Impact of psychological stress on the outcomes of assisted reproduction in Tunisian infertile women

Introduction: childlessness is an emotionally difficult experience for infertile couples. Undergoing assisted reproductive treatments (ART) could generate further stress in these patients. Studies investigating the impact of anxiety on ART outcomes have shown controversial results. Moreover, there are no publications focusing on anxiety symptomsin infertile Tunisian couples. Methods: we conducted a prospective study including 79 infertile women undergoing in vitro fertilization at the Reproductive Medicine Unit of the Farhat Hached Hospital (Tunisia). Participants were asked to answer to the Beck anxiety inventory (BAI) on the day of oocyte retrieval. Accordingly, they were classified into the 3 groups: group A: very low anxiety (n= 36; BAI<21), group B: moderate anxiety (n= 24; 22≤BAI≤35) and group C: severe anxiety (n=19; BAI≥36). For each patient, two blood samples were collected to assess free cortisol level on the day of oocyte retrieval and on the day of embryo transfer. Results: results showed that women with primary infertility were significantly more stressed than those with secondary infertility (p= 0.011). Cortisol level was significantly higher on the day of embryo transfer than on the day of oocyte pick-up (p<0.0001). A lower implantation rate was found in severely anxious patients compared with moderately anxious women (p= 0.03) and those having low levels of anxiety (p= 0.001) and was negatively correlated to BAI score (r= -0.65; p= 0.001). Both clinical pregnancy and livebirth rates were similar among the three groups. Conclusion: the day of embryo transfer is the most stressful timepoint and psychological counseling is crucial to enhance implantation rate. Hence implantation took place, no effect of stress on pregnancy and live birth was found

Pathways of cyclic FEE peptide in improving gametes and embryos quality

Le peptide FEE cyclique (FEEc) a été testé en Fécondation In Vitro humaine lors d'un essai clinique prospectif randomisé autorisé par l'Agence de la Biomédecine (ABM). Les résultats ont montré une augmentation des taux de grossesses ainsi qu'une diminution des taux de fausses couches lorsque les ovocytes ont été inséminés en présence de 1 µM de FEEc comparativement au groupe contrôle. L'hypothèse était de vérifier si la diminution des taux de fausses couches en présence du peptide serait liée à l'amélioration de la ploïdie des ovocytes et/ou des embryons. Au total 775 ovocytes immatures au stade vésicule germinative (VG) et donnés pour la recherche ont été incubé in vitro de manière randomisée en présence ou en absence du FEEc pendant 24 heures. Le taux de maturation ovocytaire était de 38,1% dans le groupe témoin et de 51,9% dans le groupe FEE (p=0,0001). Parmi les ovocytes issus de MIV, 112 (56 dans le groupe contrôle et 56 dans le groupe FEEc) ont été analysés par aCGH (array Comparative Genomic Hybridization) pour l'étude de la ploïdie. Une diminution significative des taux d'aneuploïdie dans le groupe FEEc par rapport au groupe témoin a été observée (73,2% dans le groupe contrôle vs 53,6% dans le groupe FEEc; p=0,031). Le nombre moyen de chromosomes aneuploïdes par ovocyte était par ailleurs significativement diminué dans le groupe FEEc comparativement au groupe témoin (p= 0016). L'étude transcriptomique réalisée sur des ovocytes issus de MIV (n= 11 dans chaque groupe) a montré une augmentation significative de l'expression du gène FMN1 (FC= 8,68; p= 0,012) dans le groupe FEEc. Parallèlement, l'analyse par immunofluorescence du fuseau méiotique a montré une meilleure qualité du fuseau avec un alignement adéquat des chromosomes sur la plaque équatoriale dans le groupe FEEc. La distribution des mitochondries était par ailleurs plus homogène dans les ovocytes maturés en présence du FEEc. L'étude sur l'embryon humain donné pour la recherche a été réalisée sur un total de 149 embryons. 109 embryons ayant résisté à la décongélation ont été mis de manière randomisée en culture prolongée jusqu'à J5. Les taux de blastoformation étaient de 58% dans le groupe témoin et de 62% dans le groupe FEEc avec une meilleure qualité embryonnaire dans le groupe FEEc. L'analyse de la ploïdie embryonnaire a été réalisée par aCGH et les aneuploïdies retrouvées ont été vérifiées soit par FISH ( Fluorescent in situ Hybridization) soit par PCR quantitative (quantitative Polymerase Chain Reaction). Les taux d'aneuploïdie étaient similaires dans les deux groupes (15,38% dans le groupe FEEc et 26,26% dans le groupe contrôle). Le peptide FEEc améliore les résultats en FIV humaine en diminuant les taux d'aneuploïdie ovocytaire. La modification de l'expression du gène FMN1 suggère un effet du peptide sur la modification de la tension corticale de l'ovocyte qui représente un élément crucial dans l'asymétrie de la division méiotique.The cyclic FEE peptide was found to increase pregnancy rate and to decrease miscarriage rate in In Vitro Fertilization attempts. The objective was to investigate whether the impact of the cFEE on IVF results was mediated by an improvement of oocytes and/or embryo's quality. A total of 775 human oocytes arrested in the germinal vesicle stage were included in the protocol of in vitro maturation (IVM) randomly either in the control or in the cFEE treated group (1 µM). We have shown a significant improvement in maturation rate when oocytes were incubated in culture medium supplemented with cFEE (51.9% vs 38.1%; p=0.0001). Among these, 112 (56 controls and 56 FEEc) IVM oocytes were allocated for ploïdy assessment by array Comparative Genomic Hybridization (aCGH) on single cell. A significant decrease in aneuploidy rate was shown in the cFEE group (73.2% in control group vs 53.6% in cFEE group; p=0.031). The men number of aneuploid chromosomes per oocyte was also significantly decreased when the maturation medium was supplemented with cFEE (p= 0.016). The transcriptomic analysis was performed comparing sibling IVM oocytes (11 controls and 11 cFEE) and showed an increase in FMN1 gene expression in the FEE group (FC= 8,68; p= 0.012) which corroborates with the improvement in spindle morphology and chromosome alignment as assessed by immunostaining. Mitochondria distribution was seen to be more homogenous in cFEE oocytes when compared to their control counterparts. A total of 149 human embryos donated for research purpose and frozen at cleavage stage were thawed and those who have resisted to thawing procedure (n= 109) were randomly cultured either in control (n= 62) or in cFEE (n= 47) supplemented medium until day 5. 58% of embryos reached the blastocyst stage in the control group compared to 62% in the FEE group. An improvement of blastocyst quality was shown in cFEE group. Good and top-quality blastocysts were analyzed by aCGH for ploidy status and aneuploidies were then verified either by Fluorescent in situ Hybridization (FISH) or by quantitative Polymerase Chain Reaction (qPCR). Aneuploidy rates were quite similar between the two groups (respectively 15.38% and 26.26% in control and cFEE group). The decrease in miscarriage rate could be the consequence of a better chromosomes segregation during oocyte meiosis when oocytes were incubated with cFEE peptide. The modification in FMN1 expression could be the possible pathway of FEE through a modification of oocyte cortical tension to enhance asymmetric division during meiosis

Factors associated with anxiety and depression in men undergoing fertility investigations: a cross-sectional study

Abstract Background Infertility is a real public health issue because of its medical, socio-cultural, and financial impact. It does also have heavy psychological consequences on both partners. This study aimed to assess levels of anxiety and depression among men undergoing infertility investigation and to identify their associated factors. Methods We conducted a cross-sectional study in the Laboratory of Cytogenetics and Reproductive Biology of Fattouma Bourguiba University Teaching Hospital (Monastir, Tunisia) between August 30th, 2020, and March 16th, 2021. Anxiety and depression levels were assessed using the valid Arab version of the Hospital Anxiety and Depression scale (HAD). Semen parameters were analyzed and interpreted according to 2021 World Health Organization (WHO) guidelines. Results A total of 282 men were included in the current study. The mean HAD-D (depression) and HAD-A (anxiety) scores were of 6.56 ± 3.07 (IQR [4–8]) and 7.94 ± 3.73 (IQR[5–10]) respectively. Univariate analysis showed that patients having two or more comorbidities were nearly five times more likely to be anxious than those without or with only one comorbidity (ORc = 4.71; p = 0.007). Furthermore, single patients were about four times more anxious than those in couple having primary or secondary infertility (ORc = 3.85; p = 0.027). With regards to semen parameters, patients having hypospermia were more than two times anxious compared with those with normal semen volume (ORc = 2.33; p = 0.034). As for depression, we observed that patients with an infertility history lasting for a year or more have a nine times greater risk of depression (ORc = 9.848; p = 0.007). With regards to semen parameters, patients exhibiting two or more semen abnormalities, teratozoospermia and increased MAI were more depressed (ORc = 2.478; p = 0.036; ORc = 2.549: p = 0.023; ORc = 2.762; p = 0.036). Furthermore, we found a negative correlation between HAD-A scores and patient’s age. Conclusions We pointed out through the current study the associated factors with anxiety and depression in patients under fertility management to precociously identify those who need psychological counseling and hence to better manage infertility issues

Plastic used in in vitro fertilization procedures induces massive placental gene expression alterationsResearch in context

Summary: Background: The exposure to plastic derivatives during human life is deleterious. Infants conceived using ART (IVF or ICSI) have twice as many risks of major birth defects compared to naturally conceived infants. Could plastic ware used during ART trigger defects in the fetal development? Methods: Three groups of blastocysts were transferred to pseudopregnant mice. One was obtained after IVF and embryo development in plastic ware, the second in glass ware. The third, was obtained in vivo by natural mating. On day 16.5 of pregnancy, females were sacrificed and fetal organs collected for gene expression analysis. Fetal sex was determined by RT-PCR. RNA was extracted from a pool of five placental or brain samples coming from at least two litters from the same group and analyzed by hybridisation onto the mouse Affymetrix 430.2.0 GeneChips, confirmed by RT-qPCR for 22 genes. Findings: This study highlights a major impact of plastic ware on placental gene expression (1121 significantly deregulated genes), while glassware was much closer to in vivo offspring (only 200 significantly deregulated genes). Gene Ontology indicated that the modified placental genes were mostly involved in stress, inflammation and detoxification. A sex specific analysis revealed in addition a more drastic effect on female than male placentas. In the brains, whatever the comparison, less than 50 genes were found deregulated. Interpretation: Embryos incubated in plastic ware resulted in pregnancy with massive alterations of placental gene expression profile in concerted biological functions. There were no obvious effects on the brains. Besides other effects, this suggests that plastic ware in ART could be a cause of the increased level of pregnancy disorders observed recurrently in ART pregnancies. Funding: This study was funded by two grants from the Agence de la Biomedecine in 2017 and 2019

Extended culture of poor-quality supernumerary embryos improves ART outcomes

International audiencePURPOSE:The aims of this study were to investigate the possible benefits of extending the culture of poor-quality day-2 embryos (PQE) versus good-quality embryos (GQE) and to identify factors associated with pregnancy and live birth when transferring frozen-thawed blastocysts originating from GQE and PQE.METHODS:This is a retrospective cohort follow-up study performed between November 2012 and February 2015 at the IVF Laboratory Unit of Cochin University Hospital (Paris, France) including 3108 day-2 supernumerary embryos resulting from 1237 IVF/ICSI cycles.RESULTS:Total blastulation rate was 67.2% from GQE and 48.7% from PQE. Percentage of good-quality blastocysts was 60.7 and 47.9% respectively including 14.7 and 7.3% top-quality blastocysts. A total of 150 blastocysts originating from GQE and 729 from PQE were frozen, and then, 37 and 164 were thawed and transferred respectively resulting in 19 (51.4%) and 61 (37.9%) clinical pregnancies with 13 (35.1%) deliveries from GQE and 32 (19.9%) from PQE (p = 0.046) without any difference in neonatal outcomes. Quality of blastocysts that resulted in live birth was similar in the two groups. Women < 35 years old and day-5 blastocyst expansion were predictive of pregnancy and live birth.CONCLUSIONS:(i) PQE are able to reach the blastocyst stage, to implant, and to give healthy babies and (ii) women age and day of blastocyst expansion are predictive of pregnancy and live birth

A fertilin-derived peptide improves in vitro maturation and ploidy of human oocytes

International audienceObjectiveTo analyze the effect of a cyclic fertilin-derived peptide (cFEE) on in vitro maturation of human oocytes.DesignRandomized study.SettingFertility center in an academic hospital.Patient(s)Not applicable.Intervention(s)Human immature germinal vesicle-stage oocytes (n = 1,629) donated for research according to French bioethics laws were randomly allocated to groups treated with 1 or 100 μM of cFEE or to a control group. They were incubated at 37 °C in 6% CO2 and 5% O2, and their maturation was assessed using time-lapse microscopy over 24 hours. In vitro maturated metaphase II oocytes were analyzed for chromosomal content using microarray comparative genomic hybridization, and their transcriptomes were analyzed using Affymetrix Clariom D microarrays.Main Outcome Measure(s)The percentage of oocytes undergoing maturation in vitro was observed. Aneuploidy and euploidy were assessed for all chromosomes, and differential gene expression was analyzed in oocytes treated with cFEE compared with the control to obtain insights into its mechanism of action.Result(s)cFEE significantly increased the percentage of oocytes that matured in vitro and improved euploidy in meiosis II oocytes by the up-regulation of FMN1 and FLNA genes, both of which encode proteins involved in spindle structure.Conclusion(s)cFEE improves human oocyte maturation in vitro and reduces aneuploidy. It may prove useful for treating oocytes before fertilization in assisted reproductive technology and for in vitro maturation in fertility preservation programs to improve oocyte quality and the chances for infertile couples to conceive

Additional evidence for the role of chromosomal imbalances and SOX8, ZNRF3 and HHAT gene variants in early human testis development

Abstract Background Forty-six ,XY Differences/Disorders of Sex Development (DSD) are characterized by a broad phenotypic spectrum ranging from typical female to male with undervirilized external genitalia, or more rarely testicular regression with a typical male phenotype. Despite progress in the genetic diagnosis of DSD, most 46,XY DSD cases remain idiopathic. Methods To determine the genetic causes of 46,XY DSD, we studied 165 patients of Tunisian ancestry, who presented a wide range of DSD phenotypes. Karyotyping, candidate gene sequencing, and whole-exome sequencing (WES) were performed. Results Cytogenetic abnormalities, including a high frequency of sex chromosomal anomalies (85.4%), explained the phenotype in 30.9% (51/165) of the cohort. Sanger sequencing of candidate genes identified a novel pathogenic variant in the SRY gene in a patient with 46,XY gonadal dysgenesis. An exome screen of a sub-group of 44 patients with 46,XY DSD revealed pathogenic or likely pathogenic variants in 38.6% (17/44) of patients. Conclusion Rare or novel pathogenic variants were identified in the AR, SRD5A2, ZNRF3, SOX8, SOX9 and HHAT genes. Overall our data indicate a genetic diagnosis rate of 41.2% (68/165) in the group of 46,XY DSD