Jose Antonio Castilla

Assisted reproductive technology in Europe, 2009: results generated from European registers by ESHRE

STUDY QUESTION The 13th European in vitro fertilization (IVF)-monitoring (EIM) report presents the results of treatments involving assisted reproductive technology (ART) initiated in Europe during 2009: are there any changes in the trends compared with previous years? SUMMARY ANSWER Despite some fluctuations in the number of countries reporting data, the overall number of ART cycles has continued to increase year by year and, while pregnancy rates in 2009 remained similar to those reported in 2008, the number of transfers with multiple embryos (3+) and the multiple delivery rates declined. WHAT IS KNOWN ALREADY Since 1997, ART data in Europe have been collected and reported in 12 manuscripts, published in Human Reproduction. STUDY DESIGN, SIZE, DURATION Retrospective data collection of European ART data by the EIM Consortium for the European Society of Human Reproduction and Embryology (ESHRE); cycles started between 1st January and 31st December are collected on a yearly basis; the data are collected by the National Registers, when existing, or on a voluntary basis. PARTICIPANTS/MATERIALS SETTING, METHODS From 34 countries (-2 compared with 2008), 1005 clinics reported 537 463 treatment cycles including: IVF (135 621), intracytoplasmic sperm injection (ICSI, 266 084), frozen embryo replacement (FER, 104 153), egg donation (ED, 21 604), in vitro maturation (IVM, 1334), preimplantation genetic diagnosis/screening (PGD/PGS, 4389) and frozen oocyte replacements (FOR, 4278). European data on intrauterine insemination using husband/partners semen (IUI-H) and donor (IUI-D) semen were reported from 21 and 18 countries, respectively. A total of 162 843 IUI-H (+12.7%) and 29 235 IUI-D (+17.3%) cycles were included. Data available from each country are presented in the tables; total values (as numbers and percentages) refer to those countries where all data have been reported. MAIN RESULTS AND THE ROLE OF CHANCE In 21 countries where all clinics reported to the ART register, a total of 399 020 ART cycles were performed in a population of 373.8 million, corresponding to 1067 cycles per million inhabitants. For IVF, the clinical pregnancy rates per aspiration and per transfer were 28.9 and 32.9%, respectively and for ICSI, the corresponding rates were 28.7 and 32.0%. In FER cycles, the pregnancy rate per thawing was 20.9%; in ED cycles, the pregnancy rate per transfer was 42.3%. The delivery rate after IUI-H was 8.3 and 13.4% after IUI-D. In IVF and ICSI cycles, 1, 2, 3 and 4+ embryos were transferred in 24.2, 57.7, 16.9 and 1.2%, respectively. The proportions of singleton, twin and triplet deliveries after IVF and ICSI (combined) were 79.8, 19.4 and 0.8%, respectively, resulting in a total multiple delivery rate of 20.2%, compared with 21.7% in 2008, 22.3% in 2007, 20.8% in 2006 and 21.8% in 2005. In FER cycles, the multiple delivery rate was 13.0% (12.7% twins and 0.3% triplets). Twin and triplet delivery rates associated with IUI cycles were 10.4/0.7% and 10.3/0.5%, following treatment with husband and donor semen, respectively. LIMITATIONS, REASONS FOR CAUTION The method of reporting varies among countries, and registers from a number of countries have been unable to provide some of the relevant data such as initiated cycles and deliveries. As long as data are incomplete and generated through different methods of collection, results should be interpreted with caution. WIDER IMPLICATIONS OF THE FINDINGS The 13th ESHRE report on ART shows a continuing expansion of the number of treatment cycles in Europe, with more than half a million of cycles reported in 2009. The use of ICSI has reached a plateau. Pregnancy and delivery rates after IVF and ICSI remained relatively stable compared with 2008 and 2007. The number of multiple embryo transfers (3+ embryos) and the multiple delivery rate have shown a clear decline.

Assisted reproductive technology in Europe, 2011: results generated from European registers by ESHRE

STUDY QUESTION The 15th European IVF-monitoring (EIM) report presents the results of treatments involving assisted reproductive technology (ART) initiated in Europe during 2011: are there any changes in the trends compared with previous years? SUMMARY ANSWER Despite some fluctuations in the number of countries reporting data, while the overall number of ART cycles has continued to increase year by year, the pregnancy rates in 2011 decreased slightly to those reported in 2010, and the number of transfers with multiple embryos (3+) and the multiple delivery rates declined. WHAT IS KNOWN ALREADY Since 1997, ART data in Europe have been collected and reported in 14 manuscripts, published in Human Reproduction. STUDY DESIGN, SIZE, DURATION Retrospective data collection of European ART data by the EIM Consortium for the European Society of Human Reproduction and Embryology (ESHRE); cycles started between 1 January and 31 December 2011 are collected on a yearly basis. The data are collected by National Registers, when existing, or on a voluntary basis by personal information. PARTICIPANTS/MATERIALS SETTING, METHODS From 33 countries (+2 compared with 2010), 1064 clinics reported 609 973 treatment cycles including: IVF 138 592, ICSI 298 918, frozen embryo replacement (FER) 129 693, egg donation (ED) 30 198, in vitro maturation 511, preimplantation genetic diagnosis/screening 6824 and frozen oocyte replacements 5237. European data on intrauterine insemination (IUI) using husband/partners semen (IUI-H) and donor semen (IUI-D) were reported from 861 IUI laboratories in 24 countries. A total of 174 390 IUI-H and 41 151 IUI-D cycles were included. MAIN RESULTS AND THE ROLE OF CHANCE In 17 countries where all clinics reported to the ART register, a total of 361 972 ART cycles were performed in a population of 285 million inhabitants, corresponding to 1269 cycles per million inhabitants. For all IVF cycles, the clinical pregnancy rates per aspiration and per transfer were stable with 29.1 and 33.2%, respectively, and for ICSI, the corresponding rates also were stable with 27.9 and 31.8%, respectively. In FER cycles, the pregnancy rate per thawing increased to 21.3% if compared with previous years. In ED cycles, the pregnancy rate per fresh transfer decreased to 45.8% (47.4% in 2010) and increased to 33.6% (33.3% in 2010) per thawed transfer. The delivery rate after IUI-H decreased to 8.3 (8.9 in 2010), and to 12.2% (13.8% in 2010) after IUI-D. In IVF and ICSI cycles, 1, 2, 3 and 4+ embryos were transferred in 27.5, 56.7, 14.5 and 1.3% of cycles, respectively. The proportions of singleton, twin and triplet deliveries after IVF and ICSI (added together) were 80.8, 18.6 and 0.6%, respectively, resulting in a total multiple delivery rate of 19.2% compared with 20.6% in 2010, 20.2% in 2009, 21.7% in 2008, 22.3% in 2007 and 20.8% in 2006. In FER cycles, the multiple delivery rate was 13.2% (12.8% twins and 0.4% triplets). Twin and triplet delivery rates associated with IUI cycles were 9.7/0.6% and 7.3/0.3%, following IUI-H and IUI-D treatment, respectively. LIMITATIONS, REASONS FOR CAUTION The method of reporting varies among countries, and registers from a number of countries have been unable to provide some of the relevant data such as initiated cycles and deliveries. As long as data are incomplete and generated through different methods of collection, results should be interpreted with caution. WIDER IMPLICATIONS OF THE FINDINGS The 15th ESHRE report on ART shows a continuing expansion of the number of treatment cycles in Europe, with more than 600 000 cycles reported in 2011. Since 2006, the proportion of IVF to ICSI cycles has reached a plateau after a small decrease in 2009. Pregnancy and delivery rates after IVF remained relatively stable compared with 2010 and 2009. The pregnancy rate per aspiration in ICSI cycles declined for the first time by 0.9%. The multiple delivery rate is lower than ever before. STUDY FUNDING/COMPETING INTERESTS The study had no external funding; all costs are covered by ESHRE. There are no competing interests.

External quality control program for semen analysis: Spanish experience

AbstractPurpose: Results from an external quality control programme for semen analysis carried out in Spain are analysed. Methods: Quality control materials were distributed and the following seminal parameters were determined: concentration, total motility, progressive motility, rapid progressive motility, morphology and sperm vitality. The between-laboratories coefficients of variation were assessed on different types of quality control material. Results: The majority of participating laboratories utilised manual versus computer-assisted semen analysis methods. Some between-laboratories coefficients of variation ranges were: 20.8–33.8% for concentration (semen pool suspension); 13.9–19.2% for total motility (videotapes); 54.2–70.2% for sperm morphology (strict criteria using stained smears); and 9.8–41.1% for sperm vitality (stained smears). There was an inverse relation between mean percentage of sperm and coefficients of variation between laboratories for sperm motility, morphology and vitality. Conclusions: These data highlight the urgent need for improvement in the overall quality of andrology testing.

Proposal of guidelines for the appraisal of SEMen QUAlity studies (SEMQUA)

STUDY QUESTION Is there a need for a specific guide addressing studies of seminal quality? SUMMARY ANSWER The proposed guidelines for the appraisal of SEMinal QUAlity studies (SEMQUA) reflect the need for improvement in methodology and research on semen quality. WHAT IS KNOWN ALREADY From an examination of other instruments used to assess the quality of diagnostic studies, there was no guideline on studies of seminal quality. STUDY DESIGN, SIZE AND DURATION Through systematic bibliographic search, potential items were identified and grouped into four blocks: participants, analytical methods, statistical methods and results. PARTICIPANTS/MATERIALS, SETTING AND METHODS Our findings were presented to a panel of experts who were asked to identify opportunities for improvement. Then, a checklist was designed containing the questions generated by the items that summarize the essential points that need to be considered for the successful outcome of a SEMQUA. MAIN RESULTS AND THE ROLE OF CHANCE Eighteen items were identified, from which 19 questions, grouped into four blocks, were generated to constitute the final checklist. An explanation for the inclusion of each item was provided and some examples found in the bibliographic search were cited. LIMITATIONS AND REASONS FOR CAUTION We consider that not all items are equally applicable to all study designs, and so the hypothetical results are not comparable. For that reason, a score would not be fair to critically appraise a study. This checklist is presented as an instrument for appraising SEMQUAs and therefore remains open to constructive criticism. It will be further developed in the future, in parallel with the continuing evolution of SEMQUAs. WIDER IMPLICATIONS OF THE FINDINGS The final configuration of the SEMQUA is in the form of a checklist, and includes the items generally considered to be essential for the proper development of a SEMQUA. The final checklist produced has various areas of application; for example, it would be useful for designing and constructing a SEMQUA, for reviewing a paper on the question, for educational purposes or as an instrument for appraising the quality of research articles in this field. STUDY FUNDING/COMPETING INTEREST(S) None.

Mononuclear cell subpopulations in human follicular fluid from stimulated cycles.

ABSTRACT: The study of lymphocyte subsets from human follicular fluid (FF) provides an opportunity to evaluate immunological features of the ovary. We investigated the mononuclear cell subsets in FF and peripheral blood obtained at the time of laparoscopy from ten in vitro fertilization (IVF) patients. Midcycle nonpregnant peripheral blood was used as the control. A marked increase in the proportion of monocytes (CD14+) was observed in FF. Although FF was enriched with CD8+ lymphocytes, a decrease in the proportion of CD4+ lymphocytes was observed. “Memory” T cells in FF, identified by the CD4+ CD45R− phenotype, predominated over “naive” T cells (CD4+ CD45R+) at a ratio of 2:1, which differs from the ratio yielded by control blood samples (1:1). The percentage of activated T cells (CD3+ HLA‐DR+ cell) increased significantly in FF. When lymphocyte subsets were studied in the peripheral blood of IVF patients, changes similar to but less significant than those in FF were found. These data support the concept that lymphocytes play an important role in ovarian physiology.

Sperm chromatin structure assay and classical semen parameters: systematic review

The present study is based on a PubMed search and compares the clinical validity of classical semen parameters (CSP) and the sperm chromatin structure assay (SCSA) in different clinical contexts. The PubMed database was searched using keywords on the sperm diagnostic test for pregnancy in three clinical scenarios: (i) couples attempting to conceive; (ii) couples who had been attempting to conceive for 12months without success; and (iii) couples treated with intrauterine insemination (IUI). There was a considerable heterogeneity among the studies included. For couples attempting to conceive following a SCSA that produced an abnormal result, the likelihood of male factor infertility ranged from a pre-test value of 7.5% to a post-test value of 32.1% [95% confidence interval (CI) 15.7-54.5], while after CSP with an abnormal result, the post-test probability was 17.3% (95% CI 11.8-24.5). For a pre-test prevalence of male factor infertility of 50%, the post-test probability of male factor infertility after an abnormal test is very similar for both SCSA and CSP. In couples treated with IUI, the clinical validity of SCSA is higher than that of sperm morphology alone, but not enough to introduce SCSA as a test in male infertility work-up.

Apoptosis in human granulosa cells after induction of ovulation in women participating in an intracytoplasmic sperm injection program

OBJECTIVES To investigate whether analysis of granulosa cell apoptosis can be useful in assessing follicular and oocyte maturation and the regulation of granulosa cell apoptosis by follicular fluid steroids in preovulatory follicles of stimulated women. STUDY DESIGN Apoptosis in aspirated granulosa cells (n=64) was measured using the Annexin V-affinity assay by flow cytometry. Follicular fluid steroids were determined by ELISA and RIA. Statistics were evaluated using the Levenne test, Student t-test and simple linear regression analysis. RESULTS No significant differences in the number and percentage of apoptotic granulosa cells per follicle were observed according to the maturity and fertilizability of the oocytes by intracytoplasmic sperm injection within these follicles. No correlations were found between levels of steroid hormones in follicular fluid and the number and proportion of granulosa cells undergoing apoptosis. CONCLUSIONS The percentage of apoptosis in granulosa cells is not related to oocyte maturity and fertilizability by ICSI or to follicular quality in stimulated cycles of normal women. However, the possibility cannot be discarded that this parameter may be of importance at other phases of follicular development or in natural cycles when no treatment that influences follicular physiology is being applied.

Luteal cytoplasmic estradiol and progesterone receptors in human endometrium: in vitro fertilization and normal cycles

Luteal cytoplasmic estradiol (E2) and progesterone (P) receptor levels were measured from the 22nd to the 25th days of the menstrual cycle in endometrial samples obtained from seven patients in an in vitro fertilization (IVF) program who received no embryo replacement after ovarian stimulation with clomiphene citrate/human menopausal gonadotropin/human chorionic gonadotropin, and from seven normally menstruating women. Serum levels of E2, P, follicle-stimulating hormone, luteinizing hormone, and prolactin (PRL) were measured in blood samples collected at the time of biopsy. The E2 (P

Follicular fluid α-fetoprotein, carcinoembryonic antigen, and CA-125 levels in relation to in vitro fertilization and gonadotropin and steroid hormone concentrations

OBJECTIVE To investigate the possible role of alpha-fetoprotein (AFP), carcinoembryonic antigen (CEA), and CA-125 in the ovarian follicle. DESIGN alpha-Fetoprotein, CEA, and CA-125 were measured in human follicular fluid (FF) and correlated to IVF outcome and gonadotropin and gonadal steroid hormones. SETTING The In Vitro Fertilization Program at Virgen de las Nieves Hospital, Granada, Spain. PATIENTS Thirty-six FF from 12 infertile women with irreparable tubal damage who underwent treatment for ovarian stimulation with clomiphene citrate, hMG, and hCG. INTERVENTIONS Laparoscopic follicular aspiration followed 48 hours later by ET. MAIN OUTCOME MEASURE Serum and FF AFP, CEA, and CA-125 levels. RESULTS alpha-Fetoprotein, CEA, and CA-125 were detected in all samples of serum and FF. There were no significant differences between the serum and FF levels. Follicular fluid AFP, CEA, and CA-125 of fertilized oocytes were similar to those in the unfertilized oocytes group. There were no significant correlations between the FF AFP, CEA, and CA 125 levels and gonadotropin and gonadal steroid hormone levels. CONCLUSIONS alpha-Fetoprotein, CEA, and CA-125 are present in FF after ovarian stimulation, but the mean intrafollicular levels do not differ significantly regardless of the outcome of oocyte IVF.

Expression of Transcription Factors in Endometrium During Natural Cycles

AbstractPurpose: The sex steroid control of the endometrial cycle is mediated by transcription factors, four of which are the estrogen and progesterone receptors, c-jun and c-fos, all expressed by the endometrium. The aim of this study was to analyze the distribution of the transcription factors in the different endometrial compartments during natural cycles. Methods: We studied 53 reproductively-normal women, of whom 26 were in the proliferative phase and 27 in the secretory phase. An endometrial biopsy was performed and serum values of LH, FSH, estradiol, and progesterone were determined. We studied the expression of transcription factors using monoclonal antibodies. Results: A correlation between estrogen receptor and c-jun and c-fos expression was observed in stroma and epithelia, and progesterone receptor expression correlated with c-jun expression in epithelia. C-jun and c-fos presented greater expression in the proliferative phase than in the secretory phase, in the stroma and in both epithelia. No relation was found between estradiol serum levels and any transcription factor, but progesterone serum levels correlated significantly with most such factors. Conclusion: The two proto-oncogenes could play a decisive role in regulating the endometrial cycle; they could mediate the effects induced by sex steroid, and could be related to other transcription factors.