Ovarian Growth Research Articles

Mast Cells Retard Tumor Growth in Ovarian Cancer: Insights from a Mouse Model.

Ovarian cancer has the highest mortality rate among female reproductive tract malignancies. A complex network, including the interaction between tumor and immune cells, regulates the tumor microenvironment, survival, and growth. The role of mast cells (MCs) in ovarian tumor pathophysiology is poorly understood. We aimed to understand the effect of MCs on tumor cell migration and growth using in vitro and in vivo approaches. Wound healing assays using human tumor cell lines (SK-OV-3, OVCAR-3) and human MCs (HMC-1) were conducted. Murine ID8 tumor cells were injected into C57BL6/J wildtype (WT) and MC-deficient C57BL/6-KitW-sh/W-sh (KitW-sh) mice. Reconstitution of KitW-sh was performed by the transfer of WT bone marrow-derived MCs (BMMCs). Tumor development was recorded by high-frequency ultrasonography. In vitro, we observed a diminished migration of human ovarian tumor cells upon direct or indirect MC contact. In vivo, application of ID8 cells into KitW-sh mice resulted in significantly increased tumor growth compared to C57BL6/J mice. Injection of BMMCs into KitW-sh mice reconstituted MCs and restored tumor growth. Our data show that MCs have a suppressive effect on ovarian tumor growth and may serve as a new therapeutic target.

Retraction Note: Anhuienoside C inhibits human ovarian cancer cell growth by inducing apoptosis, suppression of cell migration and invasion, and targeting PI3K/AKT/mTOR signaling pathway.

Retraction Note: Anhuienoside C inhibits human ovarian cancer cell growth by inducing apoptosis, suppression of cell migration and invasion, and targeting PI3K/AKT/mTOR signaling pathway.

A pilot in vivo study: potential ovarian cancer therapeutic by placental extracellular vesicles.

The biological links between cancer and pregnancy are of interest due to parallel proliferative, immunosuppressive, and invasive mechanisms between tumour and placental cells. However, the proliferation and invasion of placental cells are strictly regulated. The understanding of this regulation is largely unknown. Placental extracellular vesicles (EVs) may play an important role in this regulation, as placental EVs are known to contribute to maternal adaptation, including adaptation of the vascular and immune systems. We have previously reported that placental EVs significantly inhibited ovarian cancer cell proliferation by delaying the progression of the cell cycle. We therefore performed this pilot in vivo study to investigate whether placental EVs can also inhibit ovarian tumour growth in a SKOV-3 human tumour xenograft model. A single intraperitoneal injection of placental EVs at 15 days post tumour implantation, significantly inhibited the growth of the tumours in our in vivo model. Signs of cellular necrosis were observed in the ovarian tumour tissues, but not in other organs collected from mice that had been treated with placental EVs. Expression of receptor-interacting kinase 1 (RIPK1) and mixed linkage kinase domain-like (MLKL), which are mediators of necroptosis were not observed in our xenografted tumors. However, extensive infiltration of CD169+ macrophages and NK cells in ovarian tumour tissues collected from placental micro-EVs treated mice were observed. We demonstrate here that inhibition of ovarian tumour growth in our xenograft model by placental EVs involves cellular necrosis and infiltration of CD169+ macrophages and NK cells into the tumour tissues.

A Crosstalk Between Pineal and Major Extra-Pineal Sources of Melatonin and its Role in Ovarian Growth and Maturation in Fish

Pinealocytes of the pineal gland in vertebrates mainly synthesize melatonin (5-methoxy-N-acetyl-tryptamine). Moreover, melatonin is synthesized in several extra-pineal cells, including the photoreceptor cells of the retina, the cells of the gut, and the hepatocytes of the liver in different vertebrates, including fish species. One of the remarkable features of pineal and retinal melatonin is that it is produced rhythmically in synchronization with the environmental Light-Dark (LD) cycle, with a daily nighttime peak. However, the melatonin synthesis in tissue/cells from the extra-pineal and extra-retinal origin(s) may not always undergo photoperiod-regulated daily variations but is also dependent on the environmental food entrainment factors (in the gut), acting as the most reliable synchronizer(s) in its daily rhythm features. Moreover, the regulation of the liver and ovary (important for fish reproduction) is unclear. In this review, we attempt a comparative account of the nature and regulation of endogenous melatonin synthesis between a source like the pineal gland and many other nonpineal origins, which have gained serious attention in the last ten years. We also review the functions of melatonin in regulating fish ovarian growth and maturation. The physiological melatonin levels, manipulated either endogenously (by photoperiodic modulations) or exogenously (by injections or by feeds), have tremendous effects on reproductive events in fish at the age of its first maturity, as revealed in recent findings. Characterization and identification of the importance of pineal gland melatonin in the growth of the oocytes via the hypothalamic-pituitary-gonadal axis have been explored several years back. The identification of melatonin receptors about fourteen years back on the wall of developing oocyte spurt the breakthrough, which introduced the concept of direct control of melatonin on developing oocytes. Thus, this review gains uniqueness by addressing the latest developments recorded in the field of melatonin and fish reproduction, particularly in improving oocyte maturation. Nonetheless, an attempt has been made to underline approaches that need to be developed to apply the molecule in large-scale aquaculture.

Expression of genes regulating cell division in porcine follicular granulosa cells

BackgroundCell cycle regulation influences the proliferation of granulosa cells and affects many processes related to ovarian folliclular growth and ovulation. Abnormal regulation of the cell cycle can lead to many diseases within the ovary. The aim of this study was to describe the expression profile of genes within granulosa cells, which are related to the formation of the cytoskeleton, organization of cell organelles inside the cell, and regulation of cell division. Established in vitro primary cultures from porcine ovarian follicle granulosa cells were maintained for 48, 96, 144 h and evaluated via microarray expression analysis.ResultsAnalyzed genes were assigned to 12 gene ontology groups "actin cytoskeleton organization", "actin filament organization", "actin filament—based process", "cell—matrix adhesion", "cell—substrate adhesion", "chromosome segregation", "chromosome separation", "cytoskeleton organization", "DNA integrity checkpoint", "DNA replication initiation", "organelle fision", "organelle organization". Among the genes with significantly changed expression, those whose role in processes within the ovary are selected for consideration. Genes with increased expression include (ITGA11, CNN1, CCl2, TPM2, ACTN1, VCAM-1, COL3A1, GSN, FRMD6, PLK2). Genes with reduced expression inlcude (KIF14, TACC3, ESPL1, CDC45, TTK, CDC20, CDK1, FBXO5, NEK2—NIMA, CCNE2). For the results obtained by microarray expressions, quantitative validation by RT-qPCR was performed.ConclusionsThe results indicated expression profile of genes, which can be considered as new molecular markers of cellular processes involved in signaling, cell structure organization. The expression profile of selected genes brings new insight into regulation of physiological processes in porcine follicular granulosa cells during primary in vitro culture.

C1632 inhibits ovarian cancer cell growth and migration by inhibiting LIN28 B/let-7/FAK signaling pathway and FAK phosphorylation

The highly conserved RNA-binding protein LIN28B and focal adhesion kinase (FAK) are significantly upregulated in ovarian cancer (OC), serving as markers for disease progression and prognosis. Nonetheless, the correlation between LIN28B and FAK, as well as the pharmacological effects of the LIN28 inhibitor C1632, in OC cells have not been elucidated. The present study demonstrates that C1632 significantly reduced the rate of DNA replication, arrested the cell cycle at the G0/G1 phase, consequently reducing cell viability, and impeding clone formation. Moreover, treatment with C1632 decreased cell-matrix adhesion, as well as inhibited cell migration and invasion. Further mechanistic studies revealed that C1632 inhibited the OC cell proliferation and migration by concurrently inhibiting LIN28 B/let-7/FAK signaling pathway and FAK phosphorylation. Furthermore, C1632 exhibited an obvious inhibitory effect on OC cell xenograft tumors in mice. Altogether, these findings identified that LIN28 B/let-7/FAK is a valuable target in OC and C1632 is a promising onco-therapeutic agent for OC treatment.

Characteristics Sufferer Ovarian Cancer at Ibnu Sina Hospital, Makassar

Ovarian cancer is condition caused by abnormal tissue that growth in ovary . Ovary is female reproductive organs where eggs are produced. There is three Ovarian cancer is classified as an epithelial tumor that develops on the surface of the ovary, originating germ cell tumors from in producing cells egg can found in women young, and tumors of stromal origin from cells that have hormone womanhood namely estrogen and progesterone. Objective to find out the characteristics of ovarian cancer sufferers at Ibnu Sina Makassar Hospital. Method this research is a descriptive research with design studies cross sectional , the researcher will collect secondary data through record medical patient ovarian cancer at Ibnu Sina Makassar Hospital. The results shows that from 90 samples, characteristics patients based on age the most is 46-55 years totaling 29 people (32.2%), marital status is already married totaling 76 people (84.4%), symptoms clinical is stomach grow totaling 55 people (61.1%), stage of cancer is stage IIIC totaling 32 people (35.6%), type histopathology is an epithelial tumor surface totaling 73 people (81.1%), and most abdominal ultrasound images on smooth tumor walls totaling 48 people (53.3%), no septate totaling 37 people (41.1%), echogenicity mixed echoic totaling 38 people (42.2%). Can concluded that characteristics the most found at the age of 46-55 years, marital status is already married, symptom clinical is stomach enlarged, cancerous stage is stage IIIC, type histopathology is an epithelial tumor surface, and abdominal ultrasound image of the smooth tumor wall, no septate, echogenicity mixed echoic.

PPARγ activation modulates the balance of peritoneal macrophage populations to suppress ovarian tumor growth and tumor-induced immunosuppression

BackgroundOvarian adenocarcinoma (OVAD) frequently metastasizes to the peritoneal cavity and manifests by the formation of ascites, which constitutes a tumor-promoting microenvironment. In the peritoneal cavity, two developmentally, phenotypically and functionally...

Aristolochic acid I exposure triggers ovarian dysfunction by activating NLRP3 inflammasome and affecting mitochondrial homeostasis.

Aristolochic acids are widely distributed in the plants of Aristolochiaceae family and Asarum species. Aristolochic acid I (AAI) is the most frequent compound of aristolochic acids, which can accumulate in the soil, and then contaminates crops and water and enters the human body. Research has shown that AAI affects the reproductive system. However, the mechanism of AAI's effects on the ovaries at the tissue level still needs to be clarified. In this research, we found AAI exposure reduced the body and ovarian growth in mice, decreased the ovarian coefficient, prevented follicular development, and increased atretic follicles. Further experiments showed that AAI upregulated nuclear factor-κB and tumor necrosis factor-α expression, activated the NOD-like receptor protein 3 inflammasome, and led to ovarian inflammation and fibrosis. AAI also affected mitochondrial complex function and the balance between mitochondrial fusion and division. Metabolomic results also showed ovarian inflammation and mitochondrial dysfunction due to AAI exposure. These disruptions reduced the oocyte developmental potential by forming abnormal microtubule organizing centers and expressing abnormal BubR1 to destroy spindle assembly. In summary, AAI exposure triggers ovarian inflammation and fibrosis, affecting the oocyte developmental potential.

ATAD2 is a driver and a therapeutic target in ovarian cancer that functions by upregulating CENPE

Ovarian cancer is a complex disease associated with multiple genetic and epigenetic alterations. The emergence of treatment resistance in most patients causes ovarian cancer to become incurable, and novel therapies remain necessary. We identified epigenetic regulator ATPase family AAA domain-containing 2 (ATAD2) is overexpressed in ovarian cancer and is associated with increased incidences of metastasis and recurrence. Genetic knockdown of ATAD2 or its pharmacological inhibition via ATAD2 inhibitor BAY-850 suppressed ovarian cancer growth and metastasis in both in vitro and in vivo models. Transcriptome-wide mRNA expression profiling of ovarian cancer cells treated with BAY-850 revealed that ATAD2 inhibition predominantly alters the expression of centromere regulatory genes, particularly centromere protein E (CENPE). In ovarian cancer cells, changes in CENPE expression following ATAD2 inhibition resulted in cell-cycle arrest and apoptosis induction, which led to the suppression of ovarian cancer growth. Pharmacological CENPE inhibition phenotypically recapitulated the cellular changes induced by ATAD2 inhibition, and combined pharmacological inhibition of both ATAD2 and CENPE inhibited ovarian cancer cell growth more potently than inhibition of either alone. Thus, our study identified ATAD2 as regulators of ovarian cancer growth and metastasis that can be targeted either alone or in combination with CENPE inhibitors for effective ovarian cancer therapy.

Fatty acid mobilization and histological changes at different maturation stages of female hilsa, Tenualosa ilisha

Hilsa, Tenualosa ilisha is a commercially important and high-valued anadromous fish of South East Asian countries. The present study aimed to understand macro and microscopic changes in the ovary and mobilization of fatty acids during gonad development in wild-caught female hilsa. A total of 174 female fish samples of different size groups (Weight range: 68.20 ± 3.21 to 460.67 ± 29.89 g, Total length range: 18.53 ± 0.55 to 40.33 ± 1.20 cm) were collected from estuarine and freshwater zones of the Hooghly River system of India. Various biological indices, such as gonadosomatic index (GSI), hepatosomatic index (HSI), oocyte diameter (OD), viscerosomatic index (VSI) and condition factor (K) at different maturation stages (I to VI) were determined. Results of the study showed that the GSI and OD increased with ovarian growth from stage I to VI (14.50 ± 2.18%) of maturation (P < 0.05). The HSI value increased progressively from stage I to V; thereafter, it decreased (P < 0.05). Histological observations indicated different microscopic stages, namely oogonia (stage I), chromatin nucleolar oocytes, perinuclolar oocytes (stage II), cortical alveolar (III), vitellogenic (IV), post-vitellogenic (V) and hydrated/eccentric stages (VI). Total saturated fatty acid (∑SFA) decreased in the ovary and increased in the liver with the progress of maturation from stage I to VI. At the same time, total monounsaturated fatty acid (∑MUFA) in muscle remained unchanged (P > 0.05), decreased (P < 0.05) in the liver, and increased in the ovarian tissue (P < 0.05). The levels of n-3 PUFA (polyunsaturated fatty acid) and n-6 PUFA elevated in the ovarian tissue concomitantly with the ovarian growth (P < 0.05). The ratios of n-3/n-6, docosahexaenoic acid (DHA) / eicosapentaenoic acid (EPA), EPA/ arachidonic acid (AA) and DHA/AA showed a significant decreasing trend in the ovarian tissue with the progress of gonad development (P < 0.05). Accumulation of n-3 PUFAs (α-linolenic acid, ALA; EPA; DHA) and n-6 PUFAs (linoleic acid, LA and AA) in ovary suggested specific fatty acid requirements during gonad development, which will help in the formulation of a broodstock diet for hilsa.

Human-relevant exposure to di-n-butyl phthalate tampers with the ovarian insulin-like growth factor 1 system and disrupts folliculogenesis in young adult mice.

Phthalates are compounds used in consumer and medical products worldwide. Phthalate exposure in women has been demonstrated by detection of phthalate metabolites in their urine and ovarian follicular fluid. High urinary phthalate burden has been associated with reduced ovarian reserve and oocyte retrieval in women undergoing assisted reproduction. Unfortunately, no mechanistic explanation for these associations is available. In short term in vivo and in vitro animal studies modeling human-relevant exposures to di-n-butyl phthalate (DBP), we have identified ovarian folliculogenesis as a target for phthalate exposures. In the present study, we investigated whether DBP exposure negatively influences insulin-like growth factor 1 (IGF1) signaling in the ovary and disrupts ovarian folliculogenesis. CD-1 female mice were exposed to corn oil (vehicle) or DBP (10 µg/kg/day, 100 µg/kg/day, or 1000 mg/kg/day) for 20-32 days. Ovaries were collected as animals reached the proestrus stage to achieve estrous cycle synchronization. Levels of mRNAs encoding IGF1 and 2 (Igf1 and Igf2), IGF1 receptor (Igf1r), and IGF-binding proteins 1-6 (Ifgbp1-6) were measured in whole ovary homogenates. Ovarian follicle counts and immunostaining for phosphorylated IGF1R protein (pIGF1R) were used to evaluate folliculogenesis and IGF1R activation, respectively. DBP exposure, at a realistic dose that some women may experience (100 µg/kg/day for 20-32 days), reduced ovarian Igf1 and Igf1r mRNA expression and reduced small ovarian follicle numbers and primary follicle pIGF1R positivity in DBP-treated mice. These findings reveal that DBP tampers with the ovarian IGF1 system and provide molecular insight into how phthalates could influence the ovarian reserve in females.

Ginsenoside 20(S)-Rg3 reduces KIF20A expression and promotes CDC25A proteasomal degradation in epithelial ovarian cancer

BackgroundGinsenoside 20(S)-Rg3 shows promising tumor-suppressive effects in ovarian cancer via inhibiting NF-κB signaling. This study aimed to explore the downstream tumor suppressive mechanisms of ginsenoside Rg3 via this signaling pathway. Materials and methodsA systematical screening was applied to examine the expression profile of 41 kinesin family member genes in ovarian cancer. The regulatory effect of ginsenoside Rg3 on KIF20A expression was studied. In addition, we explored interacting proteins of KIF20A and their molecular regulations in ovarian cancer. RNA-seq data from The Cancer Genome Atlas (TCGA) was used for bioinformatic analysis. Epithelial ovarian cancer cell lines SKOV3 and A2780 were used as in vitro and in vivo cell models. Commercial human ovarian cancer tissue arrays were used for immunohistochemistry staining. ResultsKIF20A is a biomarker of poor prognosis among the kinesin genes. It promotes ovarian cancer cell growth in vitro and in vivo. Ginsenoside Rg3 can suppress the transcription of KIF20A. GST pull-down and co-immunoprecipitation (IP) assays confirmed that KIF20A physically interacts with BTRC (β-TrCP1), a substrate recognition subunit for SCFβ−TrCP E3 ubiquitin ligase. In vitro ubiquitination and cycloheximide (CHX) chase assays showed that via interacting with BTRC, KIF20A reduces BTRC-mediated CDC25A poly-ubiquitination and enhances its stability. Ginsenoside Rg3 treatment partly abrogates KIF20A overexpression-induced CDC25A upregulation. ConclusionThis study revealed a novel anti-tumor mechanism of ginsenoside Rg3. It can inhibit KIF20A transcription and promote CDC25A proteasomal degradation in epithelial ovarian cancer.

Cancer-associated mesothelial cells are regulated by the anti-Müllerian hormone axis

Cancer-associated mesothelial cells (CAMCs) in the tumor microenvironment are thought to promote growth and immune evasion. We find that, in mouse and human ovarian tumors, cancer cells express anti-Müllerian hormone (AMH) while CAMCs express its receptor AMHR2, suggesting a paracrine axis. Factors secreted by cancer cells induce AMHR2 expression during their reprogramming into CAMCs in mouse and human invitro models. Overexpression of AMHR2 in the Met5a mesothelial cell line is sufficient to induce expression of immunosuppressive cytokines and growth factors that stimulate ovarian cancer cell growth in an AMH-dependent way. Finally, syngeneic cancer cells implanted in transgenic mice with Amhr2-/- CAMCs grow significantly slower than in wild-type hosts. The cytokine profile of Amhr2-/- tumor-bearing mice is altered and their tumors express less immune checkpoint markers programmed-cell-death 1 (PD1) and cytotoxic T lymphocyte-associated protein 4 (CTLA4). Taken together, these data suggest that the AMH/AMHR2 axis plays a critical role in regulating the pro-tumoral function of CAMCs in ovarian cancer.

Overexpression of &lt;i&gt;MKRN2&lt;/i&gt; Inhibits the Growth of Ovarian Cancer Cells

Ovarian cancer has a high mortality with low five-year survival rates. The role of the E3 ligase Makorin ring finger protein 2 (MKRN2) in ovarian cancer is unknown. This study investigated the impact of MKRN2 on the growth of ovarian cancer. MKRN2 expression in ovarian cancer tissue was analyzed by immunohistochemistry. Overexpression of MKRN2 was induced in two ovarian cancer cell lines (SKOV3 and CAOV3) by lentivirus transfection, and expression levels were verified by western blotting. Proliferation and growth were determined by CCK-8 and colony formation assays, while migration was examined using transwell assays and apoptosis by flow cytometry. Xenograft tumors of transfected SKOV3 cells were established in mice, and immunohistochemistry and TUNEL assays measured MKRN2 levels and apoptosis in tumor cells. Reduced levels of MKRN2 in cancerous tissue relative to non-cancerous ovarian tissues. Lentiviral-based MKRN2 overexpression in SKOV3 and CAOV3 cells reduced tumor-associated behavior while inducing apoptosis in vitro. In xenograft tumors, MKRN2 overexpression inhibited ovarian cancer growth and increased apoptosis in vivo. These findings imply the MKRN2 involvement in ovarian carcinogenesis and suggest its potential for treating the disease.

Dietary yolk supplements and rate of yolk deposition do not influence sex ratios in Japanese Quail.

Birds appear to have the ability to bias offspring sex ratios in response to environmental and social conditions. The mechanisms responsible remain unknown, however one previous study indicated a link between rates of ovarian follicle growth and the sexes of resulting eggs. This could indicate that either follicles destined to retain male or female follicles grow at different rates, or the rate of ovarian follicle growth determines the sex chromosome that will be retained to influence the sex of the resulting offspring. We tested for evidence of both possibilities by staining yolk rings that are indicative of daily growth. First, we tested for a correlation between the number of yolk rings and the sexes of the resulting germinal discs collected from each egg, and second, we tested whether experimentally decreasing follicle growth rates with a dietary yolk supplement impacts the sexes of the resulting germinal discs. There was no significant correlation between the number of yolk rings and the sexes of the resulting embryos, and decreasing follicle growth rates did not impact sexes of resulting germinal discs. These results indicate that offspring sex is not related to the rate of ovarian follicle growth in quail.

TOM40 regulates the progression of nasopharyngeal carcinoma through ROS-mediated AKT/mTOR and p53 signaling

Nasopharyngeal carcinoma (NPC) is a prevalent cancer in Southern China, North Africa, and Southeast Asia. The translocase of the outer membrane (TOM) 40 is a transporter of mitochondrial proteins, and is involved in ovarian cancer cell growth. However, its role in the progression of NPC is still unclear. We found that TOM40 levels were upregulated in NPC tissues and multiple NPC cell lines. In addition, high TOM40 expression in the tumor tissues was associated with poor overall survival and disease specific survival. TOM40 knockdown in the NPC cell lines inhibited their proliferation in vitro and in vivo. Furthermore, TOM40 silencing also increased intracellular production of reactive oxygen species (ROS) and decreased mitochondrial membrane potential (MMP). Mechanistically, the anti-tumor effects of TOM40 silencing were dependent on the inhibition of AKT/mTOR signaling and activation of p53 signaling. To summarize, TOM40 mediates NPC progression through ROS-mediated AKT/mTOR and p53 signaling. Our findings highlight the potential of TOM40 as a therapeutic target for NPC.

P-647 Underlying mechanisms in clinical manifestations of polycystic ovarian morphology (PCOM) and PCOS women

Abstract Study question Does the D19S884 allele 8 (A8) equally affect the pathogenesis and ovarian gene expression of PCOS and PCOM patients? Summary answer A8-allele produces metabolic, endocrine, and ovarian alterations regardless diagnose. The mechanisms involved in PCOM alterations are different from those of the ovarian phenotype of PCOS. What is known already The specific D19S884 allele 8 of the FBN3 gene may be related to polycystic ovarian syndrome (PCOS) clinical manifestations. The A8 allele participates in alternative splicing of FBN3 and produces Asprosin-3, related to glucose modulation, and Fibrillin-3. Fibrilin-3 is an extracellular matrix protein, and together with a dysregulation of the Hippo pathway, could be responsible for constraining follicular growth in the PCOS ovaries. However, it is still unknown how these pathways act in PCOS, and whether these mechanisms are also involved in pathophysiology of polycystic ovarian morphology (PCOM) in apparently normal women with regular menses. Study design, size, duration Cross-sectional and descriptive study with 139 women (24-39 years-old) undergoing an IVF cycle between 2019-2022 at Hospital La Fe (Valencia, Spain). Thirty-one patients were considered PCOS, twenty-eight were classified as PCOM while the remaining eighty were controls. After recruitment women were screened for A8 allele, metabolic status, hormone profile, follicular fluid protein determination and expression of Hippo pathway and extracellular matrix genes. The IVF cycle parameters were recorded to determine their relationship with study variables. Participants/materials, setting, methods Women with two or more Rotterdam-criteria were considered as PCOS. PCOM patients were defined by polycystic ovarian morphology on ultrasound with regular ovulatory cycles. Genomic DNA was isolated from blood samples to assess the presence of A8 allele by capillary electrophoresis and FBN3 concentration was measured on follicular fluid by ELISA. TaqMan qPCR assay was used to analyze the expression of Hippo pathway (BIRC1 and CCN2) and extracellular matrix (ECM) genes in cumulus cells. Main results and the role of chance PCOS patients showed statistically significant metabolic and endocrine alterations with higher BMI, free androgen index (FAI), and glucose levels compared to controls. Despite having increased AMH levels (PCOS:45.0±21.7; PCOM:33.8±21.5; Control:17.6±7.19pmol/L, p &amp;lt; 0.05), and AMH/AFC ratio (PCOS:1.9±0.9; Control:1.2±0.5, p = 0.008), PCOS showed lower fertilization rates (PCOS:64±26; Control:80±18%, p = 0.008), reduced good quality (PCOS:0.6±1.0; Control:1.3±1, p = 0.003) and transferred embryos (PCOS:1.3±0.9; Control:1.9±1.0, p = 0.018). In PCOM, metabolic and androgen profiles were not different from controls. Although AMH, AFC (PCOM:24.4±13.7; Control:15.5±7.1, p = 0.007) and aspirated follicles (PCOM:17.3±5.7; Control:14.3±7.0, p = 0.026) were increased, fewer embryos were transferred (PCOM:1.2±1.1; Control:1.9±1.0, p = 0.027). PCOM women also showed ovarian downregulation of the Hippo-pathway (CCN2-PCOM:-4.8±10.8; Control:0.1±3.0, p = 0.010) and increased FBN3 concentration (PCOM:19.6±8.8; Control:14.7±5.6ng/ml, p=N.S). The A8 allele was detected in 17% of our patients: 4% PCOS, 50% PCOM and 46% controls. Overall, A8 presence associated a Hippo-pathway downregulation (BIRC1-A8+:-2.7±4.5; A8-:0.3±2.2, p = 0.034) and increased ECM expression (EMILIN1-A8+:2.1±1.0; A8-:1.2±1.5, p = 0.04). Interestingly, in controls, higher glucose (A8+:95.1±6.7; A8-:83.5±10.1mg/dl, p = 0.002), cholesterol (A8+:182.5±11.1; A8-:165.4±27.3mg/dl, p = 0.029), LDL (A8+:117.8±10.4; A8-:83.9±29.4mg/dl, p = 0.002), and DHEAS levels (A8+:2633.0±670.7; A8-:1585.8±670.0ng/ml, p = 0.026) were found, consistent with a reduction in HDL (A8+:51.8±8.8; A8-:67.4±15.43mg/dl, p = 0.026) and a downregulation of the Hippo-pathway (BIRC1-A8+:-4.8±4.6; A8-:0.2±4.6, p = 0.013). In PCOM, A8+ promoted higher FAI (A8+:1.6±1.5; A8-:0.8±0.7, p=N.S), and less embryos obtained (A8+:5.9±3.8; A8-:10.7±3.8, p = 0.04). Limitations, reasons for caution Due to the low prevalence of A8 in our PCOS population, its effects cannot be evaluated in this group. Further validation of gene expression results by RNA-seq will be required to assess the broad spectrum of ovarian transcriptomic changes induced by both, the PCOM phenotype and the A8 allele. Wider implications of the findings Polycystic ovarian morphology is not unique to clinical disorder of PCOS and the triggering mechanisms appear to be distinct from those observed in women with regular cycles. Our findings suggest that the presence of A8 allele impacts on metabolic profile, androgen levels, and ovarian pathway dysregulations, despite the clinical diagnose. Trial registration number Not applicable

P-410 First wave of primordial follicle activation during human ovarian tissue manipulation for fertility preservation

Abstract Study question Does ovarian tissue biopsy, transportation and processing for fertility preservation and restoration trigger primordial follicle activation? Summary answer Early manipulation of ovarian tissue is sufficient to trigger follicle activation by stimulating PI3K/Akt and disrupting the Hippo pathway. What is known already Primordial follicle recruitment occurs continuously in physiological conditions by modulation of autocrine and paracrine factors, like the PI3K/Akt and Hippo pathways, to ensure follicle growth over time. During fertility preservation and restoration procedures, follicle activation may be disrupted and follow nonphysiological patterns. The ability to control activation dynamics by up- or downregulation of these pathways may enhance fertility restoration outcomes in a number of ways. Indeed, downregulation of follicle activation shortly after transplantation may protect the ovarian reserve from early depletion. Conversely, ovarian tissue in vitro culture may benefit from upregulation of primordial follicle activation to boost further growth. Study design, size, duration Fresh ovarian tissue was retrieved from nine women undergoing laparoscopic surgery for benign conditions. Three timepoints were investigated. One-third of collected tissue per patient was immediately fixed in the operating room, without any manipulation (time zero, T0). The remaining tissue was transferred to the laboratory and dissected to remove any surplus medulla. It was then cut into small cortical fragments, half of which were fixed after 25 minutes(T25) and the other half after 90 minutes(T90). Participants/materials, setting, methods All cortical fragments were fixed in 4%formaldehyde and embedded in paraffin for histology. In order to explore follicle activation,markers of the PI3K/Akt and Hippo signaling pathways were immunolabeled at each timepoint, targeting: (i) phospho-Akt (p-Akt) in primordial follicles by immunohistochemistry as a marker of early PI3K/Akt pathway activation; and (ii) Yes-associated protein (YAP) cellular localizationin the granulosa cell layer of primordial follicles by immunofluorescence as a marker of Hippo disruption. Main results and the role of chance An upturn in p-Akt expression was observed at T25 (22,34 ± 0.13%; p = 0.0233) and T90 (39,01 ± 0.22%, p = &amp;lt; 0.0001) compared to T0 (2,87 ± 0,03%). In terms of YAP cellular localization, a significant nucleus-to-cytoplasm shift was detected at T25 (1.11 ± 0.09; p = 0.0428) compared to T0 (0.97 ± 0.10), while T90 (1.07± 0.14) values were similar to T25. Our data prove that ovarian tissue manipulation triggers primordial follicle activation very early, involving both the PI3K/Akt and Hippo signaling pathways, which appear to cooperate in primordial-to-primary follicle transition. Our results indicate that the first stages of any fertility preservation or restoration procedure involving ovarian tissue manipulation contribute to dysregulation of the very mechanisms responsible for the ovarian reserve maintenance and follicle growth. Additional strategies are required to gain the control of follicle activation mechanisms in nonphysiological conditions (ex vivo ovarian tissue manipulation), in order to exploit ovarian reserve dynamics to serve the need of patients. Limitations, reasons for caution Analyses in the study were limited to histology and immunolabeling to acquire a descriptive picture of pathway activation kinetics over time. Further investigations using dynamic experimental models are essential to advance our understanding of signaling pathway synergy in vivo. Wider implications of the findings Since dysregulation of follicle activation in nonphysiological conditions appears to be associated with poor oocyte quality, enhancing our ability to control the relevant signaling pathways is crucial to optimizing fertility preservation procedures. Trial registration number not applicable

O-090 Endocrine disruptors

Abstract Female mammals, including women, are exposed to endocrine disrupting chemicals (EDCs) on a daily basis. This is of concern because many EDCs inhibit ovarian follicle growth and steroidogenesis, processes that are critical for maintaining female fertility. Our research focuses on the effects of EDCs known as phthalates on female fertility. Phthalates are of interest because they are ubiquitous synthetic chemicals used as plasticizers and stabilizers in a myriad of consumer products, including everything from shower curtains to children’s toys to cosmetics and personal care products such as perfumes, nail polish, deodorants, and lotions. Phthalates are also used in pesticides, wood finishes, adhesives, solvents, lubricants, defoaming agents, and in medical devices including tubing, blood bags, surgical gloves, and dialysis equipment. Despite the widespread use of phthalates and ubiquitous human exposure, limited information is available about the effects of environmentally relevant phthalate mixtures on female reproduction. Thus, we tested the hypothesis that prenatal exposure to an environmentally relevant phthalate mixture affects female reproduction in the F1, F2, and F3 generations of mice. To test this hypothesis, pregnant CD-1 mice were orally dosed with vehicle control (corn oil) or a phthalate mixture (20µg/kg/day-500mg/kg/day) daily from gestational day 10 to birth. The mixture was based on urinary phthalate metabolite levels measured pregnant women in the United States, and consisted of 35% diethyl phthalate, 21% di(2-ethylhexyl) phthalate, 15% dibutyl phthalate, 15% di-isononyl phthalate, 8% diisobutyl phthalate, and 5% benzylbutyl phthalate. Adult F1 females born to these dams were used to create the F2 generation by mating them with unexposed males and F2 females were used to create the F3 generation by mating them with unexposed males. At selected times, estrous cyclicity and fertility indices were monitored and ovaries and sera were collected for analysis. Our data indicate that prenatal exposure to the phthalate mixture significantly decreases anogenital distance, testosterone levels, and litter size, but it significantly increases abnormal estrous cyclicity, uterine weight, the number of cystic ovaries, follicle-stimulating hormone levels, and luteinizing hormone levels compared to control in the F1 generation. Further, our data indicate that prenatal exposure to the phthalate mixture increases body weight, uterine weight, the number of cystic ovaries, and time to pregnancy and it decreases testosterone levels compared to control in the F2 generation. In addition, our data indicate that prenatal exposure to the phthalate mixture increases body weight, uterine weight, the number of cystic ovaries, and abnormal cyclicity and it decreases anogenital distance and the number of live pups compared to control in the F3 generation. Collectively, these data suggest that prenatal exposure to an environmentally relevant mixture of phthalates adversely affects reproductive function in a multi- and transgenerational manner in female mice. These findings have increased our understanding of the mechanisms by which EDCs cause female reproductive toxicity. This information eventually may lead to the development of novel targets for the prevention or treatment of infertility induced by EDCs.