New insights into the role of LH in early ovarian follicular growth: a possible tool to optimize follicular recruitment.

Evidence shows that LH participates in enhancing transition from the early stage to the antral stage of folliculogenesis. It has been demonstrated that functional LH receptors are expressed, albeit at a very low level and even in smaller follicles, during the phase that was traditionally considered to be gonadotrophin independent, suggesting a role for LH in accelerating the rate of progression of non-growing and primary follicles to the preantral/antral stage. Hypogonadotropic hypogonadism, together with other clinical conditions of pituitary suppression, has been associated with reduced functional ovarian reserve. The reduction in LH serum concentration is associated with a low concentration of anti-Müllerian hormone. This is the case in hypothalamic amenorrhoea, pregnancy, long-term GnRH-analogue therapy and hormonal contraception. The effect seems to be reversible, such that after pregnancy and after discontinuation of drugs, the functional ovarian reserve returns to the baseline level. Evidence suggests that women with similar primordial follicle reserves could present with different numbers of antral follicles, and that gonadotrophins may play a fundamental role in permitting a normal rate of progression of follicles through non-cyclic folliculogenesis. The precise role of gonadotrophins in early folliculogenesis, as well as their use to modify the functional ovarian reserve, must be investigated.

Chronic endometritis in recurrent implantation failure: Use of prednisone and IVF outcome.

In recurrent implantation failure patients (RIF), the main criteria for diagnosis of chronic endometritis, is the presence of plasma cells CD138+ in endometrial biopsy. The aim of the present study was to evaluate if treatment with prednisone, in patients with RIF and chronic endometritis, improve IVF outcome. A retrospective study was performed between 2019 and 2020. A total of 27 patients with RIF and an endometrial biopsy positive for CD56+ cells were enrolled. The treatment with prednisone 10 mg per day is began together with controlled ovarian stimulation (COS). Among endometrial biopsies, 13 (48.14%) were positive also for CD138 cells, and an antibiotic treatment was added. In all patients, after therapy, in the subsequent IVF cycle, the clinical pregnancy rate was 25.9% and the live birth rate was 22.2%. Analysing pregnancies according to the percentage of CD 56 cells on endometrial biopsy, the live birth rate in the subgroup of patients with marked endometritis (defined by the presence of >10% CD56+cells) was 29.41%, while in the subgroup with mild endometritis (CD 56 >5% and <10%) was 10%. In the subgroup with mild endometritis with CD 138 positive the live birth was 25%, while in patients with CD 138 negative no live birth were observed. In patients with RIF the count of at least two cell types (CD 138 and CD 56 cells) on endometrial biopsies is advisable. Our study suggests a benefit of prednisone and antibiotic treatment on live birth rate in a subsequent IVF cycle.

Ovarian Reserve Markers to Identify Poor Responders in the Context of Poseidon Classification.

It is well-known that poor ovarian reserve is a cause of infertility, poor response to gonadotrophin stimulation and poor success rate after in vitro fertilization (IVF) cycles. Some years ago a consensus was elaborated on precise criteria which can lead to a correct identification of poor responders (the Bologna criteria). More recently, the POSEIDON group has proposed a new stratified classification of patients with low prognosis, also with the aim of providing clinical indications for the management of these patients. A literature search was carried out for studies that investigated the ability of ovarian reserve markers, in particular AMH and AFC, to predict poor ovarian response in IVF cycles; secondly, studies regarding the Bologna criteria and their prognostic value were analyzed and available literature on POSEIDON classification was reported. The most recent markers of ovarian reserve (serum AMH and ultrasound AFC) have shown to provide a direct and accurate measurement of ovarian follicle pool. These markers have generally shown comparable predictive power for ovarian response and a number of retrieved oocytes in IVF cycles. "Abnormal ovarian reserve test" is a very important parameter both in the Bologna criteria and in the POSEIDON classification. Several studies have already been published about the reproductive outcome of patients defined as poor responders according to the ESHRE Bologna criteria: all of them agree on the poor IVF outcome and low pregnancy rate of these patients. Instead, being the POSEIDON classification of very recent publication, the efficacy of the POSEIDON approach in improving management and outcomes of POR patients has yet to be tested and validated with future prospective clinical trials. Prediction of poor response may help clinicians choose the stimulation protocol with the aim of gaining patient compliance and cost reduction, and many efforts have been made by researchers in this sense, including the formulation of the Bologna criteria and of the POSEIDON classification, in which the ovarian reserve markers (AMH and AFC) play a fundamental role.

Female age, serum antimüllerian hormone level, and number of oocytes affect the rate and number of euploid blastocysts in in vitro fertilization/intracytoplasmic sperm injection cycles.

OBJECTIVE: To study the relative role of female age and ovarian reserve, measured through serum antimüllerian hormone (AMH) in determining the rate and number of euploid blastocysts in in vitro fertilization (IVF)/intracytoplasmic sperm injection (ICSI) cycles. DESIGN: Retrospective analysis of cycles performed in 2014-2015. SETTING: Tertiary referral IVF center. PATIENT(S): A total of 578 infertile couples undergoing IVF/ICSI and preimplantation genetic screening (PGS) analysis. INTERVENTIONS(S): All embryos were cultured and biopsied at the blastocyst stage. The method involved whole-genome amplification followed by array comparative genome hybridization. Serum AMH was measured by means of the modified Beckman Coulter AMH Gen II assay. MAIN OUTCOME MEASURES: The rate and number of euploid blastocysts and their correlation with ovarian reserve and response to stimulation. RESULT(S): The mean (±SD) age of patients was 37.6 ± 4.1 years, and the mean number of blastocysts per patient was 3.1 ± 2. The total number of blastocysts available to the analysis was 1,814, and 36% of them were euploid after PGS. Age and serum AMH were significantly and independently related to the rate of euploid blastocysts available for patients. As an effect of the cohort size, the number of mature oocytes positively affected the total number of euploid blastocysts per patient. CONCLUSION(S): A strong positive age-independent relationship between AMH level and the rate of euploid blastocysts was found. This confirms that the measurement of ovarian reserve by means of AMH has high relevance when counseling infertile patients.

How to personalize ovarian stimulation in clinical practice.

Controlled ovarian stimulation (COS) in in vitro fertilization (IVF) cycles is the starting point from which couple's prognosis depends. Individualization in follicle-stimulating hormone (FSH) starting dose and protocol used is based on ovarian response prediction, which depends on ovarian reserve. Anti-Müllerian hormone levels and the antral follicle count are considered the most accurate and reliable markers of ovarian reserve. A literature search was performed for studies that addressed the ability of ovarian reserve markers to predict poor and high ovarian response in assisted reproductive technology cycles. According to the predicted response to ovarian stimulation (poor- normal- or high- response), it is possible to counsel couples before treatment about the prognosis, and also to individualize ovarian stimulation protocols, choosing among GnRH-agonists or antagonists for endogenous FSH suppression, and the FSH starting dose in order to decrease the risk of cycle cancellation and ovarian hyperstimulation syndrome. In this review we discuss how to choose the best COS therapy, based on ovarian reserve markers, in order to enhance chances in IVF.

Individualization of controlled ovarian stimulation in vitro fertilization using ovarian reserve markers.

INTRODUCTION: In assisted reproduction technologies (ART) the controlled ovarian stimulation (COS) therapy is the starting point from which a good oocytes retrieval depends. Treatment individualization is based on ovarian response prediction, which largely depends on a woman's ovarian reserve. Anti-Müllerian hormone (AMH) and antral follicle count (AFC) are considered the most accurate and reliable markers of ovarian reserve. EVIDENCE ACQUISITION: A literature search was carried out for studies that addressed the ability of AMH and AFC to predict poor and/or excessive ovarian response in IVF cycles. EVIDENCE SYNTHESIS: According to the predicted response to ovarian stimulation (poor- normal- or high-response) is today possible not only to personalize pre-treatment counseling with the couple, but also to individualize the ovarian stimulation protocol, choosing among GnRH-agonists or antagonists for endogenous follicle-stimulating hormone (FSH) suppression and formulating the FSH starting dose most adequate for the single patients. CONCLUSIONS: In this review we discuss how to choose the best COS therapy for the single patient, on the basis of the markers-guided ovarian response prediction.

Clinical application of a nomogram based on age, serum FSH and AMH to select the FSH starting dose in IVF/ICSI cycles: a retrospective two-centres study.

OBJECTIVE: To externally validate a nomogram based on ovarian reserve markers as a tool to optimize the FSH starting dose in IVF/ICSI cycles. STUDY DESIGN: A two-centres retrospective study including 398 infertile women undergoing their first IVF/ICSI cycle (June 2013-June 2014). IVF data were retrieved from two independent IVF centres in Italy (San Raffaele Hospital, Centre 1; Verona Hospital, Centre 2). A central lab for the routine measurement of AMH and FSH was used for both centres. All women were treated based on physical and hormonal characteristics according to locally adopted protocols. The nomogram was then retrospectively applied to the patients comparing the calculated starting dose to the one actually given. RESULTS: In Centre 1, 64/131 women (48.8%) had an ovarian response below the target. While 45 of these patients were treated with a maximal FSH starting dose (≥225 IU), n=19/131 (14.5%) were treated with a submaximal dose. The vast majority of them (n=17/19) would have received a higher FSH starting dose by using the nomogram. Seventeen patients (n=17/131) had hyper response and about half of them would have been treated with a reduced FSH starting dose according to the nomogram. In Centre 2, 142/267 patients (53.2%) had an ovarian response below the target. While 136 of these were treated with a maximal FSH starting dose (≥225 IU), n=6/267 were treated with a submaximal dose. The majority of them (n=5/6) would have received a higher FSH starting dose. Thirty-two (n=32/267) patients had hyper response and more than half of them would have been treated with a reduced FSH dose. CONCLUSION: In both Centres, applying the nomogram would have resulted in more appropriate FSH starting doses compared to the the ones actually given based on clinicians choices. The use of an objective algorithm based on patient's age, serum FSH and AMH levels may thus be an effective advice on the selection of the tailored FSH starting dose. Hence, the use of this easily available nomogram could increase the proportion of patients achieving the optimal ovarian response.

Live birth rates in the different combinations of the Bologna criteria poor ovarian responders: a validation study.

PURPOSE: to compare the baseline characteristics and chance of live birth in the different categories of poor responders identified by the combinations of the Bologna criteria and establish whether these groups comprise a homogenous population. METHODS: database containing clinical and laboratory information on IVF treatment cycles carried out at the Mother-Infant Department of the University Hospital of Modena between year 2007 and 2011 was analysed. This data was collected prospectively and recorded in the registered database of the fertility centre. Eight hundred and thirty women fulfilled the inclusion/ exclusion criteria of the study and 210 women fulfilled the Bologna criteria definition for poor ovarian response (POR). Five categories of poor responders were identified by different combinations of the Bologna criteria. RESULTS: There were no significant differences in female age, AFC, AMH, cycle cancellation rate and number of retrieved oocytes between the five groups. The live birth rate ranged between 5.5 and 7.4 % and was not statistically different in the five different categories of women defined as poor responders according to the Bologna criteria. CONCLUSION: The study demonstrates that the different groups of poor responders based on the Bologna criteria have similar IVF outcomes. This information validates the Bologna criteria definition as women having a uniform poor prognosis and also demonstrates that the Bologna criteria poor responders in the various subgroups represent a homogenous population with similar pre-clinical and clinical outcomes.

The menstrual cycle regularization following D-chiro-inositol treatment in PCOS women: a retrospective study.

Polycystic ovary syndrome is characterized by irregular cycles, hyperandrogenism, polycystic ovary at ultrasound and insulin resistance. The effectiveness of D-chiro-inositol (DCI) treatment in improving insulin resistance in PCOS patients has been confirmed in several reports. The objective of this study was to retrospectively analyze the effect of DCI on menstrual cycle regularity in PCOS women. This was a retrospective study of patients with irregular cycles who were treated with DCI. Of all PCOS women admitted to our centre, 47 were treated with DCI and had complete medical charts. The percentage of women reporting regular menstrual cycles significantly increased with increasing duration of DCI treatment (24% and 51.6% at a mean of 6 and 15 months of treatment, respectively). Serum AMH levels and indexes of insulin resistance significantly decreased during the treatment. Low AMH levels, high HOMA index, and the presence of oligomenorrhea at the first visit were the independent predictors of obtaining regular menstrual cycle with DCI. In conclusion, the use of DCI is associated to clinical benefits for many women affected by PCOS including the improvement in insulin resistance and menstrual cycle regularity. Responders to the treatment may be identified on the basis of menstrual irregularity and hormonal or metabolic markers.

Age-specific reference values for serum FSH and estradiol levels throughout the reproductive period.

High serum day 3 FSH levels are associated with poor ovarian reserve and reduced fertility, but the interpretation of FSH values according to age is still not univocal. The purpose of this study was to determine age-dependent reference values in women with regular menstrual cycles and FSH as a guide for specialists. The study was performed at the Department of Mother-Infant of a University-based tertiary care centre. One-hundred ninety-two healthy normal menstruating women were recruited for the study. All patients attended the department on menstrual cycle day 3 for a blood sample for FSH and estradiol determination. A linear relationship between FSH or estradiol serum levels and age was observed. The FSH level increased by 0.11 IU for every year of age (1 IU for every 9 years of age). The values of FSH and estradiol corresponding to the 5th, 25th, 50th, 75th, 95th centiles for any specific age have been calculated. Serum FSH levels need to be interpreted according to age-dependent reference values. Serum FSH levels on 95th centile for any age may represent a warning sign for reduced ovarian reserve.

Reference values in ovarian response to controlled ovarian stimulation throughout the reproductive period.

Abstract The age-related decline in ovarian response to gonadotropins has been well known since the beginning of ovarian stimulation in IVF cycles and has been considered secondary to the age-related decline in ovarian reserve. The objective of this study was to establish reference values and to construct nomograms of ovarian response for any specific age to gonadotropins in IVF/ICSI cycles. We analyzed our database containing information on IVF cycles. According to inclusion and exclusion criteria, a total of 703 patients were selected. Among inclusion criteria, there were regular menstrual cycle, treatment with a long GnRH agonist protocol and starting follicle-stimulating hormone (FSH) dose of at least 200 IU per day. To estimate the reference values of ovarian response, the CG-LMS method was used. A linear decline in the parameters of ovarian response with age was observed: the median number of oocytes decreases approximately by one every three years, and the median number of follicles >16 mm by one every eight years. The number of oocytes and growing follicles corresponding to the 5th, 25th, 50th, 75th and 95th centiles has been calculated. This study confirmed the well known negative relationship between ovarian response to FSH and female ageing and permitted the construction of nomograms of ovarian response.

How Much Does AMH Really Vary in Normal Women?

Anti-Mullerian Hormone (AMH) is an ovarian hormone expressed in growing follicles that have undergone recruitment from the primordial follicle pool but have not yet been selected for dominance. It is considered an accurate marker of ovarian reserve, able to reflect the size of the ovarian follicular pool of a woman of reproductive age. In comparison to other hormonal biomarkers such as serum FSH, low intra- and intermenstrual cycle variability have been proposed for AMH. This review summarizes the knowledge regarding within-subject variability, with particular attention on AMH intracycle variability. Moreover the impact of ethnicity, body mass index, and smoking behaviour on AMH interindividual variability will be reviewed. Finally changes in AMH serum levels in two conditions of ovarian quiescence, namely contraceptives use and pregnancy, will be discussed. The present review aims at guiding researchers and clinicians in interpreting AMH values and fluctuations in various research and clinical scenarios.

Polymorphisms in gonadotropin and gonadotropin receptor genes as markers of ovarian reserve and response in in vitro fertilization.

Since gonadotropins are the fundamental hormones that control ovarian activity, genetic polymorphisms may alter gonadal responsiveness to glycoproteins; hence they are important regulators of hormone activity at the target level. The establishment of the pool of primordial follicles takes place during fetal life and is mainly under genetic control. Consequently, single nucleotide polymorphisms (SNPs) in gonadotropins and their receptors do not seem to be associated with any significant modification in the endowment of nongrowing follicles in the ovary. Indeed, the age at menopause, a biological characteristic strongly related to ovarian reserve, as well as markers of functional ovarian reserve such as anti-Müllerian hormone and antral follicle count, are not different in women with different genetic variants. Conversely, some polymorphisms in FSH receptor (FSHR) seem to be associated with modifications in ovarian activity. In particular, studies suggest that the Ser680 genotype for FSHR is a factor of relative resistance to FSH stimulation resulting in slightly higher FSH serum levels, thus leading to a prolonged duration of the menstrual cycle. Moreover, some FSHR gene polymorphisms show a positive association with ovarian response to exogenous gonadotropin administration, hence exhibiting some potential for a pharmacogenetic estimation of the FSH dosage in controlled ovarian stimulation. The study of SNPs of the FSHR gene is an interesting field of research that could provide us with new information about the way each woman responds to exogenous gonadotropin administration during ovulation induction.

Individualization of the FSH starting dose in IVF/ICSI cycles using the antral follicle count.

BACKGROUND: The FSH starting dose is usually chosen according to women's age, anamnesis, clinical criteria and markers of ovarian reserve. Currently used markers include antral follicle count (AFC), which is considered to have a very high performance in predicting ovarian response to FSH. The objective of the present study to elaborate a nomogram based on AFC for the calculation of the appropriate FSH starting dose in IVF cycles. METHODS: This is a retrospective study performed at the Mother-Infant Department of Modena University Hospital. IVF patients (n=505) were subjected to blood sampling and transvaginal ultrasound for measurement of serum day3 FSH, estradiol and AFC. The variables predictive of the number of retrieved oocytes were assessed by backwards stepwise multiple regression. The variables reaching the statistical significance were then used in the calculation for the final predictive model. RESULTS: A model based on age, AFC and FSH was able to accurately predict the ovarian sensitivity and accounted for 30% of the variability of ovarian response to FSH. An FSH dosage nomogram was constructed and overall it predicts a starting dose lower than 225 IU in 50.2% and 18.1% of patients younger and older than 35 years, respectively. CONCLUSIONS: The daily FSH dose may be calculated on the basis of age and two markers of ovarian reserve, namely AFC and FSH, with the last two variables being the most significant predictors. The nomogram seems easily applicable during the daily clinical practice.

External validation of anti-Müllerian hormone based prediction of live birth in assisted conception.

BACKGROUND: Chronological age and oocyte yield are independent determinants of live birth in assisted conception. Anti-Müllerian hormone (AMH) is strongly associated with oocyte yield after controlled ovarian stimulation. We have previously assessed the ability of AMH and age to independently predict live birth in an Italian assisted conception cohort. Herein we report the external validation of the nomogram in 822 UK first in vitro fertilization (IVF) cycles. METHODS: Retrospective cohort consisting of 822 patients undergoing their first IVF treatment cycle at Glasgow Centre for Reproductive Medicine. Analyses were restricted to women aged between 25 and 42 years of age. All women had an AMH measured prior to commencing their first IVF cycle. The performance of the model was assessed; discrimination by the area under the receiver operator curve (ROCAUC) and model calibration by the predicted probability versus observed probability. RESULTS: Live births occurred in 29.4% of the cohort. The observed and predicted outcomes showed no evidence of miscalibration (p = 0.188). The ROCAUC was 0.64 (95% CI: 0.60, 0.68), suggesting moderate and similar discrimination to the original model. The ROCAUC for a continuous model of age and AMH was 0.65 (95% CI 0.61, 0.69), suggesting that the original categories of AMH were appropriate. CONCLUSIONS: We confirm by external validation that AMH and age are independent predictors of live birth. Although the confidence intervals for each category are wide, our results support the assessment of AMH in larger cohorts with detailed baseline phenotyping for live birth prediction.

Normal serum anti-Müllerian hormone levels in the general female population and the relationship with reproductive history.

OBJECTIVE: Anti-Müllerian hormone (AMH) has been evaluated by several groups as a potential novel clinical marker of ovarian reserve. Considering the wide use of AMH measurement in daily clinical practice and the large number of conditions in which it may be used, it is essential to establish reference values in the healthy female population. In this study we aim to calculate the age-by-age normal values of circulating AMH. In addition, we report on AMH levels in women according to BMI, smoking status and reproductive history. STUDY DESIGN: The study was performed at the Institute of Obstetrics and Gynecology, University of Modena, between January 2008 and December 2010. A total of 416 healthy women (aged 18-51) were recruited and serum AMH levels were measured for all of them. The centiles of AMH distribution were estimated with the CG-LMS method. The relationship between AMH levels and the womens' characteristics such as BMI, smoking status and reproductive history was analysed by using the uni- and multi-variable regression analysis and the Chi-square test. RESULTS: Serum AMH concentrations show a progressive decline with female ageing. Age-related nomograms for the 5th, 25th, 50th, 75th, and 95th percentiles of AMH were produced. Mean AMH concentrations were not modified by smoking habit and BMI and were independent of parity of the women. CONCLUSION: In the present study, we established age-specific reference values for circulating AMH levels in the eumenorrheic female population. AMH measurement produces new information on ovarian pathophysiology and ovarian reserve and the establishment of reference values for AMH is the first step for a correct interpretation of the assay.

Possibilities and limits of ovarian reserve testing in ART.

Markers of ovarian reserve are associated with ovarian aging as they decline with chronologic age, and hence may predict stages of reproductive aging including the menopause transition. Assessment of ovarian reserve include measurement of serum follicle stimulating hormone (FSH), anti-M�llerian hormone (AMH), and inhibin-B. Ultrasound determination of antral follicle count (AFC), ovarian vascularity and ovarian volume also can have a role. The clomiphene citrate challenge test (CCCT), exogenous FSH ovarian reserve test (EFORT), and GnRH-agonist stimulation test (GAST) are dynamic methods that have been used in the past to assess ovarian reserve. In infertile women, ovarian reserve markers can be used to predict low and high oocyte yield and treatment failure in women undergoing in vitro fertilization. However the markers may have limitations when an in depth analysis of their accuracy, cost, convenience, and utility is performed. As ovarian reserve markers may permit the identification of both the extremes of ovarian stimulation, a possible role for their measurement may be in the individualization of treatment strategies in order to reduce the clinical risk of ART along with optimized treatment burden. It is fundamental to clarify the cost/benefit of its use in the ovarian reserve testing before initiation of an IVF cycle and whether the ovarian reserve markers-determined strategy of ovarian stimulation for assisted conception may be associated to improved live birth rate.