Poor knowledge of age-related fertility decline and assisted reproduction among healthcare professionals.

Reproduction is a matter of concern for individuals and society due to the postponement of childbearing, and healthcare professionals are the main source of information and counselling. This study aims to evaluate how knowledgeable healthcare professionals are about fertility and assisted reproduction, and to explore attitudes towards social oocyte freezing. A cross-sectional study was performed with 201 professionals (gynaecologists, physicians and nurses) from four public centres in Spain. Participants completed a survey about fertility, IVF, oocyte donation (OD) and social oocyte freezing, between May 2013 and March 2014. Reported mean age limits for pregnancy were 39.5 ± 4.5 (spontaneously), 43.7 ± 5.2 (IVF) and 49.0 ± 6.5 (OD). Gynaecologists reported a younger limit for spontaneous and IVF pregnancies (P < 0.001); 36.1% reported a limit for a spontaneous pregnancy >39, compared with 77.3% of other physicians and 72.9% of nurses. Regarding social oocyte freezing, 41.8% of gynaecologists thought it should be offered to every young woman, versus 62.7% of other physicians and 48.9% of nurses (P = 0.041). In conclusion, gynaecologists are more knowledgeable about fertility and assisted reproduction, while more restrictive towards social oocyte freezing. Knowledge and attitudes could influence the quality of information and counselling given to patients.

Oocyte vitrification does not affect early developmental timings after intracytoplasmic sperm injection for women younger than 30 years old.

Oocyte vitrification causes a temporary disassembly of the metaphase plate and spindle, which needs time to recover after warming. As a consequence, early post-fertilization events-such as timing of second polar body extrusion-might be altered, with unknown effects on preimplantation development, timing to pronuclear breakdown, and timing of cleavages. The aim of this study was to evaluate if differences exist among these events when comparing embryos obtained from fresh-donated versus vitrified/warmed oocytes from young women. We performed a prospective study with 201 embryos from 100 fresh and 101 vitrified/warmed oocytes that were subsequently fertilized by intracytoplasmic sperm injection. Kaplan-Meier curves of each time period were generated, in which we observed that median developmental times did not differ between embryos from fresh versus vitrified/warmed oocytes among all the metrics assessed. Thus, for young women without fertility problems, no differences exist between the timing of early developmental milestones in embryos derived from fresh or vitrified oocytes, and vitrification does not affect the preimplantation development of the resulting embryos. Mol. Reprod. Dev. 83: 624-629, 2016. © 2016 Wiley Periodicals, Inc.

Increasing fertility knowledge and awareness by tailored education: a randomized controlled trial.

Women of reproductive age have insufficient fertility knowledge and awareness. Reproductive lifespan and assisted reproduction are the primary areas in which awareness is lacking. Relatively simple interventions can be used to increase knowledge among university students; however, no intervention has been tested to date in a population with more varied education levels. The aim of this study was to evaluate which intervention most improved fertility knowledge in women attending a fertility centre for oocyte donation. A randomized controlled trial was conducted with three intervention groups: tailored, untailored and control. A questionnaire was administered on the day of the first consultation, and again at the oocyte retrieval. Two hundred and one women were enrolled and completed the pre-test, 109 started the cycle and 90 completed the post-test. The effect of the intervention was measured as the difference between the groups in their score from the pre-test to the post test. Only the tailored group showed a significant increase (+2.5; 95% CI [1.8, 3.3]; P = 0.001). Information relating to a woman's most fertile age and limits for childbearing were the most useful. Tailored oral education, therefore, increases fertility knowledge in young women, particularly in relation to their fertility lifespan.

Semen residual viral load and reproductive outcomes in HIV-infected men undergoing ICSI after extended semen preparation.

The aim of this study was to evaluate the residual presence of the human immunodeficiency virus (HIV) following a triple gradient extended semen wash from ejaculates of serodiscordant couples, and analyse their reproductive outcomes after intracytoplasmic sperm injection (ICSI). For this purpose, a retrospective analysis of our database was performed in serodiscordant couples, with HIV-infected men and non-infected women, using fresh or frozen sperm with ICSI in oocytes from either the patients or donors from January 2006 to September 2013. Overall, the rate of positive HIV test after semen washing was 1.86%. The positive beta human chorionic gonadotrophin, clinical and ongoing pregnancy rates in patients with their own oocytes were 47.1%, 37.5% and 30.8%, respectively, and 58.6%, 50.8% and 39.1%, respectively, in oocyte donation cycles. To summarize, the described method of sperm washing based on triple gradient sperm selection coupled with extensive centrifugations is a highly reliable technique for HIV removal, as it provides lower than reported post-wash positive tests while maintaining high pregnancy rates in assisted reproduction cycles. Despite extensive personnel training and effectiveness of the washing protocol, post-wash HIV test on semen is recommended to identify residual positive samples.

Are we ready to inject? Individualized LC-CUSUM training in ICSI.

PURPOSE: The objective of this prospective, single center study was to develop a personalized training scheme for intracytoplasmic sperm injection (ICSI) through the use of learning curve-cumulative summation (LC-CUSUM), which allows to tailor training to the trainee performance, and to validate it against the performance of experienced embryologists. METHODS: Five trainees microinjected latex microspheres (LM) into vitro matured oocytes. A microinjection was considered successful when the oocyte did not lyse in the 24 h following the injection. RESULTS: Each trainee became proficient at ICSI after a variable number of injections, ranging from 35 to 80. Trainees that achieved proficiency went on to perform ICSI with human gametes in a clinical setting with proficiency comparable to that of experienced embryologists. CONCLUSIONS: We show that LC-CUSUM based personalized ICSI training is feasible and allows trainees to be as proficient as trained embryologists when treating actual patients.

Is oocyte donation a risk factor for preeclampsia? A systematic review and meta-analysis.

PURPOSE: The objective of this meta-analysis is to determine whether there is a higher incidence of preeclampsia (PE) in pregnancies achieved by oocyte donation (OD) compared with pregnancies achieved by in vitro fertilization with autologous oocytes (IVF). METHODS: A systematic review was performed to identify relevant studies published from January 1994 until April 2015 with at least an abstract in English using PubMed, ISI Web of Knowledge, and clinicaltrials.gov. The 11 studies included in this systematic review were retrospective and prospective cohort studies of women reporting results on the association between oocyte donation vs. in vitro fertilization (exposure) and preeclampsia (outcome). RESULTS: Oocyte donation is a risk factor for the development of PE compared to IVF cycles, with a weighted OR of 3.12 under a fixed effects method (FEM: no heterogeneity between the studies). The weighted OR under a random effects model was 2.9 (REM: heterogeneity between the studies). The meta-regression analysis showed that neither multiple pregnancies (estimate = 0.08; p = 0.19) nor patient age (estimate = -2.29; p = 0.13) significantly explained the variability of the effect of oocyte donation on PE. Q statistic was 12.78 (p = 0.237), suggesting absence of heterogeneity between the studies. CONCLUSIONS: Pregnancies achieved by oocyte donation confer a threefold increase in the likelihood of developing PE than those achieved by in vitro fertilization with own oocytes. Physicians should be aware of this risk in order to both counsel patients and monitor pregnancies accordingly.

PLCζ sequence, protein levels, and distribution in human sperm do not correlate with semen characteristics and fertilization rates after ICSI.

PURPOSE: Sperm-borne PLCζ protein induces Ca(2+) oscillations in the oocyte and is believed to play a major role during oocyte activation. However, its implication in fertilization failure following ICSI is still debated. We analyzed PLCζ gene sequence, protein expression level, and localization in both patients with previous failed fertilization by ICSI and sperm donors with proven fertility in order to assess the association of PLCζ with both sperm characteristics and ability to fertilize. METHODS: Semen from 15 patients and 13 sperm donors with proven fertility was included in the study. Analysis of the PLCζ gene sequence, protein expression through Western blot, and protein localization by immunofluorescence were performed. RESULTS: Two patients with total fertilization failure presented mutations in heterozygosis in the PLCζ gene. Comparison with donor sample sequences displayed comparable SNP allele frequency. Distribution pattern of PLCζ did not vary significantly between donor and patient samples. Levels of PLCζ protein in sperm cells showed an interindividual variability both in patient and donor samples. Several SNPs previously reported in infertile patients were also present in fertile men. CONCLUSION: Failed fertilization occurs even when levels and distribution of PLCζ protein are within normal range. PLCζ seems to be a necessary but not sufficient factor in determining the molecular pathway involved in oocyte activation.

Oxidative stress level in fresh ejaculate is not related to semen parameters or to pregnancy rates in cycles with donor oocytes.

PURPOSE: The purpose of the present study is to study the relationship between oxidative stress (OS) in semen, semen characteristics, and reproductive outcomes in oocyte donation intracytoplasmic sperm injection (ICSI) cycles. METHODS: OS was measured in 132 semen samples. RESULTS: OS levels were as follows: very high (1.5 %), high (43.2 %), low (30.3 %), and very low (25.0 %). Overall seminal parameters were as follows: volume (ml) = 4.2 (SD 2.1), concentration (millions/ml) = 61.6 (SD 59.8), motility (a+b%) = 47.4 (SD 18.0), and normal spermatozoa (%) = 8.2 (SD 5.1). Of the 101 cycles that reached embryo transfer, 55.4 % evolved in biochemical, 46.5 % in clinical, and 43.6 % in ongoing pregnancy. OS level does not relate to seminal parameters, fertilization rate, or pregnancy outcomes. CONCLUSIONS: OS testing by nitro blue tetrazolium (NBT) in fresh ejaculate might not be useful for all patients. Reproductive results with young oocytes and ICSI do not seem to be affected by OS-level semen.

PLCζ disruption with complete fertilization failure in normozoospermia.

PURPOSE: Intracytoplasmic sperm injection (ICSI) is widely used to achieve fertilization in the presence of severe male factor, resulting in high fertilization rates. Nevertheless, 1-3 % of couples experience complete fertilization failure after ICSI. When a male factor is identified, assisted oocyte activation (AOA) can help overcome fertilization failures. The objective of this study is to describe a case of repeated complete fertilization failures after ICSI with donor oocytes, and to investigate the molecular and functional aspects of phospholipase C zeta (PLCζ) protein in the patient semen. METHODS: The patient was a normozoospermic male who had previously fathered, through natural conception, four children by a different partner. Molecular and functional analysis of sperm-specific PLCζ in the patient and control samples by means of gene sequencing, immunocytochemistry, Western blot, mouse oocyte activation test (MOAT), and mouse oocyte calcium analysis (MOCA) were used. RESULTS: PLCζ expression levels and distribution were significantly disrupted, although MOAT and MOCA did not indicate a decrease in activation ability. CONCLUSIONS: Normozoospermic males can have disrupted expression and distribution of PLCζ, and reduced activation ability after ICSI in human oocytes, despite their normal activation potential in functional testing using mouse oocytes. Discrepancy among molecular and functional data might exist, as mutations in the gene sequence may not be the only cause of alteration in PLCζ protein related to activation failures.

Empty follicle syndrome prevalence and management in oocyte donors.

STUDY QUESTION: Is the drug used for final oocyte maturation a factor in determining the prevalence of empty follicle syndrome (EFS)? SUMMARY ANSWER: The drug used for final oocyte maturation is not a factor in determining the prevalence of EFS among women unaffected by infertility. WHAT IS KNOWN ALREADY: Despite satisfactory follicular stimulation and adequate follicular development, cases of EFS, i.e. failure to recover any cumulus oocyte complex, have been reported both with hCG and GnRH agonist triggering. No standard management protocol has been proposed so far. STUDY DESIGN, SIZE, DURATION: Retrospective analysis of oocyte donation cycles performed between August 2006 and April 2013 in a large private fertility centre. PARTICIPANTS/MATERIALS, SETTING, METHODS: The analysis included 12 483 oocyte donation cycles of which 74 were EFS cycles. All cycles were triggered with either hCG or GnRH agonists. MAIN RESULTS AND THE ROLE OF CHANCE: There were no differences in the gonadotropic stimulation, pituitary suppression and triggering drug between cycles where oocytes were recovered successfully and EFS cycles. The total prevalence of EFS was 0.59%. Given the rarity of the syndrome, caution is advised when interpreting the analysis. LIMITATIONS, REASONS FOR CAUTION: The main limitation of this study is its retrospective nature. Although this is the largest analysis of EFS in donors reported so far, its statistical power is limited because the syndrome has a low incidence. In some cycles of EFS from 2006 to 2007 there is a lack of hormone data. WIDER IMPLICATIONS OF THE FINDINGS: Our findings may be generalized to oocyte donors and IVF patients younger than 35 years old, with cycles undergoing final maturation triggering with either hCG or GnRH agonists. The generalization cannot be extended to patients with an ovarian factor as the cause of their reproductive pathology. The theoretical aetiology of a temporary hypothalamic-pituitary hyposensitivity can explain the cycles where a rescue protocol with hCG has been successful. STUDY FUNDING/COMPETING INTERESTS: This work was supported in part by funding from Fundaciò EUGIN. The authors have no conflicts to declare. TRIAL REGISTRATION NUMBER: NA.

Individualized embryo transfer training: timing and performance.

STUDY QUESTION: How long is the individualized training and the stability of competence for the embryo transfer (ET) technique? SUMMARY ANSWER: The embryo transfer technique is easy-to-learn, hardly unlearned, and training should be individualized by monitoring with learning curve-cumulative summation (LC-CUSUM) curves. WHAT IS KNOWN ALREADY: Like many medical procedures, embryo transfer is an operator-dependent technique. Individualized or standardized training of these medical procedures should be monitored to determine when competence is acquired. STUDY DESIGN, SIZE, DURATION: This prospective, monocentric study involving five embryo transfer trainees was carried out between August 2011 and November 2012. PARTICIPANTS/MATERIALS, SETTING, METHODS: The study was carried out in a large private clinic. Five gynaecologist trainees during their first year of assisted reproduction subspecialty performed embryo transfer for patients undergoing either fresh IVF, oocyte donor IVF, or frozen embryo transfer. There were 586 embryo transfers performed in 96 sessions of 3-10 embryo transfers each. An embryo transfer was considered successful if it gave rise to a positive pregnancy test 14 days later. LC-CUSUM and cumulative summation (CUSUM) curves were used to determine when competence was acquired and whether it was maintained over time, respectively. The length of time between two consecutive sessions was assessed for an effect on consolidation of the acquired competence. MAIN RESULTS AND THE ROLE OF CHANCE: We observed that all five trainees became proficient in embryo transfer by procedure 15 (after procedure 15, 9, 7, 13 and 9, respectively). Once competence was achieved, one of the five trainees showed a loss of proficiency. After having acquired competence, the median pregnancy rate per embryo transfer session was significantly lower when the interval between consecutive embryo transfer sessions was ≥10 days compared with <10 days (20.0 versus 46.7%; P = 0.006). LIMITATIONS, REASONS FOR CAUTION: The patient groups included in the study were heterogeneous (IVF, oocyte donor IVF and frozen embryo transfer) and their outcomes are very variable; thus the distribution and proportion of these groups can determine the timing of competence acquisition. Our data show that low numbers of embryo transfer are needed to acquire competence, but since a relative high percentage of embryo transfers in our practice are from oocyte donor IVF, extrapolation of the findings to other clinical context should be done with caution. WIDER IMPLICATIONS OF THE FINDINGS: Personalized embryo transfer training is feasible and useful, allowing clinics, on one hand, to offer a maximum chances of pregnancy with fully trained personnel, and the other hand, to avoid the superfluous and costly overtraining of already proficient trainees. Furthermore, it is advisable to maintain a short interval of time between consecutive embryo transfer sessions after a trainee has acquired competence, to avoid a significant drop in the resulting pregnancy rate. STUDY FUNDING/COMPETING INTEREST(S): This work was supported in part by funding from Fundació Privada EUGIN. There are no conflicts of interest to declare.

Learning curves in 3-dimensional sonographic follicle monitoring during controlled ovarian stimulation.

OBJECTIVES: Three-dimensional (3D) sonographically based automated volume calculation (SonoAVC; GE Healthcare, Zipf, Austria) is being introduced in folliculometry during ovarian stimulation; however, clear training assessments in this technique are lacking. The learning curve-cumulative summation (LC-CUSUM) test provides a quantitative tool to determine when a trainee has learned a procedure. The aim of this prospective study was to assess 3D SonoAVC LC-CUSUM curves in folliculometry. METHODS: Analyses were performed on 98 oocyte donors by capturing the ovarian image in 3D and applying the 3D SonoAVC software during ovarian stimulation cycles. Each patient was scanned by an expert operator and by a trainee. Independent LC-CUSUM tests for 4 follicular diameters tracked the competency of 3 trainees in 3D SonoAVC. RESULTS: We found that the numbers of sonographic examinations required by the 3 trainees to identify the correct number of follicles of 10 mm or larger were 38, 32, and 28, respectively; for follicles of 14 mm or larger, they were 29, 28, and 28; for follicles of 18 mm or larger, they were 24, 19, and 27; and for follicles of 21 mm or larger, they were 29, 19, and 24. CONCLUSIONS: A variable number of procedures are needed to reach proficiency in 3D SonoAVC, even for trained 2-dimensional sonographers. Assessment of learning curves should be implemented when incorporating 3D SonoAVC in reproduction units.

Racial disparity in oocyte donation outcome: a multiethnic, matched cohort study.

BACKGROUND: Race and ethnicity are one of the newly investigated patient-related prognostic factors that might affect the outcome of assisted reproduction techniques. To our knowledge no data currently are available on the effect of race on oocyte donation outcome. MATERIALS: A retrospective, matched cohort study was performed in a private infertility centre evaluating 1012 Black, South-East Asian and Caucasian recipients undergoing their first oocyte donation cycles. RESULTS: A significantly lower ongoing pregnancy rate (24.6 versus 36.8%, OR: 0.56 95% CI: 0.40-0.77, P = 0.01) was observed among Black recipients compared with their matched Caucasian counterparts. The prevalence of uterine fibroids (49.6 versus 17.1%, P < 0.0001) and previous history of tubal infertility (53.2 versus 16.5%, P < 0.0001) was significantly higher among Black women. Multiple logistic regression analysis showed that, after adjusting for confounding variables, Black race was an independent risk factor for not achieving an ongoing pregnancy (for ongoing pregnancy, adjusted OR: 0.62 95% CI: 0.43-0.89, P = 0.009). Ongoing pregnancy rate (37.2 versus 37.2%, OR: 1.0 95% CI: 0.49-2.04, P = 1.0) was not significantly different between South-East Asian and matched Caucasian patients. CONCLUSIONS: Black race was an independent risk factor for not achieving an ongoing pregnancy after oocyte donation. Although yellow race does not seem to adversely affect oocyte donation, larger studies are still warranted to draw more solid conclusions. Race should be considered as an independent prognostic factor in oocyte donation.

Triggering with HCG or GnRH agonist in GnRH antagonist treated oocyte donation cycles: a randomised clinical trial.

AIM: To compare donor and recipient outcome after inducing the final oocyte maturation with hCG or GnRH agonist in GnRH-antagonist treated oocyte donation (OD) cycles. METHODS: Two-hundred fifty-seven oocyte donors were enrolled to participate in a clinical trial in a private fertility centre. After stimulation with 225 IU rFSH and Cetrorelix 0.25 mg/day, 212 oocyte donors were randomised with sealed envelopes for triggering with recombinant hCG (Ovitrelle 250 microgr, n = 106) or a GnRH agonist (triptorelin 0.2 mg, n = 106). RESULTS: The number of retrieved COCs (12 +/- 6.3 vs 11.4 +/- 6.4), mature oocytes (8 +/- 4.6 vs 7.5 +/- 4.1), the proportion of mature oocytes (67.2 +/- 20.4% vs 67.1 +/- 20.9%) and fertilisation rates (67.8 +/- 23.5% vs 71.1 +/- 22.1%) were comparable. Clinical, ongoing pregnancy and live birth rates were not statistically different in the corresponding recipient groups. Nine cases of mild and one case of severe OHSS occurred in hCG group, whereas no cases were detected in GnRH agonist group. CONCLUSIONS: The findings of our RCT suggest that donor and recipient outcome are comparable in OD cycles triggered with hCG or a GnRH agonist. Furthermore, the risk of OHSS seems to be reduced considerably, therefore the combination of a GnRH antagonist protocol with GnRH agonist triggering constitutes a safe treatment option for egg-donors.

Decreased pregnancy rate after in-vitro fertilization in HIV-infected women receiving HAART.

A study on in-vitro fertilization (IVF) was conducted among HIV-infected women. In these patients, a reduced pregnancy rate after IVF was observed if the patient's own oocytes were used. However, no significant reduction in the pregnancy rate was found if donated oocytes were used. The CD4 lymphocyte count was independently associated with ovarian resistance to hyperstimulation. Subclinical hypogonadism mediated by immunosuppression may explain these observations, suggesting the need to optimize the immunological status of the patient before considering assisted reproduction treatments.

Characteristics and clinical outcomes of patients undergoing fertility treatment by double gamete donation.

The number of women receiving in vitro fertilization cycles with both oocyte and sperm donation (double donation; DD) has grown globally in the last decade. The aim of this retrospective study, which included 1139 DD cycles, was to describe the characteristics of patients receiving DD and the outcomes of this assisted reproductive treatment. A cluster analysis identified couple 'status' as the main variable in dividing patients into categories. Three such status groups were identified for further analysis: (i) single women (SW), that is women without a partner either male or female; (ii) women with a male partner (MP); (iii) women with a female partner (FP). SW were significantly older (43.9) than patients with a MP (40.4) and a FP (41.3). Women with a male or FP comprised fewer patients with no previous assisted reproductive technology cycles (18.4% and 25.7%, respectively) compared to SW (43.5%). The proportion of patients without children before treatment was significantly different between SW (94.7%) and women with a MP (87.2%). There were no differences in clinical outcomes among the three groups studied. Biochemical pregnancy rate was 58.2% in SW, 58.4% in women with a MP and 64.9% in women with a FP. For the same groups, clinical pregnancy rates were 50.2%, 49.4% and 55.4%, while 'take-home baby' rates were 36.6%, 38.9% and 40.3%. Multiple birth and caesarean section rates were not different among the groups, with twinning rates 21.1%, 30.4% and 36%, and caesarean section rates 25.6%, 24% and 26.4% for SW, women with MP and women with FP, respectively.

Influence of Donor, Recipient, and Male Partner Body Mass index on Pregnancy Rates in Oocyte Donation Cycles.

OBJECTIVE: A high body mass index (BMI) has been shown to associate with negative reproductive outcomes. Women with high BMI have in general lower chances of getting pregnant as well as higher risk of pregnancy complications. Several studies have described in the past the relationship between high BMI and the pregnancy outcome, however, some of them have a small sample size or fail to control for variables associated with a diminished probability of pregnancy. In the present study, we aim to analyze the role of the BMI of all parties involved in oocyte donation cycles (that is: the oocyte donor, the recipient woman, and the male partner) on pregnancy outcomes. METHODS: This study includes 1092 oocyte donation cycles. Inclusion criteria were: fertilization by ICSI, frozen semen, transfer of 2 embryos at day 3 of in vitro development. For statistical analysis, BMI was divided in: low weight (<20 kg/m2), normal (20-24 kg/m2), overweight (25-29 kg/m2) and obesity (≥30 kg/m2). Quantitative and categorical variables were assessed by squared-Chi test and one-way ANOVA. The association between the BMI (recipient, oocyte donor and partner) and pregnancy rate was assessed by multivariate logistic regression. RESULTS: Laboratory outcomes and pregnancy rates do not differ among the different BMI categories of recipient, oocyte donor or partner. After adjusted analyses (for oocyte donor age, for laboratory outcomes and for age and BMI of all the parties for pregnancy outcomes), no difference was found either. CONCLUSION: In oocyte donation cycles, where donors BMI is by law mandated to be in the 18-30 range, the pregnancy rate of the oocyte recipient does not seem to be affected by the BMI of any of the parties involved.

Fertility knowledge and awareness in oocyte donors in Spain.

OBJECTIVE: To evaluate motherhood intentions and awareness of the limits of fertility as related to menstrual cycle, female age, and assisted reproductive technologies (ART) in oocyte-donation candidates in Spain. METHODS: Cross-sectional study with 229 women seeking information about oocyte donation in March-October 2013. Women were interviewed by healthcare professionals. RESULTS: The majority of participants (95.6%) wanted to be mothers in future and 36.7% already had children. Even so, knowledge about female reproduction was low: 48.3% failed to identify the ovulation time, 48.5% missed women's fertility peak before 25, and 27.9% overestimated the age limits for ART. University education does not improve global fertility knowledge and is associated with a later intended age for childbearing (p = 0.001), which results in a twofold risk of childlessness at age of 30 (RR = 1.95, 95% CI 1.11-3.43). CONCLUSION: We conclude that fertility knowledge is insufficient but, encouragingly, nearly 30% of interviewees were proactive in seeking information from the healthcare professionals. PRACTICE IMPLICATIONS: The future fertility of young people should be protected through educational interventions emphasizing the increasing phenomenon of age-related infertility at every point of contact with a women's health professional, for instance, when oocyte-donation candidates attend a fertility center for an information visit.

Detection of DNA damage in cumulus cells using a chromatin dispersion assay.

DNA damage in cumulus cells (CCs) might be related with the developmental competence of the enclosed oocytes, however, conclusive studies are missing, partially due to the lack of a reliable, cheap, fast, and reproducible DNA damage test. We report the development of a chromatin dispersion test that allows for a fast evaluation of double strand DNA (ds-DNA) damage in CCs. The whole experiment was performed using CCs from 103 oocyte retrieval cycles evaluating the prototype D3-MAX ability (a chromatin dispersion based assay) to detect DNA breaks against in situ nick translation (ISNT) and a two tailed comet assay (TT-comet). Samples were collected from women younger than 35 years of age with a good response to stimulation. Pooled cumulus cells of MII oocytes were used. The chromatin dispersion assay results correlate with the double strand-DNA breaks values assessed by the TT-comet assay (Spearman Rho = 0.624; p = 0.003;), while the correlation was poor when compared to the single strand DNA (ss-DNA) breaks observed also with the TT-comet assay (Spearman Rho = -0.141; p = 0.554). ISNT showed a correspondence in the same cells between enzymatic incorporation of modified nucleotides and halos of chromatin dispersion. We conclude that D3-Max test detects mainly ds-DNA breaks in cumulus cells and is a reliable, fast, and easy reproducible assay suitable for routine clinical practices once the influence on oocyte quality has been established.