Human Ovary Research Articles

Retinoic acid (all-trans) presents antioxidant properties within human ovary and reduces progesterone production by human granulosa cells

Both vitamin A and E support female reproduction and embryonic development. These vitamins have been associated with decreased fertility or failure to end the pregnancy in animals. An observational study was conducted on follicular fluid (FF) samples to determine the concentrations of fat-soluble vitamins of women undergoing in vitro fertilization and its correlation with assisted reproductive technology characteristics and pregnancy outcomes. Moreover, the effects of all-trans-retinoic acid (atRA) and alpha-tocopherol on granulosa cell viability, apoptosis, autophagy and hormonal production were evaluated. No association was identified between fat-soluble vitamin concentrations in FF and infertility aetiology, body mass index or woman’s age. There were differences in follicular antioxidant profiles and ovarian response stimulation. In vitro evaluation of atRA and alpha-tocopherol reveals that, at physiological concentrations, both compounds may affect the viability of granulosa cells. In addition, these compounds are able to protect granulosa cells from oxidative stress, as well as to affect estradiol and progesterone production. Our data suggest that atRA and alpha-tocopherol levels should be well controlled as they may have implications in the function and viability of granulosa cells and highlights retinol as a marker of the oxidative defenses within ovary environment.

A predictive model of the effect of therapeutic radiation on the human ovary.

Radiation to the female pelvis as part of treatment for cancer predisposes young women to develop Premature Ovarian Insufficiency (POI). As the human female is born with their full complement of non-growing follicles which decline in an exponential fashion until the menopause, the age at which POI occurs is dependent on the age of the patient at treatment and the dose received by the ovary. A model that predicts the age at which POI occurs for a known dose at a known age will aid counselling patients on their fertility risk. Patients deemed to be at high risk of POI may be considered to be good candidates for established fertility preservation techniques. An updated and externally validated model of the age-related decline in human ovarian reserve was combined with the best available estimate of the median lethal dose LD50 for the human ovary. Using known age at diagnosis and posited radiotherapy treatment plan to estimate the dose to the least-affected ovary, we use an age-related model of the decline in ovarian reserve to generate a personalized age prediction of premature ovarian insufficiency. Our algorithm is available as an online calculator which graphs model outputs to inform discussions around survivor fertility. We report four example cases across different ages and diagnoses, each with two carefully designed photon and proton treatment plans. The treatment options are compared in terms of remaining fertile lifespan for the survivor. International oncology guidelines now mandate the consideration of later fertility when reviewing treatment options for children diagnosed with cancer. Our calculator (https://sites.cs.st-andrews.ac.uk/radiosensitivity), and the underlying algorithm and models, allow detailed predictions of the impact of various radiotherapy plans on fertility. These patient-specific data enhance pre-treatment discussions around post-treatment fertility and fertility preservation.

Correction for: Nicotine exposure impairs germ cell development in human fetal ovaries cultured in vitro.

Correction for: Nicotine exposure impairs germ cell development in human fetal ovaries cultured in vitro.

Effect of intra ovarian injection of platelet rich plasma to patients with diminished ovarian reserve (DOR)- A Prospective Study

Background: Diminished ovarian reserve (DOR) is among the common findings in infertile women with no significant underlying condition especially women with advanced age. The aim of this study was to investigate the intra-ovarian potential of platelet-rich plasma (PRP) administration on oocytesdependent variables in the DOR women. A few years ago, we used intrauterine PRP injection in patients with recurrent implantation failure (RIF) which led to noticeable findings. Nowadays, PRP is used more widely in reproductive medicine due to its regenerative potentials; however, not enough data is available on this issue. One of the new strategies for facing primary ovarian insufficiency is PRP therapy. Material and Methods: This is a Prospective cohort study conducted in three IVF Clinics Thane-India, Almaty-Kazakhstan and Tashkent- Uzbekistan from October 2021 to September 2022. All couples with failed IVF with diminished ovarian reserve as per ARSM definition between the age group of 35 to 45 (both inclusive). To standardize the definition of poor ovarian response, a European Society of Human Reproduction and Embryology Working Group convened in Bologna and proposed that two of the following criteria be present to define whether a given low response to stimulation is truly representative of poor ovarian response: 1) Advanced maternal age (R40 years). 2) A previous poor ovarian response. Result: Demographic characteristics, baseline ovarian reserve markers and ultrasound findings of women included in the analysis Table 1. Women treated with PRP had significant improvement in biochemical and ultrasound markers of ovarian reserve. Notably, AMH levels were on average 72% higher following PRP (P < 0.001). Of the 124 women who underwent IVF/ICSI following participation in the study, those with previous PRP treatment yielded an average more than 1.5× the number of oocytes collected. Conclusion: This study revealed that the injection of PRP into human ovaries is safe and improves ovarian reserve markers as measured by antral follicle count and serum levels of AMH and FSH. Nevertheless, further studies are needed to evaluate the impact of PRP on pregnancy outcomes in women undergoing ART.

Multifaceted Functions of TWSG1: From Embryogenesis to Cancer Development.

Bone morphogenetic proteins (BMPs) play an important role in development. Twisted gastrulation BMP signaling modulator 1 (TWSG1) was initially identified as a regulator of the dorsoventral axis formation in Drosophila. The mechanism of BMP signaling modulation by TWSG1 is complex. TWSG1 inhibits BMP signaling by binding to BMP ligands including BMP4, whereas it enhances signaling by interacting with Chordin, a BMP antagonist. Therefore, TWSG1 can act as both a BMP agonist and antagonist. TWSG1 has various functions ranging from embryogenesis to cancer progression. TWSG1 knockout mice showed neural, craniofacial, and mammary defects. TWSG1 also regulated erythropoiesis and thymocyte development. Furthermore, the relationship between TWSG1 and cancer has been elucidated. Allelic loss of TWSG1 was detected in colorectal cancer. TWSG1 expression was upregulated in papillary thyroid carcinoma and glioblastoma but downregulated in gastric and endometrial cancers. TWSG1 suppressed BMP7-enhanced sphere formation and migration in endometrial cancer cells, indicating its tumor-suppressive role. Further studies are required to clarify the TWSG1 function and its association with BMP signaling in cancer development. Finally, TWSG1 is abundantly expressed in human and mouse ovaries and sustains follicular growth in rodent ovaries. Thus, TWSG1 has various functions ranging from fertility to cancer. Therefore, TWSG1 signaling modulation may be beneficial in treating specific diseases such as cancer.

Single-cell RNA-sequencing of retrieved human oocytes and eggs in clinical practice and for human ovarian cell atlasing.

With the advancement of single-cell separation techniques and high-throughput sequencing platforms, single-cell RNA-sequencing (scRNA-seq) has emerged as a vital technology for understanding tissue and organ systems at cellular resolution. Through transcriptional analysis, it is possible to characterize unique or rare cell types, interpret their interactions, and reveal novel functional states or shifts in developmental stages. As such, this technology is uniquely suited for studying the cells within the human ovary. The ovary is a cellularly heterogeneous organ that houses follicles, the reproductive and endocrine unit that consists of an oocyte surrounded by hormone-producing support cells, as well as many other cell populations constituting stroma, vasculature, lymphatic, and immune components. Here we review studies that have utilized scRNA-seq technology to analyze cells from healthy human ovaries and discuss the single-cell isolation techniques used. We identified two overarching applications for scRNA-seq in the human ovary. The first applies this technology to investigate transcriptional differences in oocytes/eggs from patients undergoing in vitro fertilizationtreatments to ultimately improve clinical outcomes. The second utilizes scRNA-seq for the pursuit of creating a comprehensive single-cell atlas of the human ovary. The knowledge gained from these studies underscores the importance of scRNA-seq technologies in unlocking a new biological understanding of the human ovary.

Development of the human ovary: Fetal through pubertal ovarian morphology, folliculogenesis and expression of cellular differentiation markers

A definition of normal human fetal and early postnatal ovarian development is critical to the ability to accurately diagnose the presence or absence of functional ovarian tissue in clinical specimens. Through assembling an extensive histologic and immunohistochemical developmental ontogeny of human ovarian specimens from 8 weeks of gestation through 16 years of postnatal, we present a comprehensive immunohistochemical mapping of normal protein expression patterns in the early fetal through post-pubertal human ovary and detail a specific expression-based definition of the early stages of follicular development. Normal fetal and postnatal ovarian tissue is defined by the presence of follicular structures and characteristic immunohistochemical staining patterns, including granulosa cells expressing Forkhead Box Protein L2 (FOXL2). However, the current standard array of immunohistochemical markers poorly defines ovarian stromal tissue, and additional work is needed to identify new markers to advance our ability to accurately identify ovarian stromal components in gonadal specimens from patients with disorders of sexual differentiation.

Automatic Evaluation for Bioengineering of Human Artificial Ovary: A Model for Fertility Preservation for Prepubertal Female Patients with a Malignant Tumor.

Introduction: The in vitro culture of primordial follicles is the only available option for preserving fertility in prepubertal girls with malignant tumors. The cultivation of primordial follicles in scaffolds as artificial ovaries is a promising approach for this. Methods: Dissociated follicles were placed into an artificial ovarian scaffold composed of fibrinogen and thrombin. The follicles were cultured in a dish dedicated to live cell imaging and observed for growth using immunofluorescence and development via optical microscopy. The morphology of the follicles in the scaffold was three-dimensionally reconstructed using the Imaris software. Growth and development were also quantified. Results: The morphology of artificial ovaries began to degrade over time. Within approximately 7 days, primordial follicles were activated and grew into secondary follicles. A comparison of optical and confocal microscopy results revealed the superior detection of live cells using confocal microscopy. The three-dimensional reconstruction of the confocal microscopy data enabled the automatic enumeration and evaluation of the overall morphology of many follicles. Conclusions: The novel artificial ovary-enabled primordial follicles to enter the growth cycle after activation and grow into secondary follicles. The use of a fibrin scaffold as a carrier preserves the developmental potential of primordial germ cells and is a potentially effective method for preserving fertility in prepubertal girls.

Linifanib induces apoptosis in human ovarian cancer cells via activation of FOXO3 and reactive oxygen species

Linifanib is known as an inhibitor of receptor tyrosine kinase. Even though it has been widely recognized as efficient inhibitor of receptor tyrosine kinases, anti-carcinogenic effect has not been investigated enough in ovarian cancer. In this study, we investigated the anti-cancer effect of linifanib on human ovary cancer SKOV3 cells. WST-1, cell counting assay, and observation of morphological changes were performed to evaluate the cytotoxic effect of linifanib in SKOV3 cells. We analyzed SKOV3 cells treated with linifanib using Muse cell analyzer. We focused on investigating the effect of linifanib on DNA damage in nucleus. Additionally, intracellular reactive oxygen species (ROS) level was measured through Muse cell analyzer. Western blotting was performed to evaluate the protein expression level related to apoptosis. We found that linifanib inhibited proliferation of SKOV3 cells. Our results showed that linifanib induced apoptosis in SKOV3 cells. Additionally, linifanib induced DNA damage in SKOV3 cells. We found that intracellular ROS level increased after treatment of linifanib in SKOV3 cells. Interestingly, FOXO3 was transferred from cytosol into nucleus after linifanib treatment. Taken together, our results supported that linifanib inhibited the proliferation of human ovary cancer SKOV3 cells, which suggested that linifanib might have the potential to be developed as drugs for ovarian cancer treatment.

Different ages of development human ovaries exhibit different telomerase activity and telomere length leads to ovarian aging

Background and Aim: Telomeres are repetitive DNA sequences localized at the ends of eukaryotic chromosomes. These unique structures are composed of telomeric DNA and telomere-associated proteins. Of telomere-associated proteins, telomeric repeat factor 1 (TRF1) and 2 (TRF2) bind double-stranded telomeric DNA to regulate telomeres length throughout cellular lifespan. On the other hand, protection of telomeres 1 (POT1) interacts with single-stranded telomeric DNA and contributes to protecting genome integrity.

Anatomic nomenclature and 3-dimensional regional model of the human ovary: call for a new paradigm

The ovaries are the female gonads that are crucial for reproduction, steroid production, and overall health. Historically, the ovary was broadly divided into regions defined as the cortex, medulla, and hilum. This current nomenclature lacks specificity and fails to consider the significant anatomic variations in the ovary. Recent technological advances in imaging modalities and high-resolution omic analyses have brought about the need for revision of the existing definitions, which will facilitate the integration of generated data and enable the characterization of organ subanatomy and function at the cellular level. The creation of these high-resolution multimodal maps of the ovary will enhance collaboration and communication among disciplines and between clinicians and researchers. Beginning in March 2021, the Pediatric and Adolescent Gynecology Program of the Eunice Kennedy Shriver National Institute of Child Health and Human Development invited subject-matter experts to participate in a series of workshops and meetings to standardize ovarian nomenclature and define the organ's features. The goal was to develop a spatially defined and semantically consistent terminology of the ovary to support collaborative, team science-based endeavors aimed at generating reference atlases of the human ovary. The group recommended a standardized, 3-dimensional description of the ovary and an ontological approach to the subanatomy of the ovary and definition of follicles. This new greater precision in nomenclature and mapping will better reflect the ovary's heterogeneous composition and function, support the standardization of tissue collection, facilitate functional analyses, and enable clinical and research collaborations. The conceptualization process and outcomes of the effort, which spanned the better part of 2021 and early 2022, are introduced in this article. The institute and the workshop participants encourage researchers and clinicians to adopt the new systems in their everyday work to advance the overarching goal of improving human reproductive health.

The Continued Absence of Functional Germline Stem Cells in Adult Ovaries.

Ovaries are central to development, fertility, and reproduction of women. A particularly interesting feature of ovaries is their accelerated aging compared to other tissues, leading to loss of function far before other organs senesce. The limited pool of ovarian follicles is generated before birth and once exhausted, menopause will inevitably commence around the age of 50 years marking the end of fertility. Yet, there are reports suggesting the presence of germline stem cells and neo-oogenesis in adult human ovaries. These observations have fueled a long debate, created experimental fertility treatments, and opened business opportunities. Our recent analysis of cell types in the ovarian cortex of women of fertile age could not find evidence of germline stem cells. Like before, our work has been met with critique suggesting methodological shortcomings. We agree that excellence starts with methods and welcome discussion on the pros and cons of different protocols. In this commentary, we discuss the recent re-interpretation of our work.

Protective Effects of PollenAid Plus Soft Gel Capsules' Hydroalcoholic Extract in Isolated Prostates and Ovaries Exposed to Lipopolysaccharide.

Pollen extract represents an innovative approach for the management of the clinical symptoms related to prostatitis and pelvic inflammatory disease (PID). In this context, the aims of the present work were to analyze the phenolic composition of a hydroalcoholic extract of PollenAid Plus soft gel capsules, and to evaluate the extract’s cytotoxic effects, in human prostate cancer PC3 cells and human ovary cancer OVCAR-3 cells. Additionally, protective effects were investigated in isolated prostate and ovary specimens exposed to lipopolysaccharide (LPS). The phytochemical investigation identified catechin, chlorogenic acid, gentisic acid, and 3-hydroxytyrosol as the prominent phenolics. The extract did not exert a relevant cytotoxic effect on PC3 and OVCAR-3 cells. However, the extract showed a dose-dependent inhibition of pro-inflammatory IL-6 and TNF-α gene expression in prostate and ovary specimens, and the extract was effective in preventing the LPS-induced upregulation of CAT and SOD gene expression, which are deeply involved in tissue antioxidant defense systems. Finally, a docking approach suggested the capability of catechin and chlorogenic acid to interact with the TRPV1 receptor, playing a master role in prostate inflammation. Overall, the present findings demonstrated anti-inflammatory and antioxidant effects of this formulation; thus, suggesting its capability in the management of the clinical symptoms related to prostatitis and PID.

Spatial Proteomics for Further Exploration of Missing Proteins: A Case Study of the Ovary.

In the quest for "missing proteins" (MPs), the proteins encoded by the human genome still lacking evidence of existence at the protein level, novel approaches are needed to detect this challenging group of proteins. The current count stands at 1,343 MPs, and it is likely that many of these proteins are expressed at low levels, in rare cell or tissue types, or the cells in which they are expressed may only represent a small minority of the tissue. Here, we used an integrated omics approach to identify and explore MPs in human ovaries. By taking advantage of publicly available transcriptomics and antibody-based proteomics data in the Human Protein Atlas (HPA), we selected 18 candidates for further immunohistochemical analysis using an exclusive collection of ovarian tissues from women and patients of reproductive age. The results were compared with data from single-cell mRNA sequencing, and seven proteins (CTXN1, MRO, RERGL, TTLL3, TRIM61, TRIM73, and ZNF793) could be validated at the single-cell type level with both methods. We present for the first time the cell type-specific spatial localization of 18 MPs in human ovarian follicles, thereby showcasing the utility of the HPA database as an important resource for identification of MPs suitable for exploration in specialized tissue samples. The results constitute a starting point for further quantitative and qualitative analysis of the human ovaries, and the novel data for the seven proteins that were validated with both methods should be considered as evidence of existence of these proteins in human ovary.

Metformin modulates the gut microbiome in broiler breeder hens.

Broiler breeder hens, the parent stock of commercial broiler chickens, are genetically selected for rapid growth. Due to a longer production period and the focus of genetic selection on superior carcass traits in their progeny, these hens have the propensity to gain excess adipose tissue and exhibit severe ovarian dysfunction, a phenotype that is similar to human polycystic ovary syndrome (PCOS). Metformin is an antihyperglycemic drug approved for type 2 diabetes that is prescribed off-label for PCOS with benefits on metabolic and reproductive health. An additional effect of metformin treatments in humans is modulation of gut microbiome composition, hypothesized to benefit glucose sensitivity and systemic inflammation. The effects of dietary metformin supplementation in broiler breeder hens have not been investigated, thus we hypothesized that dietary metformin supplementation would alter the gut microbiome of broiler breeder hens. Broiler breeder hens were supplemented with metformin at four different levels (0, 25, 50, and 75 mg/kg body weight) from 25 to 65 weeks of age, and a subset of hens (n = 8–10 per treatment group) was randomly selected to undergo longitudinal microbiome profiling with 16S rRNA sequencing. Metformin impacted the microbial community composition in 75 mg/kg metformin compared to controls (adjusted PERMANOVA p = 0.0006) and an additional dose-dependent difference was observed between 25 mg/kg and 75 mg/kg (adjusted PERMANOVA p = 0.001) and between 50 mg/kg and 75 mg/kg (adjusted PERMANOVA p = 0.001) but not between 25 mg/kg and 50 mg/kg (adjusted PERMANOVA p = 0.863). There were few differences in the microbiome attributed to hen age, and metformin supplementation did not alter alpha diversity. Bacteria that were identified as differentially relatively abundant between 75 mg/kg metformin treatment and the control, and between metformin doses, included Ruminococcus and members of the Clostridia family that have been previously identified in human trials of PCOS. These results demonstrate that metformin impacts the microbiome of broiler breeder hens in a dose-dependent manner and several findings were consistent with PCOS in humans and with metformin treatment in type 2 diabetes. Metformin supplementation is a potentially promising option to improve gut health and reproductive efficiency in broiler breeder hens.

Isolation of Female Germline Stem Cells from Mouse and Human Ovaries by Differential Adhesion.

Spermatogonial stem cell (SSC) counterparts known as female germline stem cells (fGSCs) were found in the mammalian ovary in 2004. Although the existence of fGSCs in the mammalian postnatal ovary is still under controversy, fGSC discovery encourages investigators to better understand the various aspects of these cells. However, their existence is not accepted by all scientists in the field because isolation of fGSCs by fluorescent activated cell sorting (FACS) has not been reproducible. In this study, we used differential adhesion to isolate and enrich fGSCs from mouse and human ovaries and subsequently cultured them in vitro. fGSCs were able to proliferate in vitro and expressed germ cell-specific markers Vasa, Dazl, Blimp1, Fragilis, Stella, and Oct4, at the protein level. Moreover, mouse and human fGSCs were, respectively, cultured for more than four months and one month in culture. Both mouse and human fGSCs maintained the expression of germ cell-specific markers over these times. In vitro cultured fGSCs spontaneously produced oocyte-like cells (OLCs) which expressed oocyte-relevant markers. Our results demonstrated that differential adhesion allows reproducible isolation of fGSCs that are able to proliferate in vitro over time. This source of fGSCs can serve as a suitable material for studying mechanisms underlying female germ cell development and function.

S-15-04 Knowledge gaps: case studies from human fetal ovary and adrenal gland

S-15-04 Knowledge gaps: case studies from human fetal ovary and adrenal gland

Assessment Of The Follicles in Different Age Groups in Post Mortem Human Ovary

Ovary is one of the essential reproductive organs in human being. A detailed histomorphological feature of ovary is an un-debatable for diagnostic and therapeutic purpose for ovarian disease. The standardization of the morphological and histological measurements of ovary in general in Bangladeshi people is essential to increase the knowledge regarding anatomical variation in our population. So a research work on detailed anatomical parameters of ovary is an un-debatable.
 Objective: To find out the follicles in different age groups post mortem human ovary.
 Methodology: The cross-sectional observational study was carried out in the Department of Anatomy, Sylhet MAG Osmani Medical College, Sylhet in collaboration with the Department of Forensic Medicine, Sylhet MAG Osmani Medical College during the period from January 2016 to December 2016. Sixty pairs of human postmortem ovaries were collected. The inclusion criteria were dead bodies autopsied within 36 hours of death. Considerable signs of decomposition or decomposed dead body, presence of gross ovarian disease and any history of poisoning cases were excluded. All the specimens were examined for morphological parameters but only 18 specimens were examined histological parameters.
 Results: The highest total numbers of the follicles were found in the age group A and lowest total numbers of the follicles were in the age group C. The differences of total numbers of the follicles were significant between group-A and group-B (p<0.001); and group-A and group-C (p<0.001); but no significant difference was observed between group-B and group-C (p=0.080).
 Conclusion: Histomorphologically, the total number of the follicles is found maximum at reproductive years and is declined through perimenopause and postmenopause.
 Central Medical College Journal Vol 5 No 2 Jul 2021 PP 84-87

Characterization of the roles of amphiregulin and transforming growth factor β1 in microvasculature-like formation in human granulosa-lutein cells.

Vascular endothelial-cadherin (VE-cadherin) is an essential component that regulates angiogenesis during corpus luteum formation. Amphiregulin (AREG) and transforming growth factor β1 (TGF-β1) are two intrafollicular factors that possess opposite functions in directing corpus luteum development and progesterone synthesis in human granulosa-lutein (hGL) cells. However, whether AREG or TGF-β1 regulates the VE-cadherin expression and subsequent angiogenesis in the human corpus luteum remains to be elucidated. Results showed that hGL cells cultured on Matrigel spontaneously formed capillary-like and sprout-like microvascular networks. Results of specific inhibitor treatment and small interfering RNA-mediated knockdown revealed that AREG promoteed microvascular-like formation in hGL cells by upregulating the VE-cadherin expression mediated by the epidermal growth factor receptor (EGFR)-extracellular signal-regulated kinase1/2 (ERK1/2) signaling pathway. However, TGF-β1 suppressed microvascular-like formation in hGL cells by downregulating VE-cadherin expression mediated by the activin receptor-like kinase (ALK)5-Sma- and Mad-related protein (SMAD)2/3/4 signaling pathway. Collectively, this study provides important insights into the underlying molecular mechanisms by which TGF-β1 and AREG differentially regulate corpus luteum formation in human ovaries.

Extracts of Common Vegetables Inhibit the Growth of Ovary Cancer Cells.

There is recent interest in a diet that can be recommended for patients suffering from cancer. In this respect, the effects were studied of the extracts of several common fruits, herbs and vegetables on the viability of two human ovary cancer cell lines (SKOV-3 and PEO1) in vitro. Normal human MRC-5 fibroblasts were used as a control cell line. The extracts of garlic, horseradish and curly kale as well as green and black tea were the most effective in lowering the viability of ovarian cancer cells, while not affecting the viability of MRC-5 fibroblasts. Except for garlic and horseradish, the cytotoxic effects of the extracts correlated with their polyphenol content. The examination of changes in the content of ATP and glutathione, in the level of reactive oxygen species, mitochondrial potential and mitochondrial mass did not show a consistent pattern, suggesting that various extracts may act via different mechanisms. Although the extracts’ toxicity to cells in vitro is a first and direct suggestion concerning their possible anticancer effects in vivo, these results point to potential vegetable candidates to become diet components recommended for ovary cancer patients.