The utility of human two plus one small pronucleated zygotes (2.1PN) based on clinical outcomes and the focused ploidy analysis.

PURPOSE: Are human embryos arising from two plus one small pronucleated zygotes, called 2.1 pronuclei (PN), clinically useful? METHODS: In a retrospective embryo cohort study and prospective experimental study, a total of 287 cycles in which at least one 2.1PN was identified in the fertilization check were included. Embryonic development and clinical outcome were compared for the 1395 2PN zygotes and 304 2.1PN zygotes that were siblings. All embryos were individually cultured in time-lapse systems. Twenty-five 2.1PN-derived blastocysts, donated for research, were used in focused single-nucleotide variant ploidy analysis to identify the distribution pattern of heterozygosity. RESULTS: The average diameter of PN was 24.9 ± 2.4 µm for large PN and 10.2 ± 2.4 µm for small PN; 79.9% of small PN was derived from female pronuclei. Blastocyst formation rate and good-quality blastocyst rate were significantly lower with 2.1PN embryos than with 2PN embryos (40.0% vs. 57.7%, 21.4% vs. 33.5%, respectively). A total of 13 embryos derived from 2.1PN were transferred, and three healthy babies were born. In ploidy constitutions of trophectoderm (TE), 2.1PN-derived blastocyst TE was shown to be mostly diploid (95.8%, 23/24), and only one blastocyst showed triploid. CONCLUSIONS: It was suggested that 2.1PN embryos have lower embryonic developmental potential than 2PN embryos, but most of the 2.1PN were diploid, indicating that they are likely to be clinically usable. It is recommended to perform embryo transfer following a combination of PGT-A and ploidy analysis.

Micromapping testicular sperm extraction: A new technique for microscopic testicular sperm extraction in nonobstructive azoospermia.

Purpose: In microscopic testicular sperm extraction (mTESE) for nonobstructive azoospermia (NOA), sperm can be recovered relatively easily in some cases, and mTESE may be retrospectively considered excessive. However, mTESE is routinely performed in the majority of NOA patients because of the difficulty in predicting tissue status. A minimally invasive and comprehensive sperm retrieval method that allows on-the-spot tissue assessment is needed. We have developed and evaluated a novel sperm retrieval technique for NOA called micromapping testicular sperm extraction (MMTSE). Methods: MMTSE involves dividing the testis into four sections and making multiple small needle holes in the tunica albuginea to extract seminiferous tubules and retrieve sperm. The sperm-positive group by MMTSE (Group I) underwent additional tissue collection (ATC) via a small incision, whereas the sperm-negative group by MMTSE (Group 0) underwent mTESE. Results: In total, 40 NOA participants underwent MMTSE. Group I included 15 patients and Group 0 included 25 patients. In Group 1, sperm were recovered from all patients by ATC. In Group 0, sperm were recovered in 4 of 25 cases using mTESE. Conclusions: MMTSE shows promise as a simple method that comprehensively searches testicular tissue and retrieves sperm using an appropriate method while minimizing patient burden.

Chlormadinone acetate in progestin-primed ovarian stimulation does not negatively affect clinical results.

Purpose: To investigate whether progestin-primed ovarian stimulation (PPOS) with chlormadinone acetate (CMA) adversely affects clinical results and neonatal outcomes, or causes congenital deformities. Methods: This retrospective study was conducted at private IVF clinic from November 2018 to November 2021. Women underwent oocyte retrieval using gonadotropin-releasing hormone (GnRH) antagonist protocol (n = 835) or PPOS protocol (n = 57) were included. Eligible patients were normal ovarian responders (aged <40, AMH ≧1.0 ng/mL) with freeze-all cycle. Embryo developments, clinical results, or neonatal outcomes of singletons derived from transfer of frozen single blastocysts were compared within each group. Results: Patient characteristics were similar in both groups. The median LH level (mIU/mL) at trigger in the GnRH antagonist group [2.0 (1.2-3.7)] was significantly higher than in the PPOS group [0.9 (0.3-1.7)]. There was no cycle with premature LH surge in the PPOS group. Fertilization and blastocyst formation rates did not differ significantly between groups. Furthermore, clinical outcomes were also similar in the two groups. Congenital abnormality rates did not differ significantly [0.9% (3/329), 0.0% (0/17)]. Conclusions: CMA using ovarian stimulation did not negatively affect clinical results. Our data suggest that PPOS with CMA is an appropriate ovarian stimulation method for normal ovarian responders.

Clinical outcomes of personalized blastocyst embryo transfer after endometrial receptivity analysis: A multicenter, retrospective cohort study.

Purpose: To evaluate clinical outcomes after endometrial receptivity analysis (ERA). Methods: This was a multicenter, retrospective cohort study involving 861 women who underwent ERA testing at certified fertility clinics in Japan, and who received subsequent personalized blastocyst embryo transfers (ET) between 2018 and 2020. Clinical outcomes, including pregnancies, miscarriages, and live births, were evaluated according to receptivity status for ERA. Results: Mean patient age was 37.7 years (SD = 4.0), and the median number of previous ETs was 2 (interquartile range, 2-3). 41.0% (353/861) of patients were non-receptive for ERA testing. Clinical pregnancy, miscarriage, and live birth rates for personalized blastocyst ET were 44.5% (226/508), 26.1% (59/226), and 26.8% (136/508) for receptive patients, and 43.1% (152/353), 28.3% (43/152), and 28.9% (102/353) for non-receptive patients, all statistically nonsignificant. Multiple logistic regression demonstrated similar nonsignificant associations between receptivity and clinical outcomes. Greater patient age, smoking, and longer duration of infertility were significantly and negatively associated with receptivity, whereas a history of delivery was positively associated and statistically significant. Conclusions: Clinical outcomes after ERA testing were similar between receptive and non-receptive patients. Further prospective study including an appropriate comparison group are warranted to evaluate the efficacy of ERA testing.

Impact of endoplasmic reticulum stress on oocyte aging mechanisms.

Endoplasmic reticulum (ER) stress is associated with several aging-related diseases; however, the mechanism underlying age-related deterioration of oocyte quality is unclear. Here, we used post-ovulatory, in vivo aged mouse oocytes as a model. Super-ovulated oocytes harvested from the oviduct at 14 h and 20 h post-hCG injection were designated as 'fresh' and 'aged', respectively. Embryo development following IVF was compared between fresh, aged and ER stress-induced oocytes. Expression of the ER stress marker GRP78 was examined at each stage. To evaluate the effect of salubrinal, an ER stress suppressor, on embryo development following IVF, expression levels of GRP78 and phospho-eukaryotic initiation factor 2 alpha were compared between aged and salubrinal-treated aged oocytes. Embryo transfer of salubrinal-treated aged oocytes was performed to examine the safety of salubrinal. Similar to aged oocytes, ER stress-induced oocytes showed lower fertilization rates and poor embryo development. Following IVF, expression of GRP78 decreased with embryo development. GRP78 expression was significantly higher in aged oocytes than in fresh oocytes. Salubrinal lowered GRP78 levels and improved embryo development. No adverse effect of salubrinal treatment was found on the birth weight of pups or on organogenesis in mice. The limitation of this study was that protein kinase-like ER kinase was the only ER stress pathway examined; the role of IRE1 and ATF6 pathways was not considered. Nevertheless, salubrinal can significantly improve embryo development in in vivo aged oocytes undergoing ER stress. Hence, regulation of ER stress might represent a promising therapeutic strategy to overcome poor oocyte quality.

Long-term clinical outcomes of testicular sperm extraction and intracytoplasmic sperm injection for infertile men.

Purpose: To find the best methods to achieve the highest pregnancy and birth rates for couples needing testicular sperm extraction (TESE)-intracytoplasmic sperm injection (ICSI). Methods: Retrospectively studied were 801 patients with male factor infertility who had undergone TESE-ICSI between April, 1996 and July, 2016 and who had been categorized into four groups: obstructive azoospermia (OA); non-obstructive azoospermia (NOA); Klinefelter syndrome (KS); and cryptozoospermia (Crypt). The sperm retrieval rate, hormone levels, fertilization rate (FR), pregnancy rate (PR), and birth rate (BR) after ICSI among three groups were compared: fresh testicular sperm (FS)-fresh oocytes (FO) (Group I); frozen-thawed testicular sperm-FO (Group II); and FS-vitrified-warmed oocytes (Group III). Results: The testicular sperm recovery rate was 57.8% (463/801): 89.6% in the Crypt, 97.1% in the OA, 28.9% in the NOA, and 42.2% in the KS groups. The follicle-stimulating hormone levels were significantly higher in the NOA and KS groups and the testosterone levels were significantly lower in the KS group. The FR, PR, and BR were: 65.2%, 43.2%, and 28.5% in group I; 59.2%, 33.4%, and 18.7% in group II; and 56.4%, 33.8%, and 22.1% in group III. Conclusion: Intracytoplasmic sperm injection with FS-FO achieved the best PR and BR. It should be considered what to do in cases with no testicular sperm by TESE. The authors hope that ICSI with donor sperm will be allowed in Japan in the near future.

Efficacy of the endometrial receptivity array for repeated implantation failure in Japan: A retrospective, two-centers study.

Aim: This study aimed to assess the efficacy of the endometrial receptivity array (ERA) as a diagnostic tool and the impact of personalized embryo transfer (pET) for the treatment of patients with recurrent implantation failure (RIF) in Japan. Methods: Fifty patients with a history of RIF with frozen-thawed blastocyst transfers were recruited from July, 2015 to April, 2016. Endometrial sampling for the ERA and histological dating and a pET according to the ERA were performed. The receptive (R) or non-receptive (NR) status of the endometrium as a result of the first ERA, endometrial dating, and pregnancy rates after the pET were analyzed. Results: Of the patients with RIF, 12 (24%) were NR. Among them, eight (66.7%) were prereceptive. A clinical follow-up was possible in 44 patients who underwent the pET. The pregnancy rates were 58.8% per patient and 35.3% per first pET in the R patients and 50.0% per patient and 50.0% per first pET in the NR patients. Discrepancies between the ERA results and histological dating were seen more in the NR patients than in the R patients. Conclusions: For patients with unexplained RIF, there is a significance in searching for their personal window of implantation (WOI) using the ERA, considering the percentage of those who were NR and the pregnancy rates that resulted from the pET. By transferring euploid embryos in a personal WOI, much better pregnancy rates are expected.

Poor embryo development in post-ovulatory in vivo-aged mouse oocytes is associated with mitochondrial dysfunction, but mitochondrial transfer from somatic cells is not sufficient for rejuvenation.

STUDY QUESTION: Does in vivo aging of mouse oocytes affect mitochondrial function? SUMMARY ANSWER: Mitochondrial function was impaired in post-ovulatory in vivo-aged mouse oocytes and microinjection of somatic cell mitochondria did not rescue poor fertilization and embryonic development rates. WHAT IS KNOWN ALREADY: The mechanisms underlying the decline in oocyte quality associated with oocyte aging remain unknown, although studies have suggested that the decline is regulated by mitochondrial dysfunction. However, only a limited number of studies have provided direct evidence implicating mitochondrial dysfunction in oocyte quality during the aging of oocytes. STUDY DESIGN, SIZE, DURATION: We used post-ovulatory, in vivo-aged mouse oocytes as a model for studying low-quality oocytes in oocyte aging. PARTICIPANTS/MATERIALS, SETTING, METHOD: Superovulated oocytes released from the oviduct at 14 h and 20-24 h post-hCG injection were designated as 'fresh' and 'aged' oocytes, respectively. Membrane potentials and oxygen consumption in single oocytes were evaluated as measures of mitochondrial function in fresh and aged oocytes. Mitochondrial transcriptional factor A (TFAM) expression levels were examined by western blotting, and colocalization of mitochondria and TFAM was analyzed by measuring immunofluorescence in fresh and aged oocytes. IVF and blastocyst formation rates were calculated after oocyte microinjection with mitochondria derived from liver cells. MAIN RESULTS AND THE ROLE OF CHANCE: The average mitochondrial membrane potential in fresh oocytes was significantly higher than that in aged oocytes (P < 0.05). The average oxygen consumption rate in aged oocytes was significantly lower than that in fresh oocytes (P < 0.05). Although total TFAM expression was unchanged, its colocalization with mitochondria decreased in aged oocytes. IVF and blastocyst formation rates for mitochondrion-injected aged oocytes were not significantly different from those for buffer-injected aged oocytes. LARGE SCALE DATA: Not applicable. LIMITATIONS, REASONS FOR CAUTION: A limitation of this study is that we did not examine the effects of microinjecting mitochondria from other somatic cell types into aged oocytes on their fertilization and embryonic development rates. WIDER IMPLICATIONS OF THE FINDINGS: The results from the present study showed that poor embryonic development was associated with impairment of mitochondrial functions in in vivo-aged oocytes. However, the microinjection of mitochondria from liver cells did not improve the low fertilization and embryonic development rates of aged oocytes. It remains to be demonstrated whether oocyte quality can be rescued by the transfer of cytosolic factors or cellular organelles, such as the endoplasmic reticulum or mitochondria, from specific cell types. STUDY FUNDING/COMPETING INTERESTS: This study was supported by Grants-in-Aid for General Science Research to Toshifumi Takahashi (No. 25462550) and Hideki Igarashi (No. 26462474). The funding source played no role in study design in the collection, analysis, and interpretation of data; in the writing of the report; and in the decision to submit the article for publication. The authors have no conflict of interest to disclose.

Development of a new clinically applicable device for embryo evaluation which measures embryo oxygen consumption.

STUDY QUESTION: Does a new system-the chip-sensing embryo respiration monitoring system (CERMs)-enable evaluation of embryo viability for potential application in a clinical IVF setting? SUMMARY ANSWER: The system enabled the oxygen consumption rate of spheroids, bovine embryos and frozen-thawed human embryos to be measured, and this rate corresponded to the developmental potential of embryos. WHAT IS ALREADY KNOWN: To date, no reliable and clinically suitable objective evaluation methods for embryos are available, which circumvent the differences in inter-observer subjective view. Existing systems such as the scanning electrochemical microscopy (SECM) technique, which enables the measurement of oxygen consumption rate in embryos, need improvement in usability before they can be applied to a clinical setting. STUDY DESIGN, SIZE, DURATION: This is a prospective original research study. The feasibility of measuring the oxygen consumption rate was assessed using CERMs for 9 spheroids, 9 bovine embryos and 30 redundant frozen-thawed human embryos. The endpoints for the study were whether CERMs could detect a dissolved oxygen gradient with high sensitivity, had comparable accuracy to the SECM measuring system with improved usability, and could predict the development of an embryo to a blastocyst by measuring the oxygen consumption rate. The relationship between the oxygen consumption rate and standard morphological evaluation was also examined. PARTICIPANTS/MATERIALS, SETTING, METHODS: We developed a new CERMs, which enables the oxygen consumption rate to be measured automatically using an electrochemical method. The device was initially used for measuring a dissolved oxygen concentration gradient in order to calculate oxygen consumption rate using nine spheroids. Next, we evaluated data correlation between the CERMs and the SECM measuring systems using nine bovine embryos. Finally, the oxygen consumption rates of 30 human embryos, which were frozen-thawed on 2nd day after fertilization, were measured by CERMs at 6, 24, 48, 72 and 96 h after thawing with standard morphological evaluation. Furthermore, the developed blastocysts were scored using the blastocyst quality score (BQS), and the correlation with oxygen consumption rate was also assessed. MAIN RESULTS AND THE ROLE OF CHANCE: The device enabled the oxygen consumption rate of an embryo to be measured automatically within a minute. The oxygen concentration gradient profile showed excellent linearity in a distance-dependent change. A close correlation in the oxygen consumption rates of bovine embryos was observed between the SECM measuring system and CERMs, with a determination coefficient of 0.8203 (P = 0.0008). Oxygen consumption rates of human embryos that have reached the blastocyst stage were significantly higher than those of arrested embryos at 48, 72 and 96 h after thawing (P = 0.039, 0.004 and 0.049, respectively). Thus, in vitro development of frozen-thawed human embryos to the blastocyst stage would be predicted at 48 h after thawing (day 4) by measuring the oxygen consumption using CERMs. Although a positive linear relationship between BQS and the oxygen consumption rate was observed [the determination coefficient was R(2) = 0.6537 (P = 0.008)], two blastocysts exhibited low oxygen consumption rates considering their relatively high BQS. This suggests that morphology and metabolism in human embryos might not correlate consistently. LIMITATIONS, REASONS FOR CAUTION: Transfer of the embryo and pregnancy evaluation was not performed. Thus, a correlation between oxygen consumption and the in vivo viability of embryos remains unknown. Clinical trials, including embryo transfer, would be desirable to determine a threshold value to elect clinically relevant, quality embryos for transfer. We utilized frozen-thawed human embryos in this study. The effect of these manipulations on the respiratory activity of the embryo is also unknown. WIDER IMPLICATIONS OF THE FINDINGS: Selection of quality embryos, especially in a single embryo transfer cycle, by CERMs may have an impact on obtaining better clinical outcomes, albeit with clinical trials being required. Furthermore, the early determination of quality embryos by CERMs may enable the omission of long-term in vitro embryo culture to the blastocyst stage. CERMs is scalable technology that can be integrated into incubators and/or other embryo evaluation systems, such as the time-lapse systems, due to its chip-based architecture. Thus, CERMS would enable automatic measurement of oxygen consumption, under 5% CO2, in the near future, in order to reduce oxidative stress from exposure to atmospheric air. STUDY FUNDING/COMPETING INTERESTS: This study was supported by grants from the Health and Labor Sciences Research Grant (H24-Hisaichiiki-Shitei-016). The authors have no conflicts of interest. TRIAL REGISTRATION NUMBER: Not applicable.

Predictive factors for oocyte retrieval failure in controlled ovarian hyperstimulation protocols: a retrospective observational cohort study.

BACKGROUND: Oocyte retrieval failure following an ovarian hyperstimulation protocol is uncommon in assisted reproductive technology (ART) programs. We analyzed the predictive factors for oocyte retrieval failure following controlled ovarian hyperstimulation (COH) with gonadotropin-releasing hormone (GnRH) agonist and GnRH antagonist protocols in ART programs. METHODS: This study was a retrospective cohort observational study. In total, 744 cycles from 361 patients who underwent controlled ovarian hyperstimulation with GnRH agonist long protocol or antagonist protocol were analyzed. Treatment cycles with oocyte retrieval failure and with one or more oocytes retrieved were compared to determine predictive factors for oocyte retrieval failure using univariate and multilevel multivariate logistic regression analyses. RESULTS: Oocyte retrieval failure occurred in 38 cycles (5.1%). The oocyte retrieval failure rate of the GnRH antagonist protocol (8.1%) was significantly higher than that of the GnRH agonist long protocol (3.7%). On multilevel multivariate logistic analysis, cycles with GnRH antagonist protocol (odds ratio [OR] 3.06, 95% confidence interval [CI] 1.05-8.96), estradiol level on the day of human chorionic gonadotropin (hCG) injection (OR 0.997, 95% CI 0.996-0.998), and luteinizing hormone (LH) level on the day of hCG injection (OR 1.19, 95% CI 1.06-1.33) were independent predictive factors for oocyte retrieval failure. The efficacy of estradiol and LH levels on the day of hCG injection for predicting oocyte retrieval failure was evaluated using receiver operating characteristic curves. In all cycles, the areas under the curve (AUCs) for estradiol and LH were 0.84 and 0.63, respectively, for all cycles; 0.84 and 0.52, respectively, for cycles with GnRH agonist long protocol; and 0.81 and 0.82, respectively, for cycles with GnRH antagonist protocol. CONCLUSIONS: Our results suggest that in cycles with GnRH antagonist protocol, the levels of estradiol and LH on the day of hCG injection might be predictive factors for oocyte retrieval failure. This relationship may provide useful information to both patients and physicians for developing better COH protocols in ART programs.

Oocyte aging underlies female reproductive aging: biological mechanisms and therapeutic strategies.

In recent years, postponement of marriage and childbearing in women of reproductive age has led to an increase in the incidence of age-related infertility. The reproductive aging process in women is assumed to occur due to a decrease in both the quantity and quality of the oocytes, with the ultimate result being a decline in fecundity. This age-related decline in fecundity is strongly dependent on oocyte quality, which is critical for fertilization and subsequent embryo development. Aged oocytes display increased chromosomal abnormality and dysfunction of cellular organelles, both of which factor into oocyte quality. In particular, mitochondrial dysfunction has been suggested as a major contributor to the reduction in oocyte quality as well as to chromosomal abnormalities in aged oocytes and embryos. Participation of oxidative stress in the oocyte aging process has been proposed because oxidative stress has the capacity to induce mitochondrial dysfunction and directly damage many intracellular components of the oocytes such as lipids, protein, and DNA. In an attempt to improve mitochondrial function in aged oocytes, several therapeutic strategies have been investigated using both animal models and assisted reproductive technology. Here, we review the biological mechanisms and present status of therapeutic strategies in the female reproductive aging field and indicate possible future therapeutic strategies.

Effects of different trehalose concentrations in a warming medium on embryo survival and clinical outcomes in vitrified human embryos.

BACKGROUND/AIMS: In the present study, we examined the effects of different concentrations of trehalose in a warming medium on both embryo survival and clinical outcomes in vitrified-warmed embryo transfer cycles. METHODS: We retrospectively analyzed a total of 209 vitrified-warmed cycles from 177 patients who underwent in vitro fertilization or intracytoplasmic sperm injection and embryo transfer. Embryos were cryopreserved by the vitrification method and warmed in solutions containing either 0.5 or 1.0 M trehalose. We compared the 0.5 and 1.0 M trehalose warming solution groups with respect to the embryo survival rate after warming and clinical outcomes. RESULTS: The embryo survival rate in the 1.0 M trehalose group (96.5%) was significantly higher than that in the 0.5 M trehalose group (57.0%). The percentage of embryo transfers after warming in the 1.0 M trehalose group (94.3%) was significantly higher than that in the 0.5 M trehalose group (83.7%). The clinical pregnancy rate in the 1.0 M trehalose group (25.0%) was significantly higher than that in the 0.5 M trehalose group (11.1%). CONCLUSION: Embryo survival and clinical pregnancy rates were higher when a 1.0 M trehalose solution was used than when a 0.5 M trehalose solution was used during the embryo warming process.

Molecular mechanism of poor embryo development in postovulatory aged oocytes: mini review.

Oocyte quality is a key factor in determining embryo development; however, we have a poor understanding of what constitutes oocyte quality or the mechanisms governing it. Postovulatory aging of oocytes that have not been fertilized for a prolonged time after ovulation is known to significantly impair oocyte quality and subsequent embryo development after fertilization. Embryos derived from postovulatory-aged oocytes are prone to undergo apoptosis due to the decreased Bcl-2 expression. Postovulatory aging of oocytes changes the patterns of Ca(2+) oscillations at fertilization as a result of impaired Ca(2+) regulation in the endoplasmic reticulum. Moreover, postovulatory aging of oocytes impairs mitochondrial adenosine triphosphate production as a result of increasing oxidative stresses. Oxidative stresses also affect intracellular Ca(2+) regulation and impair embryo development after fertilization. Collectively, the mechanism of postovulatory oocyte aging might be involved in reactive oxygen species-induced mitochondrial injury followed by abnormal intracellular Ca(2+) regulation in the endoplasmic reticulum.

Bezafibrate restores the inhibition of FSH-induced follicular development and steroidogenesis by tumor necrosis factor-alpha through peroxisome proliferator-activated receptor-gamma pathway in an in vitro mouse preantral follicle culture.

We recently reported that bezafibrate, a lipid-lowering drug of the fibrate class, administered in addition to clomiphene citrate (CC) successfully induced ovulation in CC-resistant polycystic ovary syndrome (PCOS) patients. We hypothesized that bezafibrate may directly affect ovarian follicle development. Insulin resistance and compensatory hyperinsulinemia are important for the pathogenesis of PCOS. In this study, we first examined the effects of tumor necrosis factor-alpha (TNF), which plays a role in insulin resistance, on follicle development by using the follicle culture system. TNF significantly inhibited follicle-stimulating hormone (FSH)-induced follicle development, 17beta-estradiol (E2) secretion, and ovulation rate in a dose-dependent manner. We then examined whether bezafibrate treatment could rescue the inhibition of FSH-induced follicle development and steroidogenesis by TNF. Bezafibrate treatment rescued inhibition of follicle development, secretion of E2, and ovulation rate by TNF. We examined the expression of peroxisome proliferator-activated receptor (PPAR) subtypes in mouse preantral follicles. As the protein expression of only PPARG was observed in mouse preantral follicles, we examined whether bezafibrate could affect follicle development and steroidogenesis through PPARG pathways. Treatment with GW1929, a selective PPARG agonist, restored inhibition of FSH-induced follicle development and steroidogenesis by TNF, whereas treatment with GW9662, a selective PPARG antagonist, canceled the restorative effects of bezafibrate. Collectively, the results in this study suggest that bezafibrate may directly exhibit a restorative effect on the inhibition of ovarian follicle development and steroidogenesis by TNF through the PPARG pathway.

Cellular and molecular mechanisms of various types of oocyte aging.

It is well established that age-related decline of a woman's fertility is related to the poor developmental potential of her gametes. The age-associated decline in female fertility is largely attributable to the oocyte aging caused by ovarian aging. Age-associated oocyte aging results in a decrease in oocyte quality. In contrast to ovarian aging, there is a concept of postovulatory oocyte aging. Postovulatory aging of oocytes, not being fertilized for a prolonged time after ovulation, is known to significantly affect the development of oocytes. Both categories of oocyte aging have similar phenotypes of reproductive failure. However, the mechanisms of the decline in oocyte quality are not necessarily equivalent. An age-dependent increase in aneuploidy is a key determinant of oocyte quality. The reduced expression of molecules regulating cell cycle control during meiosis might be involved in the age-dependent increase in aneuploidy. The mechanism of age-associated oocyte aging might be involved in mitochondrial dysfunction, whose etiologies are still unknown. Alternatively, the mechanism of postovulatory oocyte aging might be involved in reactive oxygen species-induced mitochondrial injury pathways followed by abnormal intracellular Ca2+ regulation of the endoplasmic reticulum. We suggest that future research into the mechanism of oocyte aging will be necessary to develop a method to rescue the poor developmental potential of aged oocytes.

Usefulness of bezafibrate for ovulation induction in clomiphene citrate-resistant polycystic ovary syndrome patients with dyslipidemia: a prospective pilot study of seven cases.

BACKGROUND: Dyslipidemia is commonly observed in polycystic ovary syndrome (PCOS) patients. Bezafibrate is a drug for dyslipidemia acting through peroxisome proliferator-activated receptors. We investigated the effects of bezafibrate for ovulation induction in patients with PCOS with dyslipidemia who were resistant to clomiphene citrate (CC). METHODS: This was a prospective pilot study. Seven infertile, CC-resistant, PCOS patients with dyslipidemia were enrolled in this study. The participants received bezafibrate at 400 mg/day from day 1 of menses and CC at 100 mg/day from day 5 of menses simultaneously until one follicle measuring at least 18 mm in diameter was found by transvaginal ultrasound. The main outcome was ovulation rate. RESULTS: Five of 7 patients successfully ovulated. The mean number of days of menses until the follicle reached 18 mm in diameter was 16 ± 3 (range 13-20). Monofollicular development was observed in all patients that ovulated. One woman became pregnant and delivered a healthy baby. CONCLUSION: Bezafibrate may be effective for ovulation induction in CC-resistant PCOS patients with dyslipidemia.

Y chromosome microdeletion screening using a new molecular diagnostic method in 1030 Japanese males with infertility.

The azoospermia factor (AZF) region is important for spermatogenesis, and deletions within these regions are a common cause of oligozoospermia and azoospermia. Although several studies have reported this cause, the present research, to the best of our knowledge, is the first large-scale study assessing this factor in Japan. In this study, 1030 male patients with infertility who were examined for Y chromosome microdeletion using the polymerase chain reaction-reverse sequence-specific oligonucleotide (PCR-rSSO) method, a newly developed method for Y chromosome microdeletion screening, were included. The study enrolled 250 patients with severe oligospermia and 717 patients with azoospermia. Among the 1030 patients, 4, 4, 10, and 52 had AZFa, AZFb, AZFb+c, and AZFc deletions, respectively. The sperm recovery rate (SRR) of microdissection testicular sperm extraction in patients with AZFc deletions was significantly higher than that in those without AZF deletions (60.0% vs 28.7%, P = 0.04). In patients with gr/gr deletion, SRR was 18.7%, which was lower than that in those without gr/gr deletion, but was not statistically significant. In conclusion, our study showed that the frequency of Y chromosome microdeletion in male patients in Japan was similar to that reported in patients from other countries, and SRR was higher in patients with AZFc deletion.

Alterations of PLCbeta1 in mouse eggs change calcium oscillatory behavior following fertilization.

Inositol 1,4,5-trisphosphate generated by the action of a phospholipase C (PLC) mediates release of intracellular Ca2+ that is essential for sperm-induced activation of mammalian eggs. Much attention currently focuses on the role of sperm-derived PLCzeta in generating changes in egg intracellular Ca2+ despite the fact that PLCzeta constitutes a very small fraction of the total amount of PLC in a fertilized egg. Eggs express several isoforms of PLC, but a role for an egg-derived PLC in sperm-induced Ca2+ oscillations has not been examined. Reducing egg PLCbeta1 by a transgenic RNAi approach resulted in a significant decrease in Ca2+ transient amplitude, but not duration or frequency, following insemination. Furthermore, overexpressing PLCbeta1 by microinjecting a Plcb1 cRNA significantly perturbed the duration and frequency of Ca2+ transients and disrupted the characteristic shape of the first transient. These results provide the first evidence for a role of an egg-derived PLC acting in conjunction with a sperm-derived PLCzeta in egg activation.

Prognostic factors for patients undergoing vitrified-warmed human embryo transfer cycles: a retrospective cohort study.

We examined the prognostic factors for pregnancy in 210 vitrified-warmed embryo transfer (ET) cycles in 121 patients. The univariate analysis showed that age, gravida, the number of cycles associated with infertility caused by endometriosis, the number of previous assisted reproductive technology (ART) treatment cycles, and the number of ICSI procedures were significantly lower in pregnant cycles compared with non-pregnant cycles. The percentages of ET using at least one intact embryo and of ET using at least one embryo that had developed further after warming were significantly higher in pregnant cycles compared with non-pregnant cycles. Multivariate logistic regression analysis showed that previous ART treatment cycles, ET with at least one intact embryo, and ET using at least one embryo that had developed further were independent prognostic factors for pregnancy in vitrified-warmed ET cycles. We conclude that fewer previous ART treatment cycles, ET using at least one intact embryo, and ET with embryos that have developed further after warming might be favourable prognostic factors for pregnancy in vitrified-warmed ET cycles.