75 IU per adjustment and not more frequently than every 4 days. Choriongonadotrophin alfa, 250 µg s.c. (OVITRELLE; Serono), was administered to induce final follicular maturation within 1 day of observing three or more follicles of 17 mm diameter. Oocyte retrieval took place 36 h (±2 h) after hCG administration. Insemination was done via regular IVF insemination (not ICSI) at 3 h (±1 h) after oocyte retrieval. Fertilization was assessed at 20 h (±1 h), and embryo quality was assessed at 28 (±1 h), 44 (±1 h) and 68 h (±1 h) after oocyte retrieval. A top-quality embryo was defined as four to five cells on day 2, seven or more cells on day 3, equally sized blastomeres and 
20% fragmentation on day 3, and no multinucleation. The transfer of one or two embryos of minimum quality, defined as four or more cells with no cleavage arrest (i.e. cleavage must have occurred within the last 24 h) and 20% fragmentation, was done on day 3 after oocyte retrieval. Vaginal progesterone gel 90 mg/day 8% (CRINONE; Serono) for luteal support was given from the day of embryo transfer till the confirmation of clinical pregnancy (5–6 weeks after embryo transfer) or negative serum 
hCG test (13–15 days after embryo transfer). Ongoing pregnancy was determined 10–11 weeks after embryo transfer. The study procedures are described in detail in a separate publication (Nyboe Andersen et al., 2006).
Collection and handling of serum
Blood samples were obtained on day 1 of stimulation, on day 6 of stimulation, on the last stimulation day and at oocyte retrieval. The sample on day 1 was taken before the start of gonadotrophin administration, and the samples on day 6 and last stimulation day were collected at least 8 h after the previous gonadotrophin dose. Blood samples were centrifuged for 10 min at 1800 x g. Serum was stored individually at –18°C or colder at the clinic for a maximum of 2 weeks before transfer to –70°C and subsequent analysis at a central laboratory. All serum samples were analysed centrally for FSH, LH and hCG by Laboratorium für Klinische Forschung (LKF; Raisdorf, Germany) and for estradiol, progesterone, androstenedione, total testosterone and sex hormone-binding globulin (SHBG) by Capio Diagnostik (Copenhagen, Denmark).
Collection and handling of follicular fluid
Follicular fluid from at least one follicle of 17 mm from which an oocyte was retrieved was collected at oocyte retrieval. A manual was provided to all clinics with instructions on how to obtain and prepare the follicular fluid. Fluid was preferably collected from the first follicle aspirated and from follicles where flushing had not been applied. The fluid was centrifuged for 10 min at 1000 x g, after which it should be clear and not contaminated with blood. The follicular fluid was stored under the same conditions as serum. The samples were analysed for FSH, LH, hCG, estradiol, progesterone, androstenedione, total testosterone, SHBG, insulin, inhibin A, inhibin B, VEGF and IGF-1 by the hormone laboratory at Academisch Ziekenhuis Vrije Universiteit Brussel (Brussels, Belgium) and for cortisone and cortisol by the Department of Chemical Pathology, Southampton General Hospital, University of London (Southampton, UK). Fluids that were found to be contaminated either by blood cells or by flushing medium were disregarded from the analysis. Final endocrine analysis of follicular fluid was done for 335 patients in the HP-hMG group and for 341 patients in the rFSH group.
Analytical methods
An overview of analytical methods, sensitivity and total imprecision for serum and follicular fluid hormonal assessments is given in Table I. Validated laboratory immunoassay methods were selected for their sensitivity and reproducible proficiency profile to increase the precision of the measurements in serum and in follicular fluid.
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Statistical analysis
For serum and follicular fluid hormone concentrations, the treatment difference between HP-hMG and rFSH with 95% confidence interval and P-value was estimated for each parameter based on a linear model of the log-transformed data including treatment and age strata. The treatment differences estimated in this model represent the overall treatment influences, i.e. both the direct treatment effects and the effects mediated via other hormones. To evaluate the direct treatment effects on each hormone separately, we also carried out a multi-adjusted analysis. This analysis included the endocrine profile at the specific time point, the primary reason for infertility and duration of infertility as co-variates in addition to the age strata. Overall, the findings in the multi-adjusted analysis were consistent with the analysis adjusted exclusively for age strata with only minor differences. The data presentation in the Results section is focused on the age strata-adjusted analysis, but differences observed in the multi-adjusted analysis are noted in the tables. For serum estradiol and progesterone on the last stimulation day and at oocyte retrieval, the age strata-adjusted comparison between treatment groups was also adjusted for the number of follicles and oocytes retrieved, respectively.
The relationship between estradiol and testosterone and estradiol and androstenedione on day 6 and at the end of stimulation was investigated within each group using a proportionality model. Log-transformed data were analysed in a regression analysis model, and the estimates were then transformed to the original scale. The influence of the FSH concentration was explored. Data were analysed separately for each treatment group. The influence of hCG concentration on day 6 among HP-hMG-treated patients in predicting embryo quality as assessed by the local embryologists and clinical outcome was evaluated by grouping in quartiles (i.e. 25% of the patients in each group). Test for trend in hCG was conducted by assigning an ordinal score (the median) to the grouped value and then treating this score as continuous in analysis. Data were analysed using both an age strata-adjusted and a multi-adjusted (age strata, primary reason for infertility, duration of infertility and day 1 serum levels of all endocrine parameters as co-variates) approach.
The primary end-point of this clinical study was ongoing pregnancy rate (reported in Nyboe Andersen et al., 2006). No adjustment for multiplicity was performed, as there was only one primary end-point and all other end-points were considered secondary.
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Results |
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A total of 731 patients were at the end of down-regulation randomized to either HP-hMG (n = 363) or rFSH (n = 368). Demographics, baseline characteristics and the mean levels for all endocrine parameters before the start of gonadotrophin administration were comparable between the two treatment groups (Table II). All but one patient received 225 IU daily for the first 5 days. On day 6, most of the patients (60% for HP-hMG and 66% for rFSH) had the dose maintained at 225 IU, whereas 33% in the HP-hMG group and 25% in the rFSH group had the daily dose increased to 300 IU and <10% had the daily dose reduced to 150 IU. The dose of 225 IU throughout the study was maintained by 135 patients in the HP-hMG group and 171 patients in the rFSH group.
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On day 6, after stimulation with 225 IU/day, the mean number of follicles >10 mm was 4.1 in the HP-hMG group and 4.9 in the rFSH group, which was significantly different (P = 0.007). On the last stimulation day (on average day 10, irrespective of treatment group), patients in the HP-hMG group had significantly fewer follicles, both in total and by various size distributions. The total number of follicles was on average 14.8 with HP-hMG and 15.9 for rFSH, which was significantly different (P = 0.013). The mean number of oocytes retrieved was significantly (P < 0.001) higher with rFSH (11.8) compared with HP-hMG (10.0).
Serum levels during stimulation
Mean serum levels at the time points evaluated after the start of gonadotrophins (i.e. day 6 of stimulation, last stimulation day and day of oocyte retrieval) are presented in Table III, and the relative difference between treatment groups is illustrated in Figure 1. Data are presented for the population of all patients exposed to HP-hMG or rFSH. It is relevant to note that the differences observed in endocrine profile on the last stimulation day and at oocyte retrieval between HP-hMG and rFSH were also observed among those patients who maintained the dose of 225 IU throughout the study.
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FSH, LH and hCG
The serum FSH concentration was at all time points significantly higher among patients treated with HP-hMG compared with those treated with rFSH. On average, the FSH concentration was 5% higher with HP-hMG than with rFSH on day 6, 12% higher on the last stimulation day and 23% higher at oocyte retrieval. There was no difference between treatment groups in LH concentrations at any time point. Most of the patients in the HP-hMG group had circulating levels of hCG >2 IU/l on day 6 and at the end of stimulation. hCG was not detectable in serum from patients treated with rFSH at these time points. After a single administration of 250 µg of rhCG for triggering final maturation, the mean hCG concentration at oocyte retrieval was 108 and 104 IU/l in the HP-hMG and rFSH groups, respectively, which was not significantly different.
Estradiol and progesterone
The estradiol concentration on day 6 was 20% higher in the rFSH group than in the HP-hMG group, which was significant. On the last stimulation day and at the day of oocyte retrieval, the estradiol concentration was significantly higher by 10 and 16%, respectively, with HP-hMG compared with rFSH. The significantly higher estradiol level found among patients in the HP-hMG group at the end of stimulation and at oocyte retrieval was maintained after adjusting for either the number of follicles (15%) or the number of oocytes retrieved (29%). Estradiol >3500 pg/ml (12.85 nmol/l) on the last stimulation day was measured in 35 patients (10%) in the HP-hMG group and in 30 patients (8%) in the rFSH group. There was no significant difference in progesterone concentration between treatment groups on day 6, whereas the progesterone concentration was 23% higher with rFSH on the last stimulation day and 31% higher at oocyte retrieval. These significantly higher progesterone levels with rFSH were also maintained after adjusting for ovarian response: 28% higher at the end of stimulation when adjusting for the number of follicles and 29% higher at oocyte retrieval when adjusting for the number of oocytes retrieved. Progesterone >4 nmol/l on the last stimulation day was measured in 41 patients (11%) in the HP-hMG group and 85 patients (23%) in the rFSH group.
Androgens
Concentrations of androstenedione, total testosterone and FAI were significantly higher among patients treated with HP-hMG compared with those treated with rFSH at all time points after the start of stimulation. Androstenedione was on average 10% higher on day 6, 24% higher on the last stimulation day and 25% higher at oocyte retrieval with HP-hMG compared with rFSH. Total testosterone was 17, 32 and 36% higher in the HP-hMG group on day 6, on the last stimulation day and at oocyte retrieval, respectively. FAI was 22% higher with HP-hMG than with rFSH on day 6, 33% higher on the last stimulation day and 28% higher at oocyte retrieval.
Estrogen and androgen interplay
In the HP-hMG group, a proportional relationship with estradiol was observed on day 6 and at the end of stimulation for testosterone and on day 6 for androstenedione, and a linear relationship was observed at the end of stimulation for androstenedione. In the rFSH group, a proportional relationship with estradiol was observed for testosterone on day 6 and at the end of stimulation, whereas it was linear for androstenedione at both assessment time points. The relationships between estradiol and total testosterone and estradiol and androstenedione were not confounded by the FSH concentration.
Correlation between mid-follicular hCG and outcome
The associations between hCG levels during stimulation and outcome were assessed. Table IV summarizes embryo quality parameters and ongoing pregnancy by hCG concentrations on day 6 after a fixed daily dose of 225 IU HP-hMG. The ongoing pregnancy rate was significantly positively correlated with serum hCG concentrations on day 6 of stimulation (P = 0.008, trend analysis). The day 6 hCG concentration in the HP-hMG group was also significantly positively correlated with the number of top-quality embryos on day 3 after oocyte retrieval (P = 0.002, trend analysis) and the percentage of patients with top-quality embryos (P = 0.003, trend analysis).
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Follicular fluid levels
Table V summarizes the geometrical means of follicular fluid hormone concentrations for each of the treatment groups, and Figure 2 illustrates the relative difference between HP-hMG and rFSH. The concentrations of FSH, LH and hCG in follicular fluid were significantly higher among patients treated with HP-hMG compared with those treated with rFSH. On average, FSH was 63% higher, LH was 56% higher and hCG was 23% higher with HP-hMG than with rFSH. The follicular fluid concentrations of estradiol and progesterone were significantly different between groups. Estradiol was 62% higher in the HP-hMG group than in the rFSH group, and progesterone was 20% higher with rFSH compared with HP-hMG. Total testosterone and FAI were significantly higher in follicular fluid in patients stimulated with HP-hMG compared with those with rFSH. Total testosterone was 39% higher with HP-hMG compared with rFSH, whereas there was no difference in SHBG, leading to an average FAI that was 41% higher for HP-hMG compared with rFSH.
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As shown in Figure 2, the ratios of estradiol : androstenedione, estradiol : total testosterone and estradiol : progesterone were all significantly higher in the follicular fluid from patients stimulated with HP-hMG compared with those stimulated with rFSH. Estradiol : androstenedione was 24% higher with HP-hMG, estradiol : total testosterone 16% higher and estradiol : progesterone 93% higher. Concerning the inhibins, the following significant observations were made: inhibin A was higher in the rFSH group by 11%, and inhibin B was higher in the HP-hMG group by 18%, with the inhibin A : inhibin B ratio being 31% higher in the rFSH group. The follicular concentration of VEGF was 27% higher among patients stimulated with rFSH, which was significant. There were no significant differences between treatment groups in follicular fluid concentrations of IGF-1, cortisol or cortisone or in cortisol : cortisone ratio.
The differences observed in follicular fluid profile between HP-hMG and rFSH were also observed among those patients who maintained the dose of 225 IU throughout the study, with the single exception of the estradiol : total testosterone ratio, which was not significantly different between HP-hMG and rFSH in this subset of patients.
Correlation between follicular fluid and serum levels
There was a good correlation between follicular fluid levels and circulating levels of FSH, LH, hCG and SHBG at the time of oocyte retrieval. The correlation coefficients (HP-hMG and rFSH, respectively) were 0.88 and 0.81 for FSH, 0.72 and 0.72 for hCG and 0.88 and 0.89 for SHBG. Ovarian steroids in follicular fluid were poorly correlated with the serum levels at oocyte retrieval, with correlation coefficients in the range of 0.18–0.31 for estradiol, 0.19–0.26 for progesterone, 0.10–0.14 for androstenedione and 0.07–0.31 for total testosterone. These correlations were not markedly improved when the serum levels were adjusted for the number of oocytes retrieved. Serum levels of LH at the time of oocyte retrieval were below the limit of quantification in 99% of the patients, and therefore, the correlation between serum and follicular fluid levels was not calculated.
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Discussion |
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The results indicate that providing LH activity in gonadotrophin treatment from the start of stimulation, after GnRH agonist down-regulation, induces significant differences in hormone profiles in both serum and follicular fluid accompanied by differential follicle growth and selection. It is reasonable to assume that findings in follicular development and endocrine environment during ovarian stimulation until oocyte retrieval are generalizable from IVF to ICSI cycles. Despite the administration of an identical starting dose of two preparations equally calibrated for FSH bioactivity in the Steelman–Pohley assay and with similar batch-to-batch consistency (Wolfenson et al., 2005
), different effects on follicle recruitment and on circulating FSH and estradiol concentrations were observed. The FSH in the HP-hMG preparation has a longer half-life than the FSH in the rFSH preparation, as the relative difference in FSH concentrations was higher 2 days after stopping gonadotrophin treatment (time of oocyte retrieval) than at the end of stimulation. The similar LH levels between the patients exposed to HP-hMG and those to rFSH are not surprising. The timing of sampling could contribute to this finding, as LH has a very short half-life. Moreover, the HP-hMG preparation used in this study has a low LH content, with most of the LH activity derived from the hCG rather than the LH content (Wolfenson et al., 2005). Exposure to hCG from the start of stimulation might explain the differential effects observed between gonadotrophin preparations in follicle growth and selection.
It has previously been reported from the population in this study that fewer oocytes were retrieved in the HP-hMG group but that the proportion of top-quality embryos was higher than in the rFSH group (Nyboe Andersen et al., 2006). The ovarian response differed between treatment groups early after the start of stimulation. On day 6, there were more follicles in the rFSH group contributing to the significantly higher estradiol concentration compared with HP-hMG. Circulating aromatizable androgen concentrations and FAI were increased with HP-hMG, and this higher serum androgen tonus is related to the presence of circulating hCG, as the two treatments were not different with respect to LH concentration. The provision of androgens in the human ovary can be attributed to the theca cells that express functional LH receptors already during the pre-antral follicle growth stages (Erickson et al., 1985). The pivotal role of androgens for follicle survival has been studied in several animal models and also in the human (Hillier and De Zwart, 1981; Horie et al., 1992; Tetsuka and Hillier, 1997). During follicular growth, aromatizable androgens are either transformed by aromatase into estrogens or bound to the androgen receptor present on granulosa cells. Cyp19 aromatase promotor use can be triggered differentially by androgen subtypes, leading to an increased estrogen synthesis (Shaw et al., 1989; Hamel et al., 2005). Estrogens act via estrogen receptors in granulosa cells, opposing the negative effects of androgens in the follicle (Cheng et al., 2002). Androgens can, via the androgen receptor, increase the sensitivity of the follicle to FSH via up-regulation of FSH receptor (Luo and Wiltbank, 2006). However, an amplification of the FSH-stimulated cAMP-mediated postreceptor signalling at an early stage of follicular development will induce the arrest of proliferation in granulosa cells (Hillier et al., 1991). A variety of paracrine and autocrine loops can contribute to the estrogen–androgen tonus that determines the fate of the follicle (Erickson et al., 1989; Hillier et al., 1991). It is hypothesized that the shift in favour of androgens by hCG in HP-hMG from the start of stimulation led to a more selective follicle growth.
It has been speculated that elevated LH levels during ovarian stimulation have a detrimental impact on pregnancy rates (Stanger and Yovich, 1985; Howles et al., 1986; Homburg et al., 1988). It is, however, important to distinguish between exogenous LH/hCG supplementation and elevated endogenous LH levels.
In HP-hMG-treated patients, the contribution of exogenous LH activity supplementation can be evaluated independently from the confounding factor of endogenous LH levels. Among the HP-hMG-treated patients, those with the highest day 6 hCG levels had the highest ongoing pregnancy rates. Also, the number of top-quality embryos and the proportion of patients with top-quality embryos in the HP-hMG group were positively correlated with circulating levels of hCG. Oocytes in the HP-hMG group were found to have a similar fertilization rate as oocytes in the rFSH group, but the proportion of top-quality embryos was higher with HP-hMG (Nyboe Andersen et al., 2006). Improved embryo quality with exogenous LH has recently been reported from another study, where a higher incidence of grade 1 and 2 embryos was observed when supplementing the FSH stimulation with LH in women undergoing a long agonist protocol (Lisi et al., 2005). In the context of most recent studies involving LH receptor action on human cumulus cells (Assou et al., 2006), the direct effects of LH/hCG on this important cellular compartment linked to the maturing oocyte and subsequent embryo quality (McKenzie et al., 2004) could become part of the working hypothesis.
It was interesting to observe that progesterone was more elevated with rFSH-alone treatment. Several mechanisms could explain the increased progesterone that was also found in twice as many patients in the rFSH group compared with the HP-hMG group (23 versus 11%). Although increases in progesterone have been historically associated with LH in the context of premature LH surges, recent articles attribute the increase in progesterone to FSH exposure (Hofmann et al., 1993; Ubaldi et al., 1996; Filicori et al., 2002). Patients in the HP-hMG group had longer exposure time, higher serum FSH and presence of hCG, but this did not lead to increased progesterone compared with the rFSH group. The fact that there were more follicles with rFSH cannot entirely explain the increased progesterone tonus observed with this preparation, as progesterone remained significantly different between treatment groups even when adjusting for the number of developed follicles. Changes in the paracrine regulation could explain differences in progesterone production in the follicle. FSH-stimulated granulosa cells produce paracrine factors that either stimulate the production of progesterone and androgens such as IGF-1 and inhibins (Hillier et al., 1991; Smyth et al., 1993; Campbell and Baird, 2001) or decrease the action of P450c17
enzymes such as transforming growth factor (TGF)- (Magoffin et al., 1995). TGF-, whose intrafollicular production is normally inhibited by LH, acts as a strong direct suppressor of the enzyme converting progesterone to androgens (Hernandez et al., 1990; Fournet et al., 1996) and as a stimulator of the steroidogenic genes [P450scc and 3-hydroxysteroid dehydrogenase (HSD)] necessary for progesterone production (Fournet et al., 1996). TGF- has been found to be increased in follicular fluid from GnRH agonist-down-regulated patients stimulated with rFSH compared with patients stimulated with a menotrophin preparation leading to an increased progesterone accumulation (Fried et al., 1998). Although it is generally assumed that there is sufficient remaining active LH present after GnRH agonist down-regulation to support steroid production during the initial phase of follicle selection (reviewed by Fauser, 1997), this study found that supplementary hCG activity increases estrogenization of the follicle without increasing the luteinization process. The higher progesterone tonus has been shown to have a biological correlate at the endometrial echogenicity level with hyperechogenicity, leading to a reduced ongoing pregnancy rate (Nyboe Andersen et al., 2006). The propensity of rFSH to induce early luteinization and the impact on endometrial transformation would obviously be equally harmful in IVF as in ICSI cycles.
Regarding intrafollicular assessments, FSH was significantly higher in HP-hMG-stimulated patients, in line with the observations in serum FSH. LH concentrations were also significantly higher with HP-hMG, although the levels were just above the functional sensitivity of the immunoassay. Intrafollicular hCG was also significantly higher for HP-hMG, but the contribution of intrafollicular hCG during stimulation cannot be known from these measurements, as samples were taken after hCG injection for triggering ovulation massively influencing the intrafollicular hCG levels after the end of stimulation. The higher production of intrafollicular androgens and estrogens in the HP-hMG group is attributed to hCG. There was no correlation between serum estradiol, androstenedione or testosterone and follicular fluid levels for these hormones, nor when serum steroid values were adjusted by the number of oocytes retrieved. The simultaneous increase in estradiol and androgens did not induce a different adaptation in SHBG production between the two treatment groups. The calculated intrafollicular estrogen : androgen ratios demonstrate that HP-hMG induced a more estrogenic than androgenic microenvironment. These data suggest that paracrine factors induced by hCG, and mediated by androgens, activate aromatase, leading towards a more estrogenic intrafollicular climate by the end of stimulation. The estradiol : total testosterone values have been found to be significantly higher in follicular fluid from pregnancy-associated follicles than in fluid containing oocytes that did not produce viable embryos (Yding Andersen and Ziebe, 1992; Yding Andersen, 1993). The potential to effectively convert androgens to estradiol may be considered a marker of health of the follicle, and an inability to convert androgens to estradiol may represent an early atretic change.
Intrafollicular progesterone concentrations measured after hCG administration reflect the massive response of LH receptor-expressing granulosa cells. The hCG ovulatory dose induces a partial inhibition of 17-hydroxylase in theca cells; as a result, more progesterone and less androgens are produced. Immediately following the hCG injection, the pre-ovulatory granulosa cells will have a decreased production of estradiol because of a lack of androgen provision and a secondary decrease in aromatase, resulting in a change from estradiol to progesterone synthesis (Erickson et al., 1985; Couse et al., 2005). In this study, progesterone concentrations in follicular fluid were higher in rFSH-stimulated patients. Similar observations have been made previously (Westergaard et al., 2004). The estradiol : progesterone ratio in follicular fluid was significantly higher with HP-hMG compared with rFSH. It is hypothesized that providing continuous levels of hCG throughout stimulation (as is the case of HP-hMG) causes desensitization to major signalling (Hausdorff et al., 1990). An exaggerated shift in the steroid balance from androstenedione and estradiol to progesterone after the administration of hCG as ovulatory trigger has been associated with a negative effect on early embryo development (Dumesic et al., 2002, 2003). It can be speculated that hCG exposure throughout stimulation would dampen the effect of the hCG as ovulatory stimulus and would reduce the magnitude of the androgen and estrogen shift to progesterone, described to be detrimental to early embryogenesis.
The rapid growth of multiple follicles needs the development of a vascular network. VEGF is a major mediator of neovascularization and permeabilization. Its production by granulosa cells is influenced by gonadotrophins and steroids and is highly increased during luteinization (Christenson and Stouffer, 1997). The intrafollicular VEGF concentration in rFSH compared with HP-hMG might be compatible with an earlier luteinization. Inhibin A and B production in antral follicles is highly dependent on gonadotrophin stimulation and on the stage of granulosa differentiation. Inhibin B secretion by small follicles reflects the increase in the number of granulosa cells and inhibin A secretion their increased differentiation (Campbell and Baird, 2001). Assuming that inhibin A : inhibin B is a good marker for the follicle developmental progression, the data suggest that large follicles in the rFSH group might be more advanced towards end-differentiation. Considering the increase in progesterone, inhibin A : inhibin B ratio and VEGF in the rFSH group, it could be suggested that this treatment leads granulosa cells more rapidly to their end-differentiation stage or pre-luteinization.
In conclusion, the hormonal data from this comparative study show that the presence of LH activity in gonadotrophin preparations has a differential effect on the antral follicle selection process. LH activity supplementation induces a more selective follicle recruitment process, presumably because of the significant shift in the balance between estrogens and androgens. The distinct endocrine profiles between HP-hMG and rFSH appear to have genuine biological effects at the end organ level, as reflected in the embryo quality and endometrial changes reported for the population included in this trial (Nyboe Andersen et al., 2006). The intrafollicular endocrine response upon the ovulatory hCG injection supports the hypothesis that folliculogenesis in the absence of sufficient amounts of LH activity leads the granulosa cells more quickly to luteinization.
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TopAbstractIntroductionMaterials and methodsResultsDiscussionConflict of interestAcknowledgementsReferences |
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Johan Smitz and Paul Devroey have conducted clinical research sponsored by Ferring Pharmaceuticals, Serono and Organon. Anders Nyboe Andersen has conducted clinical research sponsored by Ferring Pharmaceuticals, Serono, Organon, Novo Nordisk and Medicult. Joan-Carles Arce is employee of Ferring Pharmaceuticals.
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TopAbstractIntroductionMaterials and methodsResultsDiscussionConflict of interestAcknowledgementsReferences |
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The authors thank Per Sørensen, MSc, Biometrics, Ferring Pharmaceuticals A/S for conducting the statistical analyses. The study was sponsored by Ferring Pharmaceuticals A/S, Copenhagen, Denmark. We also thank all participating centres. Belgium: AZ-VUB, Jette; Private Practice, Aalter; Centre Hospitalier Notre Dame, Charleroi; Universitair Ziekenhuis, Antwerpen; Virga Jesse Ziekenhuis, Hasselt; Universitair Ziekenhuis, Gent; Hôpital Erasme, Brussels; UZ Gasthuisberg, Leuven. Czech Republic: Veobecná Fakultní Nemocnice, Prague; ISCARE IVF a.s., Prague; Sanatorium Protnatal, Prague. Denmark: Rigshospitalet, Copenhagen; Herlev Amtssygehus, Herlev; Odense Universitetshospital, Odense; Hvidovre Hospital, Hvidovre. Finland: Väestölitto Lapsettomuusklinikka, Helsinki; Felicitas-Klinikka, Helsinki; Väestölitto, Oulu. France: Hobital Mère Enfant, Nantes; Clinique Saint Antoine, Bois Guillaume; Hobital Edoard Herriot, Lyon. Israel: Lis Maternity Hospital, Tel-Aviv; Bnai-Zion Medical Centre, Haifa; Rabin Medical Centre, Petach-Tikva; Assaf Harofe Medical Centre, Beer Yaacov; Sherba MC, Tel-Hashomer. Poland: Centrum Leczenia Nieplodnosci, Bialystok; Akademia Medyczna, Poznan; Klinika Ginekologii Akademii, Bialystok. Slovenia: General Hospital, Maribor; University Medical Centre, Ljubljana. Spain: IVI, Madrid; Institut Universitary Dexeus, Barcelona; Hospital Maternal, Valencia. Sweden: Sahlgrenska University Hospital, Gothenburg; Carl von Linné Kliniken, Uppsala; IVF Öresund, Malmö. These data were presented in part at the Annual ESHRE Meeting, Copenhagen, Denmark, 2005.
Funding to pay the Open Access publication charges for this article was provided by Ferring Pharmaceuticals A/S.
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* Menotrophin versus Recombinant FSH in vitro Fertilisation Group
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Arce J-C, Nyboe Andersen A, Collins J. (2005) Resolving methodological and clinical issues in the design of efficacy trials in assisted reproductive technologies: a mini-review. Hum Reprod 20:1757–1771.
Assou S, Anahory T, Pantesco V, Carrour TL, Pellestor F, Klein B, Reyftmann L, Dechaud H, Vos JD, Hamamah S. (2006) The human cumulus-oocyte complex gene-expression profile. Hum Reprod 21:1705–1719.
Campbell BK and Baird DT. (2001) Inhibin A is a follicle stimulating hormone-responsive marker of granulosa cell differentiation, which has both autocrine and paracrine actions in sheep. J Endocrinol 169:333–345.[Abstract]
Cheng G, Weihua Z, Mäkinen S, Mäkelä S, Saji S, Warner M, Gustafsson J, Hovatta O. (2002) A role for the androgen receptor in follicular atresia of estrogen receptor beta knockout mouse ovary. Biol Reprod 66:77–84.
Christenson LK and Stouffer RL. (1997) Follicle-stimulating hormone and luteinizing hormone/chorionic gonadotrophin stimulation of vascular endothelial growth factor production by macaque granulosa cells from pre- and periovulatory follicles. J Clin Endocrinol Metab 82:2135–2142.
Collins J. (2003) A turbulent arena. Fertil Steril 80:1117–1120.[CrossRef][Web of Science][Medline]
Costagliola S, Niccoli P, Florentino M, Carayon P. (1994) European collaborative study on luteinizing hormone assay: 2. Discrepancy among assay kits is related to variation both in standard curve calibration and epitope specificity of kit monoclonal antibodies. J Endocrinol Invest 17:407–416.[Web of Science][Medline]
Couse JF, Yates MM, Deroo BJ, Korach KS. (2005) Estrogen receptor- is critical to granulosa cell differentiation and the ovulatory response to gonadotrophins. Endocrinology 146:3247–3262.[CrossRef][Web of Science][Medline]
Dumesic DA, Schramm RD, Peterson E, Paprocki AM, Zhou R, Abbott DH. (2002) Impaired developmental competence of oocytes in adult prenatally androgenized female rhesus monkeys undergoing gonadotrophin stimulation for in vitro fertilisation. J Clin Endocrinol Metab 87:1111–1119.
Dumesic DA, Schramm RD, Bird IM, Peterson E, Paprocki AM, Zhou R, Abbott DH. (2003) Reduced intrafollicular androstenedione and estradiol levels in early-treated prenatally androgenized female rhesus monkeys receiving follicle-stimulating hormone therapy for in vitro fertilization. Biol Reprod 69:1213–1219.
Erickson GF, Magoffin DA, Dyer CA, Hofeditz C. (1985) The ovarian androgen producing cells: a review of structure/function relationships. Endocr Rev 6:371–399.
Erickson GF, Garzo VG, Magoffin DA. (1989) Insulin-like growth factor-I regulates aromatase activity in human granulosa and granulosa luteal cells. J Clin Endocrinol Metab 69:716–724.
Esposito MA, Barnhart KT, Coutifaris C, Patrizio P. (2001) Role of periovulatory luteinizing hormone concentrations during assisted reproductive technology cycles stimulated exclusively with recombinant follicle-stimulating hormone. Fertil Steril 75:519–524.[CrossRef][Web of Science][Medline]
The European Recombinant Human LH Study Group. (1998) Recombinant human luteinizing hormone (LH) to support recombinant human follicle-stimulating hormone (FSH)-induced follicular development in LH- and FSH deficient anovulatory women: a dose-finding study. J Clin Endocrinol Metab 83:1507–1514.
Fauser BC. (1997) Follicular development and oocyte maturation in hypogonadotrophic women employing recombinant follicle-stimulating hormone: the role of oestradiol. Hum Reprod Update 3:101–108 (Review).
Filicori M, Cognigni GE, Taraborrelli S, Spettoli D, Ciampaglia W, de Fatis CT, Pocognoli P. (1999) Luteinizing hormone activity supplementation enhances follicle-stimulating hormone efficacy and improves ovulation induction outcome. J Clin Endocrinol Metab 84:2659–2663.
Filicori M, Cognigni GE, Taraborrelli S, Spettoli D, Ciampaglia W, de Fatis CT, Pocognoli P, Cantelli B, Boschi S. (2001) Luteinizing hormone activity in menotrophins optimizes folliculogenesis and treatment in controlled ovarian stimulation. J Clin Endocrinol Metab 86:337–343.
Filicori M, Cognigni GE, Pocognoli P, Tabarelli C, Spettoli D, Taraborrelli S, Ciampaglia W. (2002) Modulation of folliculogenesis and steroidogenesis in women by graded menotropin administration. Hum Reprod 17:2009–2015.
Fleming R, Rehka P, Deshpande N, Jamieson ME, Yates RWS, Lyall H. (2000) Suppression of LH during ovarian stimulation: effects differ in cycles stimulated with purified urinary FSH and recombinant FSH. Hum Reprod 15:1440–1445.
Fournet N, Weitsman SR, Zachow RJ, Magoffin DA. (1996) Transforming growth factor- inhibits ovarian 17 -hydroxylase activity by a direct noncompetitive mechanism. Endocrinology 137:166–174.[Abstract]
Fried G, Wramsby H, Tally M. (1998) Transforming growth factor-1, insulin-like growth factors, and insulin-like growth factor binding proteins in ovarian follicular fluid are differentially regulated by the type of ovarian hyperstimulation used for in vitro fertilization. Fertil Steril 70:129–134.[CrossRef][Web of Science][Medline]
Hamel M, Vanselow J, Nicola ES, Price CA. (2005) Androstenedione increases cytochrome P450 aromatase messenger ribonucleic acid transcripts in nonluteinizing bovine granulosa cells. Mol Reprod Dev 70:175–183.[CrossRef][Web of Science][Medline]
Hausdorff WP, Caron MG, Lefkowitz RJ. (1990) Turning off the signal: desensitization of -adrenergic receptor function. FASEB J 4:2881–2889.[Abstract]
Hernandez ER, Hurwitz A, Payne DW, Dharmarajan AM, Purchio AF, Adashi EY. (1990) Transforming growth factor-1 inhibits ovarian androgen production: gene expression, cellular localization, mechanism(s), and site(s) of action. Endocrinology 127:2804–2811.
Hillier SG. (1994) Current concepts of the roles of follicle stimulating hormone and luteinizing hormone in folliculogenesis. Hum Reprod 9:188–191.
Hillier SG and De Zwart FA. (1981) Evidence that granulosa cell aromatase induction/activation by follicle-stimulating hormone is an androgen receptor-regulated process in-vitro. Endocrinology 109:1303–1305.
Hillier SG, Whitelaw PF, Smyth CD. (1994) Follicular oestrogen synthesis: the ‘two-cell, two-gonadotrophin’ model revisited. Mol Cell Endocrinol 10:51–54.
Hillier SG, Yong EL, Illingworth PJ, Baird DT, Schwall RH, Mason AJ. (1991) Effect of recombinant inhibin on androgen synthesis in cultured human thecal cells. Mol Cell Endocrinol 75:R1–R6.[CrossRef][Web of Science][Medline]
Hofmann GE, Bergh PA, Guzman I, Masuku S, Navot D. (1993) Premature luteinization in not eliminated by pituitary desensitization with leuprolide acetate in women undergoing gonadotrophin stimulation who demonstrated premature luteinization in a prior gonadotrophin-only cycle. Hum Reprod 8:695–698.
Homburg R, Armar NA, Eshel A, Adams J, Jacobs HS. (1988) Influence of serum luteinising hormone concentrations on ovulation, conception, and early pregnancy loss in polycystic ovary syndrome. BMJ 297:1024–1026.
Horie K, Takakura K, Fujiwara H, Suginami H, Liao S, Mori T. (1992) Immunohistochemical localization of androgen receptor in the human ovary throughout the menstrual cycle in relation to oestrogen and progesterone receptor expression. Hum Reprod 7:184–190.
Howles CM, Macnamee MC, Edwards RG, Goswamy R, Steptoe PC. (1986) Effect of high tonic levels of luteinizing hormone on outcome of in vitro fertilisation. Lancet 2:521–522.[CrossRef][Web of Science][Medline]
Humaidan P. (2006) To add or not to add LH: comments on a recent commentary. Reprod Biomed Online 12:284–285.[Web of Science][Medline]
Humaidan P, Bungum L, Bungum M, Yding Andersen C. (2002) Ovarian response and pregnancy outcome related to mid-follicular LH levels in women undergoing assisted reproduction with GnRH agonist down-regulation and recombinant FSH stimulation. Hum Reprod 17:2016–2021.
Humaidan P, Bungum M, Bungum L, Yding Andersen C. (2004) Effects of recombinant LH supplementation in women undergoing assisted reproduction with GnRH agonist down-regulation and stimulation with recombinant FSH: an opening study. Reprod Biomed Online 8:635–643.[Web of Science][Medline]
Lisi F, Rinaldi L, Fishel S, Caserta D, Lisi R, Campbell A. (2005) Evaluation of two doses of recombinant luteinizing hormone supplementation in an unselected group of women undergoing follicular stimulation for in vitro fertilization. Fertil Steril 83:309–315.[CrossRef][Web of Science][Medline]
Loumaye E, Engrand P, Shoham Z, Hillier SG, Baird DT. on behalf of the Recombinant LH and Study Group. (2003) Clinical evidence for a LH ‘ceiling’ effect induced by administration of recombinant human LH during the late follicular phase of stimulated cycles in World Health Organization type I and type II anovulation. Hum Reprod 18:314–322.
Luo W and Wiltbank MC. (2006) Distinct regulation by steroids of messenger RNAs for FSHR and CYP19A1 in bovine granulosa cells. Biol Reprod 75:217–225.
Magoffin DA, Hubert-Leslie D, Zachow RJ. (1995) Estradiol-17, insulin-like growth factor-I, and luteinizing hormone inhibit secretion of transforming growth factor by rat ovarian theca-interstitial cells. Biol Reprod 53:627–635.[Abstract]
McKenzie LJ, Pangas SA, Carson SA, Kovanci E, Cisneros P, Buster JE, Amato P, Matzuk MM. (2004) Human cumulus granulosa cell gene expression: a predictor of fertilization and embryo selection in women undergoing IVF. Hum Reprod 19:2869–2874.
McNatty KP, Hunter WM, McNeilly AS, Sawers RS. (1975) Changes in the concentration of pituitary and steroid hormones in the follicular fluid of human graafian follicles throughout the menstrual cycle. J Endocrinol 64:555–571.
Niccoli P, Costagliola S, Patricot MC, Mallet B, Benahmed M, Carayon P. (1996) European collaborative study of LH assay: 3. relationship of immunological reactivity, biological activity and charge of human luteinizing hormone. J Endocrinol Invest 19:260–267.[Web of Science][Medline]
Nyboe Andersen A, Devroey P, Arce J-C. for the MERIT (Menotrophin vs Recombinant FSH in vitro Fertilisation Trial) Group. (2006) Clinical outcome following stimulation with HP-hMG or recombinant FSH in patients undergoing IVF: a randomized assessor-blind controlled trial. Hum Reprod Advance Access published on July 27.
Platteau P, Smitz J, Albano C, Sørensen P, Arce J-C, Devroey P. (2004) Exogenous luteinizing hormone activity may influence the treatment outcome in in vitro fertilization but not in intracytoplasmic sperm injection cycles. Fertil Steril 81:1401–1404.[CrossRef][Web of Science][Medline]
Platteau P, Nyboe Andersen A, Balen A, Devroey P, Sørensen P, Helmgaard L, Arce J-C. (2006) Similar ovulation rates, but different follicular development with highly purified menotrophin compared with recombinant FSH in WHO Group II anovulatory infertility: a randomized controlled study. Hum Reprod 21:1798–1804.
van Santbrink EJ, Hop WC, van Dessel TJ, de Jong FH, Fauser BC. (1995) Decremental follicle-stimulating hormone and dominant follicle development during the normal menstrual cycle. Fertil Steril 64:37–43.[Web of Science][Medline]
Shaw HJ, Hillier SG, Hodges JK. (1989) Developmental changes in luteinizing hormone/human chorionic gonadotrophin steroidogenic responsiveness in marmoset granulosa cells: effects of follicle-stimulating hormone and androgens. Endocrinology 124:1669–1677.
Smyth CD, Miró F, Whitelaw PF, Howles CM, Hillier SG. (1993) Ovarian thecal/interstitial androgen synthesis is enhanced by a follicle-stimulating hormone-stimulated paracrine mechanism. Endocrinology 133:1532–1538.
Stanger JD and Yovich JL. (1985) Reduced in-vitro fertilization of human oocytes from patients with raised basal luteinizing hormone levels during the follicular phase. Br J Obstet Gynaecol 92:385–393.[Web of Science][Medline]
Sullivan M, Stewart-Akers A, Krasnow JS, Berga SL, Zeleznik AJ. (1999) Ovarian responses in women to recombinant follicle-stimulating hormone and luteinising hormone (LH): a role for LH in the final stages of follicular maturation. J Clin Endocrinol Metab 84:228–232.
Tetsuka M and Hillier SG. (1997) Differential regulation of aromatase and androgen receptor in granulosa cells. J Steroid Biochem Mol Biol 61:233–239.[CrossRef][Web of Science][Medline]
Ubaldi F, Camus M, Smitz J, Bennink HJTC, Van Steirteghem A, Devroey P. (1996) Premature luteinization in in vitro fertilization cycles using gonadotrophin-releasing hormone agonist (GnRH-a) and recombinant follicle-stimulating hormone (FSH) and GnRH-a and urinary FSH. Fertil Steril 66:275–280.[Web of Science][Medline]
Watson H, Kiddy DS, Hamilton-Fairley D, Scanlon MJ, Barnard C, Collins WP, Bonney RC, Franks S. (1993) Hypersecretion of luteinizing hormone and ovarian steroids in women with recurrent early miscarriage. Hum Reprod 8:829–833.
Westergaard LG, Laursen SB, Yding Andersen C. (2000) Increased risk of early pregnancy loss by profound suppression of luteinizing hormone during ovarian stimulation in normogonadotrophic women undergoing assisted reproduction. Hum Reprod 15:1003–1008.
Westergaard LG, Erb K, Laursen SB, Rasmussen PE, Rex S, Westergaard CG, Yding Andersen C. (2004) Concentrations of gonadotrophins and steroids in pre-ovulatory follicular fluid and serum in relation to stimulation protocol and outcome of assisted reproduction treatment. Reprod Biomed Online 8:516–523.[Web of Science][Medline]
Wolfenson C, Groisman J, Couto AS, Hedenfalk M, Cortvrindt RG, Smitz JEJ, Jespersen S. (2005) Batch-to-batch consistency of human-derived gonadotrophin preparations compared with recombinant preparations. Reprod Biomed Online 10:442–454.[Web of Science][Medline]
Yding Andersen C. (1993) Characteristics of human follicular fluid associated with successful conception after in vitro fertilization. J Clin Endocrinol Metab 77:1227–1234.[Abstract]
Yding Andersen C and Ziebe S. (1992) Serum levels of free androstenedione, testosterone and oestradiol are lower in the follicular phase of conceptional than of non-conceptional cycles after ovarian stimulation with a gonadotrophin-releasing hormone agonist protocol. Hum Reprod 7:1365–1370.
Zeleznik AJ and Hillier SG. (1984) The role of gonadotropins in the selection of the preovulatory follicle. Clin Obstet Gynecol 27:927–940.[CrossRef][Web of Science][Medline]
Submitted on May 29, 2006; resubmitted on September 7, 2006; resubmitted on October 10, 2006; accepted on October 19, 2006.
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