Top quality blastocyst formation rates in relation to progesterone levels on the day of oocyte maturation in GnRH antagonist IVF/ICSI cycles.

Cycles with progesterone elevation during controlled ovarian stimulation (COS) for IVF/ICSI are commonly managed with a "freeze-all" strategy, due to a well-recognized detrimental effect of high progesterone levels on endometrial receptivity. However, also a detrimental effect of elevated progesterone on day-3 embryo quality has recently been found with regards to top quality embryo formation rate. Because blastocyst culture and cryopreservation are largely adopted, we deemed relevant to determine whether this detrimental effect is also seen on blastocyst quality on day 5-6. This issue was investigated through a large two-center retrospective study including 986 GnRH antagonist IVF/ICSI cycles and using top quality blastocyst formation rate as the main outcome. Results showed that on multivariate analysis sperm motility (p<0.01) and progesterone levels at ovulation triggering (p = 0.01) were the only two variables that significantly predicted top quality blastocyst formation rate after adjusting for relevant factors including female age, BMI, basal AMH and total dose of FSH used for COS. More specifically, progesterone levels at induction showed an inverse relation with top quality blastocyst formation (correlation coefficient B = -1.08, 95% CI -1.9 to -0.02) and ROC curve analysis identified P level >1.49 ng/ml as the best cut-off for identification of patients at risk for the absence of top quality blastocysts (AUC 0.55, p<0.01). Our study is the first to investigate the top quality blastocyst formation rate in relation to progesterone levels in IVF/ICSI cycles, showing that increasing progesterone is associated with lower rates of top quality blastocyst. Hence, the advantages of prolonging COS to maximize the number of collected oocytes might eventually be hindered by a decrease in top quality blastocysts available for transfer, if increasing progesterone levels are observed. This observation extends the results of two recent studies focused on day-3 embryos and deserves further research.

A practical guide to the diagnosis and management of amenorrhoea.

For women of reproductive age, pregnancy and lactation are the two most common physiological causes of amenorrhoea. This article concentrates on pathological causes of amenorrhoea. Primary amenorrhoea refers to the absence of menarche at the age of 16 and secondary amenorrhoea is the cessation of menses for at least 6 months in already cycling women. Amenorrhoea is not a diagnosis but a symptom indicating anatomical, genetic and neuroendocrine abnormalities. It can be determined by two different groups of causes: (a) anatomical defects of the genital organs; (b) endocrine dysfunctions. Both congenital and acquired anomalies in the structure of the uterus and vagina could produce amenorrhoea; nevertheless, in the vast majority of patients, amenorrhoea is related to an ovarian malfunction. Diagnostic work-up includes history, physical examination, laboratory data and imaging. Amenorrhoea resulting from ovarian malfunction is associated with 4 distinct endocrine conditions. Hyperprolactinaemic amenorrhoea is often associated with a pituitary adenoma. Prolactin-lowering drugs, cyclical progestogen and hormone replacement therapy (HRT) are the different choices of treatment for cycle disturbance; a contraceptive pill can be used to ensure contraception, while prolactin-lowering drugs induce fertility in patients who desire pregnancy. Hypogonadotrophic amenorrhoea is frequently associated with stress and nutritional deficiency. If this is the case the patient should simply be counselled. A sequential use of estrogen and progestogen can be suggested to prevent estrogen deficiency or for psychological reasons. If contraception is needed, oral contraception may be the choice for both cycle and fertility control. If the patient desires pregnancy, ovulation may be induced with pulsatile gonadotrophin-releasing hormone (GnRH) in patients with hypothalamic disfunction and with gonadotrophins in patients with pituitary failure. Hypergonadotrophic amenorrhoea is the result of an ovarian failure. There is no curative therapy for these amenorrhoeas. However, a long term hypoestrogenic condition should be treated with estrogen to cure symptoms and to prevent an increased risk of cardiovascular disease and osteoporosis. Normogonadotrophic amenorrhoea is caused by some disturbance in the pattern of pulsatile GnRH secretion. Since these women have some ovarian activity, they are not hypoestrogenic and will bleed in response to progestogen withdrawal. Most of these patients are likely to have polycystic ovarian disease (PCO). Menstrual bleeding can be induced in these women by cyclical progestogen administration or the sequential use of estrogen plus progestogen. Oral contraception is indicated not only in patients who desire to be protected against pregnancy but also in women with acne and hirsutism. These frequently present signs of hyperandrogenism are consistently improved by the ovarian suppression induced by the contraceptive pill. The beneficial effect of the pill can be reinforced by the simultaneous use of antiandrogens. Women with normogonadotrophic amenorrhoea and desiring pregnancy have a less favourable response to all forms of ovulation induction (antiestrogen, GnRH and gonadotrophin preparations.

PIP: Amenorrhea, characterized by anatomical, genetic, and neuroendocrine abnormalities, occurs frequently in adolescents and in 10-20% of women presenting with subfertility. It can be caused by anatomical defects of the genital organs or endocrine dysfunctions. A correct diagnosis, reached through history, physical examination, laboratory data, and imaging, is essential to proper treatment. Amenorrhea resulting from ovarian malfunction, the most common presentation, is associated with four distinct endocrine conditions. Hyperprolactinemic amenorrhea, often linked to a pituitary adenoma, is treated with prolactin-lowering drugs or cyclical progestogen and hormone replacement therapy. Hypogonadotrophic amenorrhea, frequently associated with stress and nutritional deficiency, is generally addressed through counseling and sequential use of estrogen and progestogen. Hypergonadotrophic amenorrhea, the result of ovarian failure, has no curative therapy; however, long-term hypoestrogenicity should be treated with estrogen to prevent an increased risk of osteoporosis and cardiovascular disease and cure symptoms. Finally, in normogonadotrophic amenorrhea, caused by a disturbance in the pattern of pulsatile gonadotropin-releasing hormone, menstrual bleeding can be induced by cyclical progestogen administration or the sequential use of estrogen plus progestogen.

Premature ovarian failure.

Secondary amenorrhoea with elevated gonadotrophins occurring under the age of 40 (premature ovarian failure (POF)), and at the age between 41 and 44 years (early menopause (EM)), respectively, affects 1-2% and 5% of women in the general population. Objective of this study was to evaluate the prevalence of familial cases of POF and EM and to assess the clinical and genetic characteristics of these patients. One hundred and sixty women with idiopathic secondary amenorrhoea before the age of 45 and serum follicle-stimulating hormone (FSH) levels greater than or equal to 40 IU/l were included in the study. Tests performed on patients included complete medical history, pedigree's analysis, clinical pelvic examination, gonadotrophins and thyroid assessment, chromosomal analysis. The 160 patients included in the study showed idiopathic POF (n=130) or EM (n=30). Following pedigree assessment, we were able to identify an incidence of familial cases of 28.5% in the POF group (n=37) and of 50% in the EM group (n=15). POF and EM condition were often present in the same family. There were no differences between POF and EM patients and between familial and sporadic cases regarding age at menarche, personal history, gynaecological history, weight, height and diet habits. There was a statistically significant difference between sporadic and familial cases in age at POF onset: 32.0+/-7.3 years (12-40) compared to 35. 0+/-5.8 (18-40), respectively (P<0.05). The POF and EM families identified showed two or more affected females and transmission through either maternal or paternal relatives; in four families both maternal and paternal transmission was observed. This study suggests that idiopathic POF and EM conditions, differing only in age of menopause onset, may represent a variable expression of the same genetic disease. The different age of menopause onset in these patients may be explained by genetic heterogeneity and/or by different environmental factors. Our results indicate a high rate of familial transmission of the condition. Pedigree's analysis suggests an autosomal or an X-linked dominant sex-limited pattern of inheritance for POF and EM.

Reliability of ovulation tests in infertile women.

OBJECTIVE: To assess the reliability of the most widely used clinical methods for predicting or confirming ovulation. METHODS: We monitored spontaneous cycles in 101 infertile women using basal body temperature (BBT), transvaginal ultrasound, a urinary stick system for LH surge, and three serum progesterone measurements in the midluteal phase. Transvaginal ultrasound monitoring was standard for ovulation detection and sensitivity. We calculated specificity and accuracy of each method compared with that standard. RESULTS: Follicular development and ultrasound evidence of ovulation were confirmed in 97 of 101 cycles (96%). Urinary LH surge preceded follicular rupture assessed by ultrasonography in all cycles and showed concordance with ultrasound-evidenced ovulation in 98 of 101 cases. The timing of BBT nadir had wide variability, and BBT and ultrasonography agreed in a similar percentage of cases (74%). Midluteal serum progesterone assessments showed ovulatory values in 93 subjects, and ovulation was concordant with ultrasonography in 90 subjects. CONCLUSION: Urinary LH was accurate in predicting ovulation with ultrasonography as the standard for detection, but time varied widely. The nadir of BBT predicted ovulation poorly. The BBT chart was less accurate for confirming ovulation, whereas a single serum progesterone assessment in midluteal phase seemed as effective as repeated serum progesterone measures.

Ovarian activity during regular oral contraceptive use.

The aim of this study was to assess whether during regular OC use ovarian activity might lead to ovulation, as assessed by ultrasound (US) evaluation of follicular growth and blood levels of 17-beta-estradiol and progesterone. A total of 51 healthy women with normal menstrual cycles (28 +/- 3 days) and no gynecological symptoms were recruited. A total of 22 patients were given a triphasic OC pill containing 35 mg ethinyl estradiol (EE) and 50 mg desogestrel (DSG) in the first seven tablets; 30 mg EE and 100 mg DSG in tablets 8 to 14, and 30 mg EE and 150 mg DSG in tablets 15 to 21; 29 patients received one of two OC pills, both containing 20 mg EE plus 150 mg DSG (15 patients) or 75 mg of gestodene (14 patients). A total of 86 cycles were monitored: 51 during the 3rd-4th cycle and 35 during the 6th-8th cycle of OC treatment. Follicular-like structures were observed in nine patients. The frequency of follicular-like structures was similar during the 3rd-4th cycle (9%) and during the 6th-8th cycle (11%). There was no relationship between follicular growth and blood levels of E2 and progesterone, which always appeared suppressed. In conclusion, the results of this study suggest that during OC use (even with low dose of ethinyl estradiol), a little ovarian activity may be present without ovulation.

PIP: Several studies have described a 10-20% incidence of folliculogenesis during low-dose oral contraceptive (OC) use. This study used ultrasound evaluation of follicular growth and blood levels of 17-beta-estradiol and progesterone to determine whether OC-induced ovarian activity led to ovulation in 51 healthy women with regular menstrual cycles. Subjects were given a new triphasic OC containing 35 mg ethinyl estradiol (EE) and 50 mg desogestrel (DSG) in the first 7 tablets, 30 mg EE and 100 mg DSG in tablets 8-14, and 30 mg EE and 150 mg DSG in tablets 15-21 (22 women); an OC containing 20 mg EE and 150 mg DSG (15 women); or an OC comprised of 20 mg EE and 75 mg gestodene (14 women). A total of 86 cycles were monitored: 51 during the third and fourth cycles and 35 during the sixth through eighth cycles of treatment. Follicular-like structures were observed in 9 patients. Follicular growth occurred in 4 cycles (11%) in the triphasic group, 3 (13%) in the EE-DSG group, and 2 (7%) in the EE-gestodene group. The frequency of follicular-like structures was similar during the third-fourth cycle (9%) and the sixth-eighth cycle (11%). There was no association between follicular growth and blood levels of estradiol and progesterone. These results are compromised by the fact that only 35 of the 51 subjects completed 2 cycles. Nonetheless, they tend to confirm the observation that even a low dose of ethinyl estradiol can produce some ovarian activity without ovulation.

Two-year treatment with oral contraceptives in hyperprolactinemic patients.

The aim of this prospective study was the follow-up for 2 years in symptoms, serum prolactin (PRL) levels, and radiological aspects of a group of young patients using oral contraceptives (OC) with hyperprolactinemia. A total of 16 hyperprolactinemic women (eight with idiopathic hyperprolactinemia and eight with pituitary microadenoma) who started OC use were admitted in the study. After 2 years of OC use, the assessable patients showed a nonsignificant decrease in plasma PRL level (26.8 +/- 29.4 micrograms/mL, range 4.2-97.1 micrograms/mL vs 56.3 +/- 31.5 micrograms/mL, range 23.5-144 micrograms/mL). No patient experienced any radiological changes during OC treatment. In conclusion, although the number of observations is limited, the data suggest that after 2 years of follow-up, no harmful effect of OC use was observed in these patients.

PIP: Recent case-control studies have failed to document any growth of pituitary adenomas following oral contraceptive (OC) use. The present study, involving 16 hyperprolactinemic OC users (8 with idiopathic and 8 with pituitary microadenoma) from Milan, Italy, also suggested exogenous estrogen has no harmful effects on these patients. Study participants underwent two blood collections before OC initiation for measurement of basal prolactin levels as well as a pituitary computed tomography or nuclear magnetic resonance scan. During OC use, prolactin measurements were taken between days 5-10 during cycles 6, 12, 18, and 24. At the end of the 24-month treatment period, all women underwent a second radiologic examination. After 2 years of OC use, women showed a nonsignificant decrease in plasma serum prolactin levels (median, 26.8 +or- 29.4 mcg/ml; range, 23.5-144 mcg/ml). No radiologic changes occurred. No patient experienced a prolactinoma enlargement during OC use. Despite a lack of evidence, OC administration is often considered contraindicated in hyperprolactinemic women.

[Induction of monofollicular cycles]. / Induzione di cicli monofollicolari.

The therapy of anovulatory infertility is not meant to obtain a pregnancy at any cost, but to restore an ovulation as physiological as possible. This involves the use of drugs and therapeutical protocols to obtain monofollicular cycles. Monofollicularity reduces the two main risks of induction of ovulation: ovarian hyperstimulation syndrome and multiple pregnancy. The aim of this study is a review of the Literature on ovulation induction and a comparison with the data of our Sterility Service. The importance of the question will be examined together with the most used ovulation induction drugs: clomiphene citrate, gonadotrophins and pulsatile GnRH. The parameters considered are: the number of follicles, single or multiple pregnancies and ovarian hyperstimulation. After a review about ovarian stimulation, the results of our Sterility Service are presented: 364 cycles of ovulation induction with clomiphene citrate, low-dose gonadotrophins or pulsatile GnRH were monitored; monofollicularity was obtained in 58,48% of ovulatory cycles. Differences between drugs will be described in the text. The therapy of anovulatory infertility aims to restore a physiological ovulation and to obtain a single pregnancy, not a pregnancy at any cost.

Influence of intermediate and uninterrupted FMR1 CGG expansions in premature ovarian failure manifestation.

BACKGROUND: Studies attempting to precisely define the range of fragile mental retardation 1 (FMR1) expansions and its inf luence in premature ovarian failure (POF) manifestation are partially lacking. To this aim, we evaluated a large cohort of POF patients for the size and, in selected cases, for the sequence of the CGG expansion. Furthermore, the correlation between POF and X-inactivation was investigated in FRAXA families. METHODS: By fluorescent PCR, 190 POF and 200 control women were sized for the CGG tract; some subjects were also characterized by sequencing and for the FMR1 activation ratio. RESULTS AND CONCLUSION: We found a significant association (19/190, 10%, P < 1 x 10(-6)) between POF and FMR1 premutation (range 63-163 repeats) and a significant enrichment (9/190, 4.7%, P = 0.021) of POF carriers of intermediate expansions (range 41-58 repeats). Interestingly, intermediate alleles were entirely composed of CGG repeats. Furthermore, the analysis of three pairs of siblings with similar FMR1 expansions and discordant for the POF phenotype showed a direct correlation between the expression of the intermediate/premutated allele and POF manifestation. The results obtained strengthen the correlation between FMR1 expansion and POF and suggest that the manifestation of the ovarian dysfunction could be influenced both by the pattern of interruption of the CGG repeat and by X-inactivation.

Comparison of GnRH agonists and antagonists in assisted reproduction cycles of patients at high risk of ovarian hyperstimulation syndrome.

BACKGROUND: During IVF or ICSI cycles, ovarian hyperstimulation syndrome (OHSS) is a major problem. The aim of this prospective, multicentre, comparative study (using historical controls) was to assess the efficacy of a GnRH antagonist protocol in preventing OHSS in selected patients who had experienced OHSS or had been at risk of OHSS in their previous IVF/ICSI attempt. METHODS AND RESULTS: Patients underwent a new cycle where the same gonadotrophin protocol was used [same dose of recombinant FSH (rFSH)] but a different protocol was used for pituitary desensitization: cetrorelix 0.25 mg multiple-dose antagonist instead of GnRH agonist long protocol. Cetrorelix 0.25 mg was administered daily, starting when the leading follicle reached a diameter of 14 mm. In other words, rFSH was administered in the new cycle according to the dosage and the step-up or step-down modalities used during the previous cycle, independently of ultrasound findings and serum estradiol (E(2)) levels. Eighty-seven patients entered the study. Out of the 87 cycles involving GnRH agonists, 49 (56.3%) were cancelled and out of the 87 involving GnRH antagonists 28 (32.2%) were cancelled [McNemar's test; 95% confidence interval (CI) -35.8% to -11.2%; P < 0.001]. After GnRH agonist cycles, we recorded 24 cases of OHSS (18 moderate and six severe; 27.6%), whereas after the GnRH antagonist cycles there were 10 cases of OHSS (nine moderate and one severe; 11.5%) (95% CI-26.4% to -5.7%; P = 0.006). There was a statistically significant reduction in the total number of follicles with a diameter >10 mm (Wilcoxon's test; Z = 6.1; P < 0.001) and of E(2) levels on the day of HCG administration (2538 versus 4322.4 pg/ml; P < 0.001) in the GnRH antagonist cycles versus GnRH agonist cycles. Twenty-nine patients had an embryo transfer in the first cycle (76.3% of oocyte retrievals) and 57 in the cycle using GnRH antagonist (96.6%). This 20.3% difference was also significant (Z-test; 95% CI 6.8-36.0%; P = 0.003). After the antagonist cycles, 18 pregnancies (20.7 per initiated cycle; 31.6% per embryo transfer) were obtained. CONCLUSIONS: Although this study presents some limitations owing to the use of historical controls, our data show a favourable effect of GnRH antagonists in reducing the incidence of OHSS and the number of assisted fertilization cycles cancelled because of the risk of OHSS in high responder patients. As a consequence, GnRH antagonist plus gonadotrophin administration could also increase the percentage of oocyte retrievals and embryo transfers in this high risk group of patients.

Management of anovulatory infertility.

Chronic anovulation is probably the major cause of human infertility and is essentially associated with four distinct endocrine conditions; hyperprolactinemic anovulation, hypogonadotrophic anovulation, normo-gonadotrophic anovulation and hypergonadotrophic anovulation. Hyperprolactinaemia and microprolactinoma are frequent findings in young women and excessive prolactin secretion impairs ovarian function causing anovulatory subfertility. Dopaminergic treatment restores ovarian function and shrinks prolacinoma. In these patients restoration of fertility with prolactin lowering drugs does not increase the incidence of multiple pregnancies or early pregnancy loss. In the vast majority of hyperprolactinemic women pregnancy is safe and could be beneficial. Cabergoline is the most effective and tolerated of the antiprolactinemic drugs. Hypogonadotrophic anovulation is frequently associated with acute or chronic emotional stress and in this case the patient should be counselled. Explanation and reassurance are the first important management steps. The use of pulsatile gonadotrophin-releasing hormone is the best strategy to induce fertility. Patients with normogonadotrophic anovulation are likely to have polycystic ovary. The most cost effective profertility treatment is the administration of an anti-oestrogen such as clomiphene or tamoxifen. The second choice therapy for patients with normogonadotrophic anovulation is ovarian stimulation with human gonadotrophin preparations. Low dose modifications give pregnancy rates lower than that with the traditional high-dose step-up protocol and intensive monitoring is required, but multiple pregnancies are less frequent. No treatment is available to enable women with hypergonadotrophic anovulation to conceive. Fertility in these patients can be promoted only by an egg donation programme.

Hormonal contraception and ovarian pathology.

Despite sporadic ovarian follicle development, hormonal contraception consistently and uniformly prevents steroidogenesis and ovulation. For their suppressive activity on ovarian androgen production, oral contraceptives remain the treatment of choice for acne and hirsutism in most hyperandrogenic women. Inhibition of the synthesis of endometrial estrogen receptors explains the effectiveness of hormonal contraception in the therapy of dysfunctional uterine bleeding and in the treatment of pain associated with pelvic endometriosis. Through the inhibition of ovarian cyclicity, the contraceptive pill lowers the incidence of functional ovarian cysts, benign breast disease, dysmenorrhea and premenstrual syndrome and shows a consistent and long-lasting protection against ovarian and endometrial cancer.

Does laparoscopic excision of endometriotic ovarian cysts significantly affect ovarian reserve? Insights from IVF cycles.

BACKGROUND: Residual ovarian function after laparoscopic excision of endometriotic ovarian cysts is a major and still unsolved topic. Ultrasonographic evaluation of ovarian response to ovulation stimulation represents a simple yet poorly employed tool to assess residual ovarian function after surgery. METHODS: Data from patients referred for IVF or ICSI between January 2001 and December 2002 were reviewed. Patients were included who previously underwent laparoscopic excision of a monolateral endometriotic ovarian cyst. The operated ovary and contralateral intact ovary were compared in terms of number of follicles with a mean diameter >15 mm at the time of hCG administration. Basal volume of the two ovaries before initiating stimulation was also compared. A paired Student's t-test was used to investigate differences between the two ovaries. RESULTS: In total, 32 patients and 46 cycles were identified. The mean (+/- SD) number of follicles >15 mm was 4.2 +/- 2.5 in the control ovary and 2.0 +/- 1.5 in the previously operated ovary (P < 0.001); this corresponded to a mean reduction of 53% (95% CI 35-72%) but did not seem to be related to the dimension of the excised ovarian cyst. The basal volume of the operated ovaries was also statistically significantly diminished, though this reduction was less relevant. CONCLUSIONS: Excision of endometriotic ovarian cysts is associated with a significant reduction in ovarian reserve. Further studies are required to clarify whether the damage is related to the surgical procedure or to the previous presence of a cyst.

An open randomized comparative study of an oral contraceptive containing ethinyl estradiol and cyproterone acetate with and without the GnRH analogue goserelin in the long-term treatment of hirsutism.

A multicenter randomized study was carried out to compare the efficacy of combined therapy with a GnRH analog (goserelin) + an oral contraceptive (OC) containing ethinyl estradiol and cyproterone acetate and same OC alone in the treatment of severe hirsutism. The effect of these two therapies was assessed in a subjective and an objective evaluation of hair growth. According to the subjective evaluation, judged by physician and patient. 95% of patients obtained a partial response. The objective response was assessed by measuring the mean diameter of hair from 3 different areas and 1 control area. The decrease in hair diameter compared to pretreatment was statistically significant for both treatments, mainly for the abdomen and face. The difference between the two groups did not reach statistical significance. Therefore, we assume that OC alone remains the treatment of choice for hirsutism. However, the addition of the GnRH analog to OC needs further investigation and could be justified for patients with no response to standard monotherapy.

Ovarian stimulation with low-dose pure follicle-stimulating hormone in polycystic ovarian syndrome anovulatory patients: effect of long-term pretreatment with gonadotrophin-releasing hormone analogue.

A randomised clinical trial was performed to evaluate the effect of a 3-month gonadotrophin-releasing-hormone analogue (GnRH-a) in one cycle of ovulation induction with low-dose pure follicle-stimulating hormone (pFSH) in patients with polycystic ovarian syndrome (PCOS) anovulation. Twenty patients with chronic anovulation due to PCOS were randomised to ovulation induction with pFSH administered in a low-dose schedule with (10 patients) and without (10 patients) a 3-month pretreatment with GnRH-a. Ultrasound scan only monitoring of follicular growth, evaluation of plasmatic oestradiol at the day of triggering of ovulation with human chorionic gonadotrophin 5,000 IU and evaluation of plasmatic progesterone 8 days after were the main outcome measures. Ovulation occurred in 9 patients treated with pFSH and in 2 patients treated with GnRH-a plus pFSH. Five pregnancies in the pFSH group and no pregnancy in the GnRH-a group were obtained. Five cycles were stopped due to multifollicular growth in the GnRH-a group and 1 in the pFSH group. Pretreatment with a 3-month administration of a GnRH-a did not improve the ovulation rate and pregnancy rate in PCOS patient ovulation induction with low-dose pFSH.

Mutation analysis of the inhibin alpha gene in a cohort of Italian women affected by ovarian failure.

BACKGROUND: Premature ovarian failure (POF) is a secondary hypergonadotrophic amenorrhoea affecting 1-3% of females, whose aetiology is almost unknown. However, inhibin alpha gene (INHalpha) has recently been indicated as candidate in POF pathogenesis. METHODS: We analysed patients affected by POF (n = 157) for the missense mutation (769G-->A transition) in the exon 2 of the INHalpha gene. The same analysis was carried out on early menopause (EM) (n = 36) and primary amenorrhoea (n = 12) patients. RESULTS: The incidence of the mutation was significantly more frequent within both POF (7/157, 4.5%) (Fisher's exact test, P = 0.030) and primary amenorrhoea (3/12, 25%) (Fisher's exact test, P < 0.001) patients, compared with the control population of women (0/100), who experienced physiological menopause. No mutation was found in EM patients. Furthermore, the likelihood of finding the mutation was statistically significant in familial (5/65; 7.7%) (Fisher's exact test, P < 0.01) but not in sporadic (2/92; 2.2%) (Fisher's exact test, P = not significant) POF, compared with the control group. The analysis of pedigrees showing the inheritance of the 769G-->A mutation and POF strengthens the concept of the disease heterogeneity, since the POF phenotype was not always associated with the mutation. Moreover, a higher prevalence of the C allele of a single nucleotide polymorphism (129C-->T), located in the 5'-UTR of the INHalpha gene, was observed in POF patients (80.3%) than in the control group (66.7%) (Fisher's exact test, P = 0.014). CONCLUSION: These data strengthen the concept of the INHalpha gene as a candidate for ovarian failure.

Endometrial morphology and ultrasound vascular findings. A randomized trial after intramuscular and vaginal progesterone supplementation in IVF.

Luteal-phase supplementation has proved necessary in Gn-RH analog and human gonadotropin-stimulated cycles. We studied the effects of vaginally and intramuscularly delivered progesterone on the endometrium. Thirty patients enrolled in an IVF program without embryo transfer due to absence of fertilization were included in the study. Patients were randomly allocated to two treatment groups. Group A (n = 15) was administered 200 mg progesterone b.i.d. by the vaginal route (Esolut, Angelini) starting on the day of oocyte pick up and group B (n = 15) was given 100 mg intramuscular progesterone once daily (Prontogest, Amsa). Six days after HCG administration, biopsies were obtained for endometrial histological maturation and estrogen (ER) and progesterone (PR) receptor analyses. In addition, ultrasound measurements of endometrial thickness were made and uterine and myometrial artery flow was determined. Serum concentrations of estriol and progesterone were measured on the day of HCG, at oocyte pick up and at endometrial biopsy. The two treatment groups were similar in terms of follicular phase parameters during superovulation with Gn-RH analog and gonadotropin. Histologic, receptor and ultrasonographic analyses showed no significant differences between the two treatment groups. Our results indicate that both intramuscular and vaginal progesterone are equally effective on the endometrium.

Effects of different amounts of LH on superovulation induction with long-acting GnRH in an IVF program.

OBJECTIVE: To evaluate whether different doses of exogenous LH affect IVF outcome. DESIGN: Retrospective analysis of two consecutive trials. SETTING: Infertility Unit of 1st Department of Obstetrics and Gynecology, University of Milan, Italy. PATIENTS: Two groups of 40 eumennorrheic patients matched for age and indications for in vitro fertilization. INTERVENTION: After suppression with a GnRH analogue, both groups received equal amounts of FSH in the first five days of stimulation; after this, the proportions of LH administered were doubled in one group because a different preparation was in use. MAIN OUTCOME MEASURES: The following observations were compared: number of scans (days to hCG), number of oocytes retrieved, fertilization rate, quality and number of embryos transferred, pregnancy and abortion rate. RESULTS: Doubling the amount of LH administered does not affect any outcome measure except the number of days of stimulation needed prior to hCG administration. CONCLUSIONS: No significant difference was seen in terms of number of oocytes, embryo number and quality, pregnancy and abortion rate. The only difference was the length of the stimulation phase.

Comparison of luteal phase profile in gonadotrophin stimulated cycles with or without a gonadotrophin-releasing hormone antagonist.

BACKGROUND: The aim of our study was to explore luteal phase hormone profiles in gonadotrophin-stimulated cycles with or without gonadotrophin-releasing hormone (GnRH) antagonist therapy during intrauterine insemination (IUI). Forty-one infertile couples were recruited in this randomized clinical study. METHODS: The 19 patients included in group A were treated for 21 cycles with recombinant FSH 150 IU/day starting from day 3 of the cycle and with the GnRH antagonist cetrorelix at the dose of 0.25 mg/day starting from the day in which a follicle with a mean diameter of > or =14 mm was seen at ultrasound scan. Cetrorelix was administered until human chorionic gonadotrophin (HCG) administration. The 22 patients included in group B were administered recombinant FSH alone at the same dosage for 27 cycles. RESULTS: The two treatment groups showed a similar increase in progesterone concentration during the luteal phase. In the mid-luteal phase (day 6 after HCG), oestradiol concentrations in group B were significantly higher compared with group A (P < 0.05) but the oestradiol:progesterone ratio was similar in the two groups. Serum LH was completely suppressed during the follicular phase only in group A, concomitantly with GnRH antagonist administration. A total of six pregnancies, all ongoing, were achieved (14.3% per patient and 12.2% per cycle), equally distributed in group A and in group B. CONCLUSION: GnRH antagonists can be safely administered in gonadotrophin-stimulated IUI cycles without luteal phase supplementation because no deleterious effects of GnRH antagonist administration were noted on luteal progesterone concentration or on the duration of the luteal phase.

Semen parameters and conception rates after intraperitoneal insemination.

OBJECTIVE: To analyze the reproductive outcome in infertile couples which underwent intraperitoneal insemination (IPI). METHODS: We analyzed a series of 216 couples who underwent IPI. Indications for treatment were unexplained infertility in 51 couples and male factor in 165. The 51 couples with unexplained infertility underwent a total of 71 cycles (20 couples underwent a second IPI cycle). The 165 couples with male factor underwent 243 cycles (165 first cycles and 78 second cycles). RESULTS: Out of the 314 cycles performed, a total of 41 clinical pregnancies were observed, with a corresponding conception rate of 13.1%. The values of conception rates for unexplained and male factor infertility were 21.1 (based on 15 pregnancies) and 10.7% (based on 26 pregnancies), respectively. Out of the 41 pregnant women, 26 gave birth to a child, thus the overall livebirth rate was 8.3% (12.7 and 7.0%, respectively, for unexplained infertility and male factor diagnostic subgroups). Considering the couples with unexplained infertility, the conception rates were 9.0, 30.8 and 20.0% for strata of < 5, 5-10 and > or = 10 millions of inseminated spermatozoa. The corresponding values were 6.8, 12.5 and 18.5%, respectively, in couples with male factor infertility (chi(2)1 trend p < 0.05). CONCLUSION: In conclusion, this series provides quantitative estimates of pregnancy rates after IPI in Italian couples with unexplained infertility or male infertility and suggested that the number of motile sperm inseminated is a determinant of pregnancy with this technique.

Preimplantation genetic diagnosis and sperm analysis by fluorescence in-situ hybridization for the most common reciprocal translocation t(11;22).

In this study we describe the pre-clinical development and clinical application of preimplantation genetic diagnosis (PGD) by fluorescence in-situ hybridization (FISH) for two non-related carriers (one male and one female) of the most common balanced reciprocal translocation: t(11;22)(q25;q12). For the couple with the female carrier, enumeration of the sex chromosomes in the embryos was also indicated (husband: 47,XXY karyotype). Four-colour FISH analysis was performed on six blastomeres from three embryos. No embryo transfer was possible because all the embryos were unbalanced. Three PGD cycles, with two-colour FISH, were carried out for the couple with the male translocation carrier. A total of 35 embryos were biopsied and diagnosed by FISH; nine out of the 35 embryos (25. 7%) were normal and seven of them were transferred (two embryos from the first and four from the third cycle), six out of 35 embryos (17%) were unbalanced, three out of 35 embryos (5.7%) were triploid or polyploid, 10 out of 35 embryos (28.6%) were mosaic and seven out of 35 embryos (20%) were chaotic. Diagnosis failed in 2.9% of the embryos. The spermatozoa of the male carrier were also analysed using three-colour FISH. Only 29.1% of the sperm cells seemed to be balanced or normal. By choosing probes lying on both sides of the breakpoints and by using a combination of sub-telomeric or locus-specific probes and centromeric probes, the use of three-colour FISH enabled detection of all the imbalances in sperm and/or cleavage-stage embryos in the patients. This may improve risk assessment and genetic counselling in the future for translocation carriers.