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Mitochondria is a powerhouse organelle involved in ATP synthesis, calcium signaling, reactive oxygen species (ROS) by oxidative stress production, cell cycle arrest via apoptosis and sex steroid hormones biosynthesis. Improvement of sperm parameters such as motility, capacitation, acrosome reaction, and oocyte interaction, involve regulation of ROS levels by the mitochondria. In human, the relation between the quantitative level of mitochondrial DNA (mtDNA), oocyte cytoplasm maturation and fertilization potential, is not clear. It has been hypothesized that oocytes without sufficient wild type mtDNA and therefore able to generate ATP, would not normally be ovulated. This is reflected in the low numbers of mtDNA observed in degenerate oocytes obtained through super ovulation protocols during assisted reproductive technology programs. Different theories place mitochondria in a central role of oxidative damage to cells and tissues related to infertility declining and aging. Mitochondria-dependent apoptosis seems to be responsible for the pre and post-natal decline in germ cells, embryo development, implantation failure, and miscarriages. [ABSTRACT FROM AUTHOR]

Objective: To evaluate sperm DNA fragmentation in normozoospermic male partners of couples undergoing infertility evaluation. Design: Retrospective cohort study. Setting: Clinical andrology laboratory. Patient(s): A total of 1,974 consecutive normozoospermic men selected from a larger cohort of 4,345 consecutive, nonazoospermic men presenting for infertility evaluation. Intervention(s): None. Main Outcome Measure(s): Clinical parameters, conventional semen parameters, and sperm DNA fragmentation assessed by flow cytometry-based TUNEL assay and reported as percent sperm DNA fragmentation (%SDF). Result(s): The mean (±SD) %SDF and the proportion of men with high %SDF (>30%) were significantly lower in the normozoospermic compared with the entire cohort of 4,345 evaluable infertile men (17.6% ± 10.1% vs. 20.7% ± 12.4% and 11% vs. 20%, respectively). In the group of 1,974 normozoospermic men, %SDF was positively correlated with paternal age (r = 0.17) and inversely correlated with progressive motility (r = −0.26). In the subset of normozoospermic men with sperm parameters above the 50th percentile (≥73 × 106 sperm/mL, ≥55% progressive motility, and ≥14% normal forms, World Health Organization 2010 guidelines), 5% (4 of 83) had elevated %SDF (>30%). Conclusion(s): In this large cohort of normozoospermic men presenting for infertility evaluation, DNA fragmentation level is related to sperm motility and paternal age, and 11% of these men have high levels of sperm DNA fragmentation. Furthermore, the data indicate that a nonnegligible proportion (5%) of normozoospermic men with high-normal sperm parameters may also have significant sperm DNA fragmentation. [ABSTRACT FROM AUTHOR]

Objective: To investigate the relationship between MTHFR and MTRR genetic variants with respect to both blood plasma homocysteine concentration and sperm counts. Design: Polymerase chain reaction followed by specific enzymatic digestion to determine the genotype of the individuals and blood plasma homocysteine quantification by high-performance liquid chromatography. Setting: Research laboratory. Patient(s): Two hundred sixty-eight men seeking infertility counseling and 254 partners of infertile women. Intervention(s): We studied three MTHFR (c.1286A→C, c.665C→T and c.203G→A) and two MTRR (c.66A→G and c.524C→T) single-nucleotide polymorphisms and characterized sperm parameters in both oligozoospermic and normospermic men. A cohort of 522 men was examined for this study. A subgroup of 103 men was constituted for quantification of Hcy levels. Main Outcome Measure(s): Semen samples were collected for determinations of sperm concentration, motility, and morphology according to World Health Organization guidelines as well as for DNA isolation. Blood samples of the corresponding individuals were obtained to quantify plasma homocysteine levels. Result(s): We did not observe a relationship between homocysteinemia and sperm counts. The MTHFR c.665C→T variant is associated with mild hyperhomocysteinemia in blood plasma in the TT homozygous state. Conclusion(s): No association was found between MTHFR/MTRR genetic variants and sperm counts. Although no association was observed with reduced sperm counts, the MTHFR 665TT genotype is associated with a significant increase in blood plasma homocysteine levels. [ABSTRACT FROM AUTHOR]

Objective: To investigate whether homocysteine, a well known inhibitor of methylation, which is produced after imprinting and other methylation processes, can be recycled to methionine in the oocyte, at least until the stage of maternal to zygotic transition (i.e., four- to eight-cell stage); before this stage, most of the biochemical processes are carried out with the use of maternal stores of protein and mRNA. Design: A first approach using microarrays and then reverse-transcription polymerase chain reaction (RT-PCR) for methionine synthase (5-methyltetrahydrofolate-homocysteine methyltransferase [MTR]), betaine-homocysteine methyltransferase (BHMT), and cystathionine-beta synthase (CBS). Setting: Two private hospitals. Patient(s): Patients involved in IVF/ICSI procedures. Intervention(s): Germinal vesicle oocytes collected at the time of oocyte retrieval, RNA extraction amplification, RT-PCR, microarrays. Main Outcome Measure(s): mRNA expression of all the enzymes involved in the chain of methylation and recycling of homocysteine to methionine. Result(s): All of the enzymes required for methylation are present in the oocyte. Homocysteine can be recycled with BHMT and MTR. Conclusion(s): The human oocyte is able to regulate its Hcy level via remethylation using MTR and BHMT but not CBS. This aspect is important, because recent studies have shown that controlled ovarian hyperstimulation affects the homocysteine concentration in follicular fluid. This may regulate, at least in part, the risk of imprinting problems during IVF procedures. [Copyright &y& Elsevier]

Objective: To assess cytologic and cytogenetic abnormalities following round spermatid injection.Design: Prospective analysis.Setting: In vitro fertilization centers.Patient(s): Fourteen couples accepted to a round spermatid injection (ROSI) and preimplantation genetic diagnosis (PGD) program after appropriate counseling.Intervention(s): ROSI, PGD, with fluorescence in situ hybridization for chromosome enumeration.Main outcome measure(s): Cytologic and cytogenetic abnormalities in oocytes, zygotes, and blastomeres.Result(s): The fertilization rate following ROSI was 36%. Only 11 of 143 (7.7%) oocytes developed to have several blastomeres. Cytologic and cytogenetic abnormalities accounted for the vast majority of blockage at oocyte, zygote, and early mitotic division stages. Four biopsied embryos were normal. These and seven others were implanted, but no pregnancy was achieved.Conclusion(s): A PGD diagnosis for common aneuploidies and blastocyst stage transfer is feasible for ROSI cases. Failure with ROSI is cause primarily by chromosome abnormalities, so use of ROSI in assisted reproductive technologies should be limited. [Copyright &y& Elsevier]

In this study, laser-assisted intracytoplasmic sperm injection (ICSI) improved the fertilization rate and the embryo quality in patients with a history of poor ICSI outcome and with limited metaphase II oocytes. This technique is less traumatic to the oocytes during the procedure, and the use of the technique may be expanded. [ABSTRACT FROM AUTHOR]

To investigate the effect of hysteroscopic endometrial injury for treatment of recurrent implantation failure (RIF). This prospective and randomized controlled trial included 239 patients who had failed to achieve a clinical pregnancy after the transfer of at least four good-quality embryos in a minimum of three fresh or frozen–thawed embryo transfer cycles and were under the age of 40 years, who were randomized into two groups. The injury group (n = 124) received endometrial injury during their hysteroscopic procedure, whereas the control group (n = 115) did not. Patients who had endometrial pathologies were excluded from the study. There were no statistically significant differences in duration of gonadotrophin use (8.23 versus 8.30 days), total dose of gonadotrophins (2330 versus 2338 IU), number of oocytes (7.03 versus 8.21), number of mature oocytes (5.27 versus 6.02), number of fertilized oocytes (4.19 versus 4.55), number of good-quality embryos (2.07 versus 2.43), number of embryos transferred (1.97 versus 1.93) or endometrial thickness (9.04 versus 9.35 mm) between the injury group and control group, respectively. Clinical pregnancy rates (25.8% versus 15.6%, P = 0.047), live birth rates (21.8% versus 12.2%, P = 0.049) and implantation rates (14.2% versus 8.8%, P = 0.036) were significantly different, favouring the injury group. This study suggests that endometrial injury is beneficial in RIF patients to increase the odds of implantation, clinical pregnancy and live birth. [ABSTRACT FROM AUTHOR]

Objective: To study the effectiveness of a new in vitro maturation (IVM) approach based on heterologous follicular fluid (HFF) and supernatant of cumulus-granulosa cells (CGCs) mimicking the intact follicular microenvironment to rescue immature denuded oocytes (IDOs) of patients with polycystic ovary syndrome (PCOS) whose IVM or IVF outcomes remain poor.Design: Randomized controlled trial.Setting: University-affiliated private center.Patient(s): One hundred fifty-nine IDOs were obtained from 47 patients with PCOS. First, a simple IVM system (S-IVM; 40 IDOs; control group) was compared with different protocols based on the addition of autologous follicular fluid (AFF-IVM; 44 IDOs), HFF (HFF-IVM; 42 IDO), or HFF with CGC isolated from seven women without PCOS and presenting 100% in vivo oocyte maturation (HFF/CGC-IVM; 33 IDOs).Intervention(s): None.Main Outcome Measure(s): IVM outcomes were compared among the four groups (S-IVM, AFF-IVM, HFF-IVM, HFF/CGC-IVM); then the vitro and in vivo maturation results (from controlled ovarian stimulation of PCOS patients) were compared for each group.Result(s): The HFF/CGC-IVM method gave the best yield of developed blastocysts per IDO compared with S-IVM, AFF-IVM, and HFF-IVM (27% vs. 2%, 2%, and 12%, respectively). The IVM rate with the HFF/CGC-IVM method was even higher than that compared with the in vivo maturation rate (79% vs. 42%), with significant improvement in the cleavage rate (71% vs. 61%).Conclusion(s): This adapted IVM system could be used to reach an acceptable result in meiotic competence and competent metaphase II oocytes capable of developing into intact embryos after fertilization and before transfer. [ABSTRACT FROM AUTHOR]

Objective To evaluate the influence of body mass index (BMI) on semen characteristics. Design Cohort study. Setting Single private andrology laboratory. Patient(s) All patients (n = 10,665) consulting for a semen analysis from October 9, 2010, to October 8, 2011. When analyses were repeated on the same patient, only the first was included. Intervention(s) Recording of self-reported weight and height and of semen analysis. Main Outcome Measure(s) All parameters of standard semen analysis: pH, volume, sperm concentration per mL, total sperm count per ejaculate, motility (%) within 1 hour after ejaculation (overall and progressive), viability (%), and normal sperm morphology (%). Parametric and nonparametric statistical methods were applied, and results are given either with mean ± SD, or 10th, 50th, and 90th percentiles. Result(s) Semen volume decreased from 3.3 ± 1.6 to 2.7 ± 1.6 mL when BMI increased from normal (20–25 kg/m 2 ) to extreme obesity (>40 kg/m 2 ). The same was true for semen concentration (56.4 ± 54.9 to 39.4 ± 51.0 million/mL), total sperm count (171 ± 170 to 92 ± 95 million), and progressive motility (36.9 ± 16.8% to 34.7 ± 17.1%). The percentage of cases with azoospermia and cryptozoospermia increased from 1.9% to 9.1% and from 4.7% to 15.2%, respectively. The other semen characteristics were not affected. Multivariate models including age and abstinence duration confirmed these results. Conclusion(s) In this study, on a large patient sample size, increased BMI was associated with decreased semen quality, affecting volume, concentration, and motility. The percentage of normal forms was not decreased. [ABSTRACT FROM AUTHOR]

Abstract: The recent trend toward delayed parenthood raises major safety concerns because of the adverse effects of aging on couple fertility. Studies have demonstrated that aging clearly affects female fertility, but can also affect male fertility. Although several theories have been proposed, the exact mechanisms responsible for the observed age-related decline in male fertility remain to be elucidated. It has been shown that advanced paternal age (PA) is associated with reduced semen volume as well as, reduced sperm count, motility and morphology. Recent studies have also reported that paternal aging is associated with a significant increase in the prevalence of both genomic and epigenomic sperm defects. In the context of natural and intrauterine insemination (IUI) conception, advanced paternal age has been associated with lower pregnancy rates and increased rates of spontaneous abortion (independent of maternal age). In IVF and oocyte donation programs, a significant decrease in late blastocyst development has been seen in those cycles using spermatozoa of men older than 55. However, no significant relationship between paternal age and IVF or ICSI pregnancy rates has been observed. Although there are no treatments that can fully restore the age-related decline in male fertility, various measures have been shown to optimize male fertility potential. Specific therapies (e.g. varicocelectomy) and lifestyle changes (e.g. dietary antioxidant supplements) may help minimize some of the age-related deleterious effects on spermatogenesis, such as, oxidative stress and endocrine abnormalities. [Copyright &y& Elsevier]

Abstract: Women's fertility potential is declining with age because of multiples intrinsic and extrinsic factors such as life style, oxidative stress and/or endocrine disruptors and is affecting the ability of these women to conceive naturally. This declining fertility potential and the late age of motherhood is increasing significantly the number of patients consulting infertility specialists. Different strategies of investigation and management are proposed to patients over 40 in order to overcome their infertility and improve the live birth rate in these patients. Intra Uterine Insemination (IUI) in women over 40 is associated with a low rate of ongoing pregnancy and IUI should not therefore be offered always as the first line of treatment. When the predictive factors are positive IVF/ICSI seem to be good alternatives until 43 years of age. Customized ovarian stimulation and flexible laboratory methods such as in vitro maturation (IVM), preimplantation genetic diagnosis (PGD), embryo vitrification and transfer after thawing in subsequent natural or artificial cycles can improve the success rate of ART in patients over 40. Meanwhile, oocyte and embryos donation remain good options for patient over 40 with a bad prognosis and can lead to successful ongoing pregnancies until 45 years of age. Ovarian tissue cryopreservation, oocyte vitrification at the germinal vesicle (GV) stage or metaphase II stage present a breakthrough for fertility preservation but the ideal age for starting fertility preservation is still debated as well as the minimum number of oocytes to be vitrified in order to optimize the chances of pregnancy when needed at an older age. This manuscript reports the results of our own experience from patients older than 40 in the light of the published data and discusses the different therapeutic alternatives which can be proposed to patients over 40 consulting ART centres. [Copyright &y& Elsevier]

Objective: To characterize a potential genetic cause for methylation errors described in oligozoospermia. Design: Analysis of PEG1/MEST-DMR and H19-DMR methylation level in sperm, in parallel with the study of several genes on the Y chromosome, DNMT3A, and DNMT3L. Clinical outcome was also looked at regarding PEG1/MEST-DMR and H19-DMR methylation level in sperm. Setting: Research and diagnostic laboratories. Patient(s): One hundred nineteen normospermic and 175 oligozoospermic men consulting for couple infertility. Intervention(s): We studied PEG1/MEST-DMR and H19-DMR methylation profiles in 294 men. We searched for Y chromosome gene aberrations and for mutations in both DNMT3A and DNMT3L genes in men showing epimutations. Assisted reproductive technology (ART) outcomes were also investigated. Main Outcome Measure(s): Sperm samples were collected from 294 volunteers for genomic DNA isolation that was used to study methylation profiles in imprinted loci and Y chromosome SMCY, DNMT3A, and DNMT3L genes. Pregnancy rate was also studied after ART treatment using sperm showing epimutations. Result(s): Epimutations in H19-DMR and PEG1/MEST-DMR were found in 20% and 3% of oligozoospermic men, respectively. We identified an amino acid change in DNMT3A in one case and in DNMT3L in eight men with altered methylation profiles. No mutations were detected in SMCY or in selected Y chromsome genes. No correlation between ART outcome and epimutations was found. Conclusion(s): We observed epimethylations in spermatozoa of oligozoospermic individuals, but no association was found with genetic variants or in the ART outcome. [ABSTRACT FROM AUTHOR]

Objective: To evaluate the effectiveness of IV calcium infusion on prevention of ovarian hyperstimulation syndrome (OHSS) in patients with polycystic ovary syndrome undergoing assisted reproductive techniques cycles. Design: A retrospective comparative study. Setting: Assisted reproduction techniques centre in Turkey. Patient(s): Four hundred fifty-five women with high risk for OHSS. Intervention(s): The patients in group I (n = 84) were administered IV calcium gluconate for prevention of OHSS, and the patients in group II (n = 371) comprised the control group, with no manipulation for prevention of OHSS and were age- and body mass index–matched with the study group. Main Outcome Measure(s): Ovarian hyperstimulation syndrome rate, clinical pregnancy rate. Result(s): Mean (±SD) ages of the women in the calcium infusion group (group I) and the control group (group II) were comparable (30.5 ± 4.3 vs. 31.4 ± 3.9, respectively). Ovarian hyperstimulation syndrome was found in 16.2% (60 patients) in group II, whereas in group I, only 3 patients (3.6%) developed OHSS. Interestingly, all the hyperstimulation cases in group I were mild, and there was no severe effect. Implantation rates were similar in both groups. Furthermore, we obtained clinical pregnancy in nearly 40.5% in group I and 28.8% in group II. The live-birth rate was 38.1% in the calcium infusion group and 24.8% in the control group. Conclusion(s): Intravenous calcium infusion resulted in a significantly lower rate of development of OHSS for patients with polycystic ovary syndrome and high risk of OHSS. This novel therapy may be used for prevention of OHSS effectively. [ABSTRACT FROM AUTHOR]

Objective: To investigate DNA fragmentation by using terminal deoxyribonucleotidyl transferase–mediated dUTP nick-end labeling in relation to World Health Organization parameters and computer-aided sperm analysis (CASA) in sperm to determine the possibility of obtaining a correlation among CASA parameters, sperm morphology, and DNA fragmentation. Design: Sperm analysis according to World Health Organization parameters, terminal deoxyribonucleotidyl transferase–mediated dUTP nick-end labeling (TUNEL) for sperm DNA fragmentation, and CASA for sperm movement. Prospective study. Setting: All the patients were under clinical management, consulting for hypofertility at a fertility center in France. Patient(s): One thousand six hundred thirty-three men who were referred for infertility investigation, including a complete sperm analysis. Intervention(s): Sperm analysis and DNA damage testing. Main Outcome Measure(s): Sperm morphology, DNA fragmentation, and movement characteristics. Result(s): One third of the patients had a TUNEL rate of >30%. Analysis of the 21 semen parameters tested revealed that 7 of them were significantly correlated with the TUNEL results. Conclusion(s): World Health Organization sperm parameters and DNA damage are complementary, rather than strongly linked. This should be considered to more fully understand the paternal contribution in assisted reproductive technologies failures. [Copyright &y& Elsevier]