Number Of Antral Follicles Research Articles

Altered m6A modification is involved YAP-mediated apoptosis response in 4-vinylcyclohexene diepoxide induced ovotoxicity

4-Vinylcyclohexene diepoxide (VCD), an industrial occupational health hazard chemical associated with premature ovarian insufficiency (POI) and reproductive failure. Recently, investigators have paid an increasing attention on VCD model of menopause recapitulates the natural, physiological transition through perimenopause to menopause. The current study sought to examining the mechanisms of follicular loss and exploring the effect of the model on systems outside of the ovaries. In this study, 28 days female SD rats were injected with VCD (160 mg/kg) vehicle for 15 consecutive days, euthanized in the diestrus phase approximately 100 days after the onset of treatment. Reproductive system injury, Neuroendocrine, sex hormone levels and receptor were observed, the levels of N6-methyladenosine (m6A) RNA modification and the expression of modulator genes were first measured. The VCD treated rats showing irregular estrous cycles, significantly reduced in the number of primordial follicles, the preantral and antral follicles also decreased significantly, accompanied by the plasma level of FSH increased and anti-Mullerian hormone (AMH) were decreased. The total m6A level was significantly decreased after exposure to VCD. Moreover, ALKBH5-mediated YAP m6A modification changed in VCD - induced premature ovarian insufficiency. These present work provides a new perspective on m6A modification in the VCD-induced POI rat model, which could provide valuable insights into the mechanisms underlying follicle development and finding new therapeutic targets for follicle prematurely exhausted. Also provide novel methodological guidance and endocrine basis to guide research and extend the applications in premature ovarian insufficiency model.

The Effects of Coenzyme Q10 on Hormonal and Histopathological Changes in Female Wistar Rats with Polycystic Ovary Syndrome

Background: Polycystic ovary syndrome (PCOS) is one the most common diseases in women in the fertility age. PCOS alters ovarian follicles and affects blood lipid profiles, liver enzymes and hormones. In this study, we tried to investigate the effects of coenzyme Q10 (CoQ10) on PCOS pathological conditions. Methods: Twenty-four female rats were randomly divided into the three groups; 1: control group, 2: PCOS group (received 1 mg/ kg letrozole daily for 21 days, PO) and 3: PCOS+CoQ10 group (received 1 mg/kg letrozole with 10 mg/kg of CoQ10 for 21 days orally). Blood lipid profiles, liver enzymes, blood glucose and testosterone levels were measured and ovarian histopathology was evaluated. Results: Histological examination showed reduced number of antral follicles in PCOS+CoQ10 group as compared with PCOS group (P<0.001). PCOS increased the alanine aminotransferase (ALT) enzyme by acting on the liver (P<0.01). Administration of CoQ10 during PCOS induction was able to reduce ALT levels (P<0.01). PCOS increased the cholesterol (P<0.05), fasting blood sugar (FBS) (P<0.05), triglyceride (P<0.001), low-density lipoprotein (LDL) (P<0.05) and testosterone levels (P<0.001) compared to the control group. However, CoQ10 treatment significantly reduced the above-mentioned parameters compared to the PCOS group. Conclusion: The results of the present study confirm that CoQ10 has a therapeutic potential for PCOS-induced lipid and hormonal changes and ovarian tissue damages. CoQ10 supplementation and its concomitant use with letrozole could inhibit the development of PCOS in rats. Testosterone reduction could be an important mechanism for CoQ10 beneficial effects.

P-615 Ovarian rejuvenation with human placenta hydrolyzate

Abstract Study question Is it possible to rejuvenate the ovaries with human placenta hydrolyzate? Summary answer Injecting human placenta hydrolyzate into the ovaries can activate dormant follicles and produce mature oocytes in patients with ovarian insufficiency. What is known already Patients with low ovarian reserve and/or ovarian failure are a difficult category for reproductive specialists. Attempts to rejuvenate the ovaries have been made for several years. The use of adjuvant therapy such as PRP can activate dormant follicles. Human placenta hydrolyzate contains growth factors, hormones, trace elements, vitamins. Administration of human placenta hydrolyzate directly to the ovaries can also activate folliculogenesis and oogenesis. Study design, size, duration A cohort study was conducted, which involved 20 women with low ovarian reserve who underwent IVF / ICSI at the Person ICCR in 2022. AMH, FSH, the number of antral follicles, the number of received oocytes were determined before the injection of the human placenta hydrolyzate (JAPAN BIO PRODUCTS Co, LTD, Japan) into the ovaries and 1-3 months after procedures. Participants/materials, setting, methods All patients had previously undergone IVF/ICSI. 20 women with infertility, mean age was 39±6 years. In September-October 2022, we received a human placenta hydrolyzate intraovarially. The human placenta hydrolyzate was injected into the ovaries as in a transvaginal puncture with a 20 G needle, 1 ml into each ovary. All participants gave their voluntary informed consent to the procedure. Main results and the role of chance On the 2-3rd day of the cycle before the procedure, the average level of AMH was 0.11±0.09 ng/ml, FSH 38.11±10.86 mIU/ml, the number of antral follicles 1.18±0, 38, the number of mature oocytes obtained on TVP, 0.68±0.19. After the introduction of the human placenta hydrolyzate into the ovaries, measurements were repeated 1-3 months later: AMH was 0.46±0.18 ng/ml, p = 0.09; FSH was 13.54±7.86 mIU/ml, p = 0.074; CAF 1.96±0.17, p = 0.068; oocytes M2 1.51±0.38, p = 0.058. Due to small cohort size and short follow-up time, we believe that ovarian activation by placental hydrolyzate takes place, this is especially noticeable in the number of obtained mature oocytes that went for freezing or fertilization: 4 patients received embryos suitable for PGT-A, one the embryo (mosaic) was transferred into the uterine cavity and pregnancy occurred. There was no any side effect after the injection. Limitations, reasons for caution Limitations relate to study design. More patients and longer follow-up are also needed. Wider implications of the findings Ovarian rejuvenation is currently an adjuvant therapy, high-level evidence is not yet available. However, for women with POI and POR, any opportunity to improve the chances of obtaining their own oocytes is valuable. Trial registration number NO APPLICABLE

O-206 Growth hormone supplementation reduces aneuploidy and improves the quality of aged oocytes by activating the JAK2-ERK1/2 pathway

Abstract Study question Does growth hormone (GH) supplementation improves the quality of oocytes in aged mice via reducing aneuploidy? Summary answer Daily injections of GH for 8 weeks reduces aneuploidy by activating JAK2-ERK1/2 pathway and improves the quality of oocyte in aged mice. What is known already Aging-induced low-quality oocytes, which are mainly caused by aneuploidy, decrease female fertility. However, few treatments are available to improve the poor quality of oocytes in aged women. Age-related decline in GH levels may be related to the insufficient reproductive potential in aged women. Several studies have reported that GH improves assisted reproductive technology outcomes in poor ovarian response patients, although its role is still controversial. In animal models, GH supplementation improved the oocyte quality by enhancing mitochondrial function. However, whether GH reduces the aneuploidy rate in aged oocytes, and if so the underlying mechanisms, are unclear. Study design, size, duration The aged (8-month-old) C57BL/6J female mice were used in the study. For the in vivo experiments, the aged mice were intraperitoneally injected with GH (GenSci, Changchun, China; 1.6 mg/kg body weight) or an equivalent volume of normal saline for 8 weeks; For the in vitro experiments, germinal vesicle (GV) oocytes from aged mice were treated with GH (200 ng/mL) or PBS in M16 medium. Participants/materials, setting, methods We counted oocytes by immunohistochemistry to assess ovarian reserve. The expression of growth hormone receptor and mitochondrial genes were measured by quantitative real-time PCR. Time–lapse incubator was utilized to record the developmental potential of early embryos. Immunofluorescence was performed to assess parameters of oocyte quality (mitochondrial functions, spindle/chromosome defects, and DNA damage). Chromosome spread and DNA sequencing were used to analyze aneuploidy rate. We performed quantitative proteomics analysis to identify the potential targets of GH. Main results and the role of chance Firstly, we found that GH promoted GHR expression in aged oocytes (30.64±1.70 vs 21.68±1.08, P < 0.001). GH ameliorated aging-induced decline in ovarian reserve of aged mice, with increased ovarian index (0.040%±0.0031 vs 0.023%±0.0019, P < 0.01), and number of antral follicles (8.67±1.2 vs 3.33±0.88, P < 0.05) compared to the controls. In addition, GH improved the quality of aged oocytes, representing restored mitochondrial functions (P < 0.05), decreased DNA damage (23.37±1.76 vs 38.35±2.52, P < 0.001) . As expected, GH promoted the fertilization rate (39.9%±1.10 vs 14.4%±0.62, P < 0.001) and early embryo development (P < 0.01) of aged oocytes. Of note, GH recovered the spindle/chromosome defects (42.6%±2.12 vs 63.9%±1.83, P < 0.01, and reduced aneuploidy rate (20.9%±1.61 vs 39.3%±1.84, P < 0.01) in aged oocytes. GH activated ERK1/2 expression to decrease aneuploidy rate in aged oocytes. In addition, JAK2 might be involved in the regulation of GH on ERK1/2 expression in aged oocytes. Altogether, GH restored age-related decline in oocyte number, and decreased the occurrence rate of aneuploidy by activating JAK2-ERK1/2 pathway, thereby improving the quality of aged oocytes. Limitations, reasons for caution The molecular action mechanism of GH to regulate aneuploidy remains to be determined. Besides, future work should be extended to human oocyte to determine whether this mechanism is conserved between mice and humans Wider implications of the findings Our work expounds a theoretical basis for application of GH to improve the fertility of aged women. Besides, the results also feed new ideas for the prevention and treatment of oocyte quality decline in assisted reproductive technology. Trial registration number not applicable

P-684 Subclinical hypothyroidism and ovarian reserve indices in women with normogonadotropic anovulation

Abstract Study question Does subclinical hypothyroidism (SCH) affect ovarian reserve indices in women with Polycystic Ovary syndrome (PCOS) or other forms of Hypothalamic-Pituitary-Ovarian Axis Dysfunction (HPOD)? Summary answer SCH increases Anti-Müllerian hormone (AMH) concentration, but has no influence on Follicle-stimulating hormone (FSH). What is known already In PCOS, anovulation is caused by the arrest of growing follicles due to increased frequency and amplitude of luteinizing hormone (LH) pulses, resulting in increased number of pre-antral and antral follicles, and concentration of AMH. In other cases of HPOD, the causes of anovulation are more diverse and have not yet been clearly identified. Thyroid hormones can modify the function of the HPO axis, affecting the maturation of follicles. While overt hypothyroidism is treated to improve ovulation and fertility, the effect of subclinical hypothyroidism (SCH) and the presence of circulating antithyroid antibodies (ATA) on ovarian function is uncertain. Study design, size, duration A prospective cohort tertiary single-center study (consent no. 1072.6120.172.2022) included women aged 18-45, examined due to menstrual disorders and/or infertility, from July to August 2022. Only women without previously diagnosed thyroid dysfunction were included. All enrolled women gave informed written consent to participate in the study. Participants/materials, setting, methods Women underwent routine gynecological examination and pelvic ultrasound. Blood sample was tested for AMH, FSH, Luteinizing hormone (LH), Thyroid-stimulating hormone (TSH), anti-thyroid peroxidase (anti-TPO) and anti-thyroglobulin (anti-TG) antibodies concentrations. The Rotterdam-ESHRE-ASRM criteria were used to diagnose PCOS and its phenotypes. HPOD was diagnosed according to WHO classification. SCH was defined as TSH > 2.5 uIU/ml with normal thyroid function. The above parameters within subpopulations were calculated using regression analysis, Kruskal-Wallis and post-hoc tests. Main results and the role of chance The study included 51 euthyroid women aged 18-40. PCOS was diagnosed in 42/51 (82.35%) women, including PCOS-A phenotype in 31/51 (60.78%), PCOS-B in 5/51 (9.8%), PCOS-D in 6/51 (11.76%) women, and HPOD in 9/51 (17.65%) women. There was a significant positive correlation between the concentration of TSH and AMH in the studied population - with the increase of TSH value, the concentration of AMH increased (r = 0.4, p = 0.0035). The mean AMH concentration significantly differed between the groups and equaled to 55.4, 39.4, 31.1, 24.0 pmol/l in PCOS-A, PCOS-B, PCOS-D and HPOD (p = 0.05), respectively. The mean anti-TPO concentration was significantly different between the groups and was equal to 18.2, 19.0, 14.6, 10.9 U/ml in PCOS-A, PCOS-B, PCOS-D and HPOD (p = 0.05), respectively. There was no significant difference in FSH, LH, TSH, anti-TG concentrations between the groups. Limitations, reasons for caution The limitations of the study are small study group and single-center nature. Wider implications of the findings SCH increases the concentration of AMH, which in PCOS may exacerbate the symptoms of anovulation. Whether treatment of subclinical hypothyroidism affects ovarian reserve indices and improves ovarian function remain a subject of further research. Trial registration number 1072.6120.172.2022

O-068 Transdermal testosterone gel (TTG) pre-treatment duration in improving ivf outcome in patients with poor prognosis (poseidon group 3 and 4): A randomised controlled trial.

Abstract Study question To compare the most effective duration of pre-treatment with transdermal testosterone gel in improving IVF outcomes using GnRH antagonist protocol in poor prognosis patients. Summary answer Pre-treatment with testosterone gel for 28 days had higher clinical pregnancy rate, more mature oocytes in lesser days and lower total dose of gonadotropins. What is known already Poor prognosis patients undergoing IVF exhibits subnormal ovarian response resulting in higher IVF cycle cancellation and lower clinical pregnancy. Amongst various adjuvants regimens use of androgen supplementation improves clinical pregnancy and probability of live birth in such patients. Androgen stimulates early stages of follicular growth, increase the number of primary, pre-antral and antral follicles. It increases FSH receptor expression in granulosa cells, in turn leading to better oocyte yield and pregnancy rate. We aim to compare the most effective duration of pre-treatment with transdermal testosterone gel in improving IVF outcomes using GnRH antagonist protocol in poor prognosis patients. Study design, size, duration A prospective, randomised controlled trial was carried out from 1st September 2021 to 31st August 2022 at a tertiary infertility centre in India. 100 poor prognosis patients POSEIDON group 3 and 4 (age 21-40 years, AFC <5 and AMH ≤1.2 ng/ml) were enrolled. They were randomized into 4 groups of 25 each by computer based program. Participants/materials, setting, methods Group 1, control received no pretreatment, Group 2, Group 3 and Group 4 received 1% TTG one actuation (12.5mg testosterone daily) for 2, 3 and 4 weeks respectively before start of ovarian stimulation. Patients received a starting dose of HP HMG dose 300 IU and dose was adjusted. Inj. Cetrorelix 0.25 mg started when lead follicle was 14 mm and Oocyte retrievals were performed 35 hours after 250 mcg r-HCG trigger. Main results and the role of chance All groups were comparable with regards to baseline FSH, AFC and AMH levels. Only one cycle was cancelled in control group because of lack of response to gonadotropins till day 5 of stimulation. Requirement of total days (9.68 Vz 8.95 days) and total dose of gonadotropins was significantly lower in group 4 as compared to control group 1. This showed that cases with pre-treatment of transdermal testosterone gel for longer duration, required less amount of gonadotropin (2826 IU Gp 1 Vz 2329 IU Gp 4) treatment (p < 0.01) and also the mean number of mature oocytes retrieved (0.80 Gp 1 Vz 3.38 Gp 4) were significantly higher (p < 0.01). Day 3 fresh embryo transfers were performed for all. Incidence of clinical pregnancy rate was 4.2%, 8%, 12% in group 1, 2, and 3 respectively which increased to 20% in group 4 (p-0.33). Miscarriage rate was 4.2% , 4%, 12% and 4% in group 1 ,2, 3 and 4 respectively (p-0.71). Ongoing pregnancy rate was 4% in group 1, while it was 7.7 %, 10.6 % and 19.2% in group 2,3 and 4 respectively (p-0.33). Limitations, reasons for caution The study was done at a single centre with small sample size, replication with more subjects and in different centers is needed. The long term effects of testosterone pretreatment requires further studies. Wider implications of the findings Pre-treatment with testosterone gel in poor prognosis patients improves response to gonadotrophin, reducing amount and days to complete stimulation and results in higher number of mature oocytes and improved clinical pregnancy rates. Transdermal testosterone treatment is advantageous as it is not first metabolized in the liver, convenient to the patient. Trial registration number CTRI/2022/03/040793

O-244 Mitochondrial fission factor (MFF) is required for female fertility and oocyte development

Abstract Study question To explore the role of Mitochondrial fission factor (MFF) in female fertility and oocyte development using a Mff knockout mouse model. Summary answer Mff deficiency influences mitochondrial dynamics in oocytes at germinal vesicle stage (GV), and results in premature ovarian failure (POF) and female infertility. What is known already Mff regulates mitochondrial quality control (MQC) by influencing mitochondrial dynamics. MQC has critical effects on oocyte development and POF. However, little is known about the role of Mff in oocyte development and female fertility. Study design, size, duration The Mff knockout (Mff-/-) mice were generated via the CRISPR-Cas9 system. Adult Mff-/- female mice were compared to wildtype (WT) female mice. Female mice at 2-month old from each group (Mff-/- or WT, n = 5) were used to assess the fecundity for 10 months. The follicle development (n = 4 mice), the number of GV oocytes (n = 5 mice) and metaphase II (MII) oocytes (n = 5 mice) were determined. Participants/materials, setting, methods Female mice from each group were mated with adult WT males to assess the fecundity. Follicle development was evaluated in ovaries after fixation, paraffin embedding, and sectioning, followed by hematoxylin and eosin staining. MFF and DRP1 protein expression was evaluated by western blot. Oocyte mitochondria were stained with the Mito-Tracker Red CMXRos. Main results and the role of chance The Mff deficient ovaries had significantly lower levels of MFF (0.056 vs. 1, p < 0.01) than the WT ovaries. Mff-/- female mice were infertile (0 vs. 39, p < 0.05). Mature (8-week-old) Mff deficient ovaries had significantly lower weight (0.0043 vs. 0.011, p < 0.05) compared to the WT. The 2-month-old female Mff-/- mice had a lower number of primordial (220 vs. 775, p< 0.05), primary (128 vs. 580, p< 0.05), secondary (29 vs. 82, p< 0.05) and antral follicles (5 vs. 23, p< 0.05). The numbers of GV oocytes (18 vs. 37, p< 0.05) and MII oocytes (2 vs. 16, p< 0.001) were lower in the Mff-/- mice than the WT mice. In addition, the Mff-/- ovaries expressed significantly less mitochondrial fission key regulator DRP1 (0.54 vs. 1, p< 0.05). Mff-/- oocytes contained elongated and aggregated mitochondria compared to the discrete and evenly distributed mitochondria in the ooplasm of WT oocytes. Herein, we found that the Mff played a concerted role in oocyte and follicle development. Limitations, reasons for caution The present observations in mice may not be applicable to humans. The results are derived global knockout of Mff. It is not known which kind of cell plays a vital role in fertility. Further investigation is needed to explore the regulatory mechanism of Mff in oocyte development and fertility regulation. Wider implications of the findings This study provides important insights into the relationship between mitochondrial dynamics and female fertility. Trial registration number not applicable

Ovarian morphology and follicular dynamics associated with ovarian aging in Bos indicus beef cows

This study characterizes the ovarian volume, antral follicle count (AFC), and ovarian dynamics of cows at different ages. Nellore cows (Bos taurus indicus) were used in two experiments. In Experiment 1, 57 lactating cows ranging in age from 3 to 23 years had their estrous cycle synchronized based on progesterone/estradiol treatments. The ovaries were scanned by ultrasound to record ovarian volume and AFC prior to follicle aspiration (AFC1). A second AFC (AFC2) was performed 5 days later to count only the growing antral follicles. In Experiment 2, six long-lived (14- to 23-year-old) and three young (4- to 8-year-old) non-lactating cows were submitted to daily ovarian scanning ultrasound during an interovulatory interval. Blood samples were collected during the estrous cycle to assess serum progesterone concentration. Data were analyzed using Student's t-test, ANOVA, and regression analysis tests. In Experiment 1, there were more (P < 0.05) antral follicles in AFC1 (31.4 ± 3.5) than in AFC2 (22.6 ± 2.4). In AFC1, the volume of the right ovary (6.03 ± 0.5 cm3) was greater (P < 0.01) than that of the left one (4.53 ± 0.4 cm3), although the AFC did not differ between the two ovaries (15.3 ± 1.8 and 16.3 ± 1.8, respectively). In both AFC1 and AFC2, there was a decline in the number of antral follicles as the age of the cow increased (P < 0.01). Ovarian volume (average of both ovaries) was related (P < 0.01) with AFC (R² = 0.1499) and cow age (R² = 0.0911). In Experiment 2, young and old cows under the age of 20 had a pattern of follicular growth waves, while cows over 20 years old did not have waves of follicular growth. The progesterone profiles and corpus luteum size during the estrous cycle did not differ between the groups. In conclusion, although cows had increased ovarian volume and decreased follicular population as they age, the follicular growth pattern and corpus luteum functionality appear to be unaffected by age.

Different types of bisphenols alter ovarian steroidogenesis: Special attention to BPA

Endocrine disruptors such as bisphenol A (BPA) and some of its analogues, including BPS, BPAF, and BPE, are used extensively in the manufacture of plastics. These synthetic chemicals could seriously alter the functionality of the female reproductive system. Although the number of studies conducted on other types of bisphenols is smaller than the number of studies on BPA, the purpose of this review study was to evaluate the effects of bisphenol compounds, particularly BPA, on hormone production and on genes involved in ovarian steroidogenesis in both in vitro (human and animal cell lines) and in vivo (animal models) studies.The current data show that exposure to bisphenol compounds has adverse effects on ovarian steroidogenesis. For example, BPA, BPS, and BPAF can alter the normal function of the hypothalamic-pituitary-gonadal (HPG) axis by targeting kisspeptin neurons involved in steroid feedback signals to gonadotropin-releasing hormone (GnRH) cells, resulting in abnormal production of LH and FSH. Exposure to BPA, BPS, BPF, and BPB had adverse effects on the release of some hormones, namely 17-β-estradiol (E2), progesterone (P4), and testosterone (T). BPA, BPE, BPS, BPF, and BPAF are also capable of negatively altering the transcription of a number of genes involved in ovarian steroidogenesis, such as the steroidogenic acute regulatory protein (StAR, involved in the transfer of cholesterol from the outer to the inner mitochondrial membrane, where the steroidogenesis process begins), cytochrome P450 family 17 subfamily A member 1 (Cyp17a1, which is involved in the biosynthesis of androgens such as testosterone), 3 beta-hydroxysteroid dehydrogenase enzyme (3β-HSD, involved in the biosynthesis of P4), and cytochrome P450 family 19 subfamily A member 1 (Cyp19a1, involved in the biosynthesis of E2). Exposure to BPA, BPB, BPF, and BPS at prenatal or prepubertal stages could decrease the number of antral follicles by activating apoptosis and autophagy pathways, resulting in decreased production of E2 and P4 by granulosa cells (GCs) and theca cells (TCs), respectively. BPA and BPS impair ovarian steroidogenesis by reducing the function of some important cell receptors such as estrogens (ERs, including ERα and ERβ), progesterone (PgR), the orphan estrogen receptor gamma (ERRγ), the androgen receptor (AR), the G protein-coupled estrogen receptor (GPER), the FSHR (follicle-stimulating hormone receptor), and the LHCGR (luteinizing hormone/choriogonadotropin receptor). In animal models, the effects of bisphenol compounds depend on the type of animals, their age, and the duration and dose of bisphenols, while in cell line studies the duration and doses of bisphenols are the matter.

Brown adipose tissue-derived exosomes delay fertility decline in aging mice.

Ovarian aging has steadily grown to be a significant health issue for women as a result of the increase in average life expectancy and the postponement of reproductive age. One of the important pathological foundations of ovarian aging is formed by mitochondrial dysfunction, which causes decreases in follicle quantity and oocyte quality. In recent years, brown adipose tissue (BAT) transplantation has been proven as an effective treatment for aging-related diseases, such as ovarian aging. However, BAT transplantation is an invasive operation with long-term risks. Therefore, we need to find an alternative strategy. We injected BAT-derived exosomes into eight-month-old C57BL/6 female mice. The fertility was detected by the estrous cycle and mating test. The changes of ovary and oocyte were measured by ovarian volume, organ coefficient, follicle counting, and oocyte maturation rate. ROS level, mitochondrial membrane potential and ATP level were measured to analyze the mitochondrial function of oocytes. The changes in metabolism were explored by cold stimulation test, body weight and blood sugar. The possible molecular mechanism was further investigated by RNA sequencing. In terms of fertility, the estrous cycle of aging mice after BAT-derived exosome intervention was more regular, and the number of progenies and litters was increased. At the tissue level, the ovaries in the BAT-exosome group were larger, and the number of primordial follicles, secondary follicles, antral follicles and total follicles increased. At the cellular level, BAT-derived exosomes improved the maturation of oocytes in vivo and in vitro, increased the mitochondrial membrane potential and ATP levels of oocytes, and decreased ROS levels. Besides, BAT-derived exosomes ameliorated the metabolism and viability of aging mice. Furthermore, mRNA sequencing showed that BAT exosomes altered the expression levels of genes related to metabolism and the quality of oocytes. BAT-derived exosomes enhanced mitochondrial function, promoted follicle survival, improved fertility, and extended ovarian lifespan in aging mice.

Chemotherapy impairs ovarian function through excessive ROS-induced ferroptosis

Chemotherapy was conventionally applied to kill cancer cells, but regrettably, they also induce damage to normal cells with high-proliferative capacity resulting in cardiotoxicity, nephrotoxicity, peripheral nerve toxicity, and ovarian toxicity. Of these, chemotherapy-induced ovarian damages mainly include but are not limited to decreased ovarian reserve, infertility, and ovarian atrophy. Therefore, exploring the underlying mechanism of chemotherapeutic drug-induced ovarian damage will pave the way to develop fertility-protective adjuvants for female patients during conventional cancer treatment. Herein, we firstly confirmed the abnormal gonadal hormone levels in patients who received chemotherapy and further found that conventional chemotherapeutic drugs (cyclophosphamide, CTX; paclitaxel, Tax; doxorubicin, Dox and cisplatin, Cis) treatment significantly decreased both the ovarian volume of mice and the number of primordial and antral follicles and accompanied with the ovarian fibrosis and reduced ovarian reserve in animal models. Subsequently, Tax, Dox, and Cis treatment can induce the apoptosis of ovarian granulosa cells (GCs), likely resulting from excessive reactive oxygen species (ROS) production-induced oxidative damage and impaired cellular anti-oxidative capacity. Thirdly, the following experiments demonstrated that Cis treatment could induce mitochondrial dysfunction through overproducing superoxide in GCs and trigger lipid peroxidation leading to ferroptosis, first reported in chemotherapy-induced ovarian damage. In addition, N-acetylcysteine (NAC) treatment could alleviate the Cis-induced toxicity in GCs by downregulating cellular ROS levels and enhancing the anti-oxidative capacity (promoting the expression of glutathione peroxidase, GPX4; nuclear factor erythroid 2-related factor 2, Nrf2 and heme oxygenase-1, HO-1). Our study confirmed the chemotherapy-induced chaotic hormonal state and ovarian damage in preclinical and clinical examination and indicated that chemotherapeutic drugs initiated ferroptosis in ovarian cells through excessive ROS-induced lipid peroxidation and mitochondrial dysfunction, leading to ovarian cell death. Consequently, developing fertility protectants from the chemotherapy-induced oxidative stress and ferroptosis perspective will ameliorate ovarian damage and further improve the life quality of cancer patients.

The effects of decreased ovarian reserve on in vitro fertilization outcome

Objective: In this study, we aimed to compare the effects of microdose and antagonist regimens on IVF-ICSI outcome. &#x0D; Material and Methods: The study was conducted at the XXXX University Medical Faculty Training and Research Hospital IVF Clinic during the period of 2018 and 2022. The files of the patients were reviewed retrospectively and age, duration of infertility, basal FSH level, body mass index, antral follicle number (AFC), antimullerian hormon (AMH) level, retrieved oocyte number, metaphase 2 oocyte number, fertilization and pregnancy rates were evaluated. The patients in Group 1 (n=124) were treated by using gonadotropin releasing hormone (GnRH) microdose protocol and in Group 2 (n=136) were treated by using antagonist protocol. &#x0D; Results: There was no significant difference between the groups in terms of the characteristics of the patients such as age, infertility duration, basal FSH level body mass index, AFC and AMH level. The retrieved oocyte number, metaphase 2 oocyte number, fertilization rates and pregnancy rates were also similar.&#x0D; Conclusion: Our results demonstrated that microdose and antagonist regimens have similar effect in poor reponder women on IVF-ICSI outcome.

Evaluation of ovarian reserves in women suffering from an autoimmune disease vitiligo.

In this study, we aimed to investigate whether vitiligo, an autoimmune disease, affects the ovarian reserves of women suffering from this disease. This case-control study included 27 vitiligo patients and 44 healthy participants with regular menstrual cycles. The total number of participants was 71. We conducted the study in a tertiary hospital between June 2022 and November 2022. We carried out the study after receiving the Ethics Committee Approval of the same institute, numbered KAEK/2022.04.88. Before the study, we obtained informed consent from all the participants included in our study. We complied with the Declaration of Helsinki at all stages of the study. We compared the study groups' demographic information, hormonal parameters, ovarian volumes, and antral follicle numbers. We performed an independent t-test to compare group means. We used SPSS for Windows 24.0 (IBM Corp., Armonk, NY, USA) for the analyses. We considered the p-value lower than 0.05 to be statistically significant. AMH values were 2.66±1.76 pmol/L in the control group and 1.61±0.86 pmol/L in the patient group. The total number of antral follicles was 10.25±2.13 (n) in the control group and 9.26±2.97 (n) in the study group. Ovarian volume was 11.57±1.37 ml in the control group and 10.63±1.96 ml in the study group. These results were statistically significantly different. We detected one premature ovarian failure (POI) in the study group. Although there was only one case, this difference between the groups was statistically significant. Our study has proven that the ovaries of women of reproductive age suffering from any disease with an autoimmune etiology are affected by this autoimmune process. Therefore, ovarian reserves should be evaluated in all fertile women with autoimmune diseases. Due to the evaluation, appropriate treatment and follow-up plans should be made in patients with infertility or premature ovarian failure (POF) risk.

Hipogonadotropik Hipogonadizmli Kadınlarda İn Vitro Fertilizasyon Sonuçlarının Değerlendirilmesi

Aim: Hypogonadotropic hypogonadism (HH) is a rare clinical condition resulting from gonadal insufficiency due to low pituitary gonadotropin levels. Since ovulation occurs rarely in these patients, the probability of spontaneous pregnancy is very low. The study aimed to evaluate the in vitro fertilization (IVF) treatment outcomes in patients with HH and to compare these results with that of patients with unexplained infertility (UI) who underwent IVF treatment. Material and Methods: In this study, 36 cycles of 28 HH patients who underwent IVF treatment and 72 cycles of 68 patients who underwent IVF treatment for UI were included. Demographic data, ovarian hyperstimulation and cycle outcomes, clinical pregnancy rates, and predictive factors for clinical pregnancy were evaluated retrospectively, and the two groups were compared. Results: In the HH group, clinical pregnancy, and live birth rates per cycle after IVF were significantly higher compared with the UI group (n=16, 44.4% vs. n=17, 23.6%, p=0.027; and n=14, 38.9% vs. n=14, 19.4%; p=0.030, respectively). Although the number of antral follicles (p=0.001) and retrieved oocytes (p=0.042) were significantly higher in the UI group, the number of mature oocytes and grade I-II embryos were similar in the HH and UI groups. The total gonadotropin dose used and duration of stimulation in the HH group were significantly higher than in the UI group (both p=0.001). Conclusion: HH patients responded well to IVF treatment and had better IVF outcomes compared to women who underwent IVF for UI. No prognostic factor that affected pregnancy success in HH patients was detected.

Zebrafish ambra1b knockout reveals a novel role for Ambra1 in primordial germ cells survival, sex differentiation and reproduction

BackgroundAMBRA1 is an intrinsically disordered protein, working as a scaffold molecule to coordinate, by protein-protein interaction, many cellular processes, including autophagy, mitophagy, apoptosis and cell cycle progression. The zebrafish genome contains two ambra1 paralogous genes (a and b), both involved in development and expressed at high levels in the gonads. Characterization of the zebrafish paralogous genes mutant lines generated by CRISPR/Cas9 approach showed that ambra1b knockout leads to an all-male population.ResultsWe demonstrated that the silencing of the ambra1b gene determines a reduction of primordial germ cells (PGCs), a condition that, in the zebrafish, leads to the development of all-male progeny. PGC reduction was confirmed by knockdown experiments and rescued by injection of ambra1b and human AMBRA1 mRNAs, but not ambra1a mRNA. Moreover, PGC loss was not rescued by injection with human AMBRA1 mRNA mutated in the CUL4-DDB1 binding region, thus suggesting that interaction with this complex is involved in PGC protection from loss. Results from zebrafish embryos injected with murine Stat3 mRNA and stat3 morpholino suggest that Ambra1b could indirectly regulate this protein through CUL4-DDB1 interaction. According to this, Ambra1+/− mice showed a reduced Stat3 expression in the ovary together with a low number of antral follicles and an increase of atretic follicles, indicating a function of Ambra1 in the ovary of mammals as well. Moreover, in agreement with the high expression of these genes in the testis and ovary, we found significant impairment of the reproductive process and pathological alterations, including tumors, mainly limited to the gonads.ConclusionsBy exploiting ambra1a and ambra1b knockout zebrafish lines, we prove the sub-functionalization between the two paralogous zebrafish genes and uncover a novel function of Ambra1 in the protection from excessive PGC loss, which seems to require binding with the CUL4-DDB1 complex. Both genes seem to play a role in the regulation of reproductive physiology.

BDNF promotes mouse follicular development and reverses ovarian aging by promoting cell proliferation

BackgroundBrain-derived neurotrophic factor (BDNF) plays an important role in ovarian function including follicle development and oocyte maturation, and embryonic development. However, whether BDNF treatment can reimpose ovarian aging and impaired fertility remains elusive. In this study, we investigated the reproductive outcomes of BDNF treatment and potential mechanisms in aged mice.Method“Aged” mice (35–37 weeks old, n = 68) were treated with recombinant human BDNF protein (rhBDNF, 1 µg/200 µL) through daily intraperitoneal (IP) injection for 10 days with/without ovulation induction. Reproductive age mice (8–10 weeks old, n = 28) were treated with ANA 12 (a selective BDNF receptor, TrkB antagonist) through daily IP injection for 5 days with/without ovulation induction. Ovarian function was assessed by ovarian weight, number of follicles, and sex hormone productions. Following induction of ovulation, the number of total oocytes or abnormal oocytes, and blastocyst formation were assessed. Reproductive functions of the mice were evaluated, including pregnancy rate, mating duration for conception, implantation sites, litter size, and weight of offspring. Finally, the molecular mechanism of the effects of BDNF on ovarian cell functions in mice were examined by Western blot and immunofluorescence.ResultsrhBDNF treatment increased the ovarian weight, number of follicles, number and quality of oocytes including increased blastocysts formation, blood estrogen levels, and pregnancy rate in 35-37-week-old mice. Conversely, BDNF receptor antagonist, ANA 12, treatment decreased the ovarian volume and number of antral follicles and increased the proportion of abnormal oocytes in 8-10-week-old mice. We further demonstrated that BDNF treatment promoted ovarian cell proliferation as well as activation of TrkB and cyclinD1-creb signalling.ConclusionWe demonstrated that ten consecutive days of daily IP injection of rhBDNF rescued ovarian function in aged mice. Our results further indicate that TrkB and cyclin D1-creb signaling may underlie the BDNF function in ovaries. Targeting BDNF-TrkB signaling is a potential novel therapeutic strategy to reverse ovarian aging.

Participation of LIF in follicular development and steroidogenesis in rat ovaries.

Leukaemia inhibitory factor (LIF) is a cytokine belonging to the IL-6 family that is important at the reproductive level in the uterine implantation process. However, there is very little evidence regarding its effect at the ovarian level. The aim of this work was to study the local involvement of the LIF/LIFRβ system in follicular development and steroidogenesis in rat ovaries. To carry out this research, LIF/LIFR/GP130 transcript and protein levels were measured in fertile and sub-fertile rat ovaries, and in vitro experiments were performed to assess STAT3 activation. Then, in in vivo experiments, LIF was administered chronically and locally for 28 days to the ovaries of rats by means of an osmotic minipump to enable us to evaluate the effect on folliculogenesis and steroidogenesis. It was determined by q-PCR and WB that LIF and its receptors are present in fertile and sub-fertile ovaries, and that LIF varies during the oestrous cycle, being higher during the oestrus and meta/dioestrus stages. In addition to this, it was found that LIF can activate STAT3 pathways and cause pSTAT3 formation. It was also observed that LIF decreases the number and size of preantral and antral follicles without altering the number of atretic antral follicles, and can increase the number of corpora lutea, with a notable increase in the levels of progesterone. It is therefore possible to infer that LIF exerts an important effect in vivo on folliculogenesis, ovulation and steroidogenesis, specifically the synthesis of progesterone.

Lovastatin, an Up-Regulator of Low-Density Lipoprotein Receptor, Enhances Follicular Development in Mouse Ovaries.

Ovarian aging hampers in vitro fertilization in assisted reproductive medicine and has no cure. Lipoprotein metabolism is associated with ovarian aging. It remains unclear how to overcome poor follicular development with aging. Upregulation of the low-density lipoprotein receptor (LDLR) enhances oogenesis and follicular development in mouse ovaries. This study investigated whether upregulation of LDLR expression using lovastatin enhances ovarian activity in mice. We performed superovulation using a hormone and used lovastatin to upregulate LDLR. We histologically analyzed the functional activity of lovastatin-treated ovaries and investigated gene and protein expression of follicular development markers, using RT-qPCR and Western blotting. Histological analysis showed that lovastatin significantly increased the numbers of antral follicles and ovulated oocytes per ovary. The in vitro maturation rate was 10% higher for lovastatin-treated ovaries than for control ovaries. Relative LDLR expression was 40% higher in lovastatin-treated ovaries than in control ovaries. Lovastatin significantly increased steroidogenesis in ovaries and promoted the expression of follicular development marker genes such as anti-Mullerian hormone, Oct3/4, Nanog, and Sox2. In conclusion, lovastatin enhanced ovarian activity throughout follicular development. Therefore, we suggest that upregulation of LDLR may help to improve follicular development in clinical settings. Modulation of lipoprotein metabolism can be used with assisted reproductive technologies to overcome ovarian aging.

Evaluation of the relationship between ovarian reserve with congenital anomalies and intramural uterine leiomyoma among infertile women: a cross-sectional study

BackgroundOvarian reserve is a crucial indicator of a woman’s fertility potential, which is determined by the quality and quantity of antral follicles and oocytes. However, certain factors such as endometriosis, pelvic inflammatory disease, myoma, and the natural process of aging can lead to a poor ovarian response to stimulation, reducing a woman’s chances of conceiving.ObjectiveTo evaluate the effect of uterus congenital anomalies and uterine leiomyoma are associated on ovarian reserve.MethodsThe present cross-sectional study was performed on 321 infertile women in three groups consisted of 97 infertile women with intramural uterine leiomyoma and 81 infertile women with uterine anomalies and 143 infertile women without uterine anomalies and uterine leiomyoma during 2017–2019 in Royan Center. Sampling method was continuous and available. Data collection tool in this study was a questionnaire which was in two parts of individual variables and the second part was related to ultrasound results (number of antral follicles and ovarian volume) and laboratory tests (Anti-Mullerian Hormone (AMH) and Follicle-stimulating Hormone (FSH)). Ovarian reserve parameters were measured in three groups on the third day of the cycle in both groups. Data analysis was performed using SPSS software version 21. Quantitative variables were analyzed using t-test, qualitative variables were analyzed using chi-square test.ResultsThe results of in laboratory parameters showed that there was no statistically significant difference between the three groups in FSH (2.35 ± 1.55, 2.07 ± 1.81, 2.31 ± 1.93) and AMH (6.84 ± 2.75,7.52 ± 3.14,6.93 ± 3.04), respectively (P > 0.05). The results of sonographic variables also showed that the variables include number of antral follicles in right ovarian, number of antral follicles in left ovarian have statistically significant between the three groups (5.73 ± 2.69,4.84 ± 3.14,6.66 ± 3.13), respectively (P < 0.05).ConclusionThe results of the present study showed that uterine abnormalities and uterine leiomyoma with different mechanisms such as reduce of antral follicle numbers and the effect on uterine and ovarian blood flow lead to a decrease in ovarian reserve and infertility. Therefore, treatment and surgery can reduce these effects and improve the fertility of the affected women.

The influence of reproductively significant autoantibodies determined in the follicular fluid on the quality of the obtained oocytes and embryos, as well as on the implantation rate in assisted reproductive technology cycles

BACKGROUND: According to reports, the efficiency of in vitro fertilization and intra cytoplasmic sperm injection protocols is decreased in patients positive for various autoantibodies, as opposed to autoantibody negative patients. However, there are contradictory data indicating no autoantibody effect on the outcome of infertility treatment using assisted reproductive technology.&#x0D; AIM: The aim of this study was to evaluate the embryological outcome and clinical efficiency of infertility treatment in in vitro fertilization and intra cytoplasmic sperm injection protocols in women in the presence of reproductively significant autoantibodies.&#x0D; MATERIALS AND METHODS: This prospective study enrolled 90 infertile patients undergoing assisted reproductive technology treatment. The follicular fluid obtained on the day of oocyte retrieval was evaluated for a wide autoantibody panel using commercial ELISA kits. The main group (n = 52) included women with autoantibody levels determined in the follicular fluid of more than three standard deviations from the mean values determined among all patients. The comparison group consisted of 38 women with autoantibody levels of less than three standard deviations from all subjects. The intergroup comparative analysis included clinical and anamnestic data, hormonal parameters, ovarian reserve, embryological data, and in vitro fertilization and intra cytoplasmic sperm injection efficiency.&#x0D; RESULTS: Reliably lower ovarian reserve parameters (anti-Mullerian hormone levels 1.9 (1.4; 4.0) vs. 3.3 (2.2; 6.5) ng/ml; p = 0.005; number of antral follicles 8.5 (6.0; 12.0) vs. 11.0 (9.0; 17.0); p = 0.003) have been noted in the main group relatively to the comparison group. The autoantibodies to thyroid peroxidase and cardiolipin content in the follicular fluid has been shown to be negatively associated with the number of two-pronuclear zygotes, the presence of autoantibodies to aromatase correlating negatively with the fertilization rate. Furthermore, the follicular fluid levels of autoantibodies to thyroid peroxidase (105 IU/ml) and cardiolipin (5.1 IU/ml) are reliably associated with a higher frequency of a suboptimal response to previous controlled ovarian stimulation, a lower incidence of high quality embryos on days 3 and 4 of in vitro cultivation, a decreased number of top-quality blastocysts, and the clinical efficiency of in vitro fertilization and intra cytoplasmic sperm injection programs.&#x0D; CONCLUSIONS: The presence of reproductively significant autoantibodies is an independent risk factor for reducing the total efficiency of assisted reproductive technology treatment.