Primary oocytes with cellular senescence features are involved in ovarian aging in mice.

In mammalian females, quiescent primordial follicles serve as the ovarian reserve and sustain normal ovarian function and egg production via folliculogenesis. The loss of primordial follicles causes ovarian aging. Cellular senescence, characterized by cell cycle arrest and production of the senescence-associated secretory phenotype (SASP), is associated with tissue aging. In the present study, we report that some quiescent primary oocytes in primordial follicles become senescent in adult mouse ovaries. The senescent primary oocytes share senescence markers characterized in senescent somatic cells. The senescent primary oocytes were observed in young adult mouse ovaries, remained at approximately 15% of the total primary oocytes during ovarian aging from 6 to 12 months, and accumulated in aged ovaries. Administration of a senolytic drug ABT263 to 3-month-old mice reduced the percentage of senescent primary oocytes and the transcription of the SASP factors in the ovary, in addition, led to increased numbers of primordial and total follicles and a higher rate of oocyte maturation. Our study provides experimental evidence that primary oocytes, a germline cell type that is arrested in meiosis, become senescent in adult mouse ovaries and that senescent cell clearance reduced primordial follicle loss and mitigated ovarian aging phenotypes.

Reduced anti-Müllerian hormone action in cumulus-oocyte complexes is beneficial for oocyte maturation without affecting oocyte competence.

Anti-Müllerian hormone (AMH) is a key paracrine/autocrine factor regulating folliculogenesis in the postnatal ovary. As antral follicles mature to the preovulatory stage, AMH production tends to be limited to cumulus cells. Therefore, the present study investigated the role of cumulus cell-derived AMH in supporting maturation and competence of the enclosed oocyte. Cumulus-oocyte complexes (COCs) were isolated from antral follicles of rhesus macaque ovaries for in vitro maturation with or without AMH depletion. Oocyte meiotic status and embryo cleavage after in vitro fertilization were assessed. In vitro maturation with AMH depletion was also performed using COCs from antral follicles of human ovarian tissue. Oocyte maturation and morphology were evaluated. The direct AMH action on mural granulosa cells of the preovulatory follicle was further assessed using human granulosa cells cultured with or without AMH supplementation. More macaque COCs produced metaphase II oocytes with AMH depletion than those of the control culture. However, preimplantation embryonic development after in vitro fertilization was comparable between oocytes derived from COCs cultured with AMH depletion and controls. Oocytes resumed meiosis in human COCs cultured with AMH depletion and exhibited a typical spindle structure. The confluency and cell number decreased in granulosa cells cultured with AMH supplementation relative to the control culture. AMH treatment did not induce cell death in cultured human granulosa cells. Data suggest that reduced AMH action in COCs could be beneficial for oocyte maturation. Cumulus cell-derived AMH is not essential for supporting oocyte competence or mural granulosa cell viability.

Estrogen influences the transzonal projection assembly of cumulus-oocyte complexes through G protein-coupled estrogen receptor during goat follicle development.

Estrogen is an important hormone that plays a role in regulating follicle development and oocyte maturation. Transzonal projections (TZPs) act as communication bridges between follicle somatic cells and oocytes, and their dynamic changes are critical for oocyte development and maturation. However, the roles and mechanisms of estrogen in regulating TZPs during follicular development are not yet understood. We found that the proportion of oocytes spontaneously resuming meiosis increases as the follicle grows, which is accompanied by rising estrogen levels in follicles and decreasing TZPs in cumulus-oocyte complex. To further explore the effect of elevated estrogen levels on TZP assembly, additional estrogen was added to the culture system. The increased estrogen level significantly decreased the mRNA and protein expression levels of TZP assembly-related genes. Subsequent research revealed that TZP regulation by estrogen was mediated by the membrane receptor GPER and downstream ERK1/2 signaling pathway. In summary, our study suggests that estrogen may regulate goat oocyte meiosis arrest by decreasing TZP numbers via estrogen-mediated GPER activation during follicle development.

Single-cell RNA-seq reveals the effects of the FecB mutation on the transcriptome profile in ovine cumulus cells.

Genetic variations in the ovine ovulation rate, which are associated with the FecB mutation, provide useful models by which to explore the mechanisms regulating the development of mammalian antral follicles. In order to study the effects of the FecB mutation on cumulus cell differentiation, preovulatory follicles were aspirated and cumulus cells were isolated from three FecB genotypes (homozygous, heterozygous and wild type) of Small Tail Han (STH) sheep superstimulated with FSH. Transcriptome information from tens of thousands of cumulus cells was determined with the 10 × Genomics single-cell RNA-seq technology. Under the superovulation treatment, the observed number of preovulatory follicles in the ovaries of FecB carriers was still significantly higher than that in the wild-type (P < 0.05). The expression patterns of cumulus cells differed between FecB carriers and wild-type ewes. The screened cumulus cells could also be further divided into different cell clusters, and the differentiation states and fates of each group of cumulus cells also remained different, which supports the notion that heterogeneity in gene expression is prevalent in single cells. The oxidative phosphorylation pathway was significantly enriched in differentially expressed genes among the cell differentiation branch nodes of cumulus cells and among the differentially expressed genes of cumulus cells from the three genotypes. Combined with the important role of oxidative phosphorylation in the maturation of COCs, we suggest that the oxidative phosphorylation pathway of cumulus cells plays a crucial role in the differentiation process of cumulus cells and the mutation effect of the FecB gene.

The structural and functional modularity of ovarian follicle epithelium in the pill-millipede Hyleoglomeris japonica Verhoeff, 1936 (Diplopoda: Glomerida: Glomeridae).

Ovarian somatic tissues typically surround developing oocytes and play a crucial role in oogenesis across various metazoans, often displaying structural properties specific to their functions. However, there is an absence of evident structural modularity in the follicle epithelium of Myriapoda. We report here two structurally and developmentally distinct domains within the follicle epithelium of the Japanese pill millipede, Hyleoglomeris japonica. The follicle epithelium of H. japonica exhibits a thick cell mass at the apex of the follicle. These cells harbor abundant rough endoplasmic reticulum, mitochondria, Golgi complexes, and numerous microvilli, indicative of synthetic/secretory activities. Moreover, their height increases as oogenesis progresses. In contrast, another region of the epithelium lacks these features. Our findings highlight the presence of structural and functional modularity in the follicle epithelium of H. japonica. We suggest classifying the follicle epithelium of Myriapoda into three types: homogenous epithelia with enhanced synthetic activities, homogenous epithelia with diminished such activities, and heterogeneous epithelia with varying synthetic activities. These findings prompt a reevaluation of the nature of ovarian somatic tissues in Myriapoda as well as in Arthropoda.

Mesenchymal stem cells promote ovarian reconstruction in mice.

BACKGROUND: Studies have shown that chemotherapy and radiotherapy can cause premature ovarian failure and loss of fertility in female cancer patients. Ovarian cortex cryopreservation is a good choice to preserve female fertility before cancer treatment. Following the remission of the disease, the thawed ovarian tissue can be transplanted back and restore fertility of the patient. However, there is a risk to reintroduce cancer cells in the body and leads to the recurrence of cancer. Given the low success rate of current in vitro culture techniques for obtaining mature oocytes from primordial follicles, an artificial ovary with primordial follicles may be a good way to solve this problem. METHODS: In the study, we established an artificial ovary model based on the participation of mesenchymal stem cells (MSCs) to evaluate the effect of MSCs on follicular development and oocyte maturation. P2.5 mouse ovaries were digested into single cell suspensions and mixed with bone marrow derived mesenchymal stem cells (BM-MSCs) at a 1:1 ratio. The reconstituted ovarian model was then generated by using phytohemagglutinin. The phenotype and mechanism studies were explored by follicle counting, immunohistochemistry, immunofluorescence, in vitro maturation (IVM), in vitro fertilization (IVF), real-time quantitative polymerase chain reaction (RT-PCR), and Terminal-deoxynucleotidyl transferase mediated nick end labeling(TUNEL) assay. RESULTS: Our study found that the addition of BM-MSCs to the reconstituted ovary can enhance the survival of oocytes and promote the growth and development of follicles. After transplanting the reconstituted ovaries under kidney capsules of the recipient mice, we observed normal folliculogenesis and oocyte maturation. Interestingly, we found that BM-MSCs did not contribute to the formation of follicles in ovarian aggregation, nor did they undergo proliferation during follicle growth. Instead, the cells were found to be located around growing follicles in the reconstituted ovary. When theca cells were labeled with CYP17a1, we found some overlapped staining with green fluorescent protein(GFP)-labeled BM-MSCs. The results suggest that BM-MSCs may participate in directing the differentiation of theca layer in the reconstituted ovary. CONCLUSIONS: The presence of BM-MSCs in the artificial ovary was found to promote the survival of ovarian cells, as well as facilitate follicle formation and development. Since the cells didn't proliferate in the reconstituted ovary, this discovery suggests a potential new and safe method for the application of MSCs in clinical fertility preservation by enhancing the success rate of cryo-thawed ovarian tissues after transplantation.

Age-dependent high-yield isolation of primordial, primary, and early secondary follicles from the bovine ovarian cortex.

In brief: Preantral follicles constitute the largest follicle reserve in the mammalian ovary. This study assesses a mechanical isolation method to maximize the number of follicles retrieved from a defined cortex volume. Abstract: Primordial, primary, and secondary follicles (collectively defined as preantral follicles) constitute the most abundant source of gametes inside the mammalian ovarian cortex. The massive isolation of preantral follicles and the refinement of stage-specific protocols for in vitro follicle growth would provide a powerful tool to boost the rescue and restoration of fertility in assisted reproduction interventions in human medicine, animal breeding, and vulnerable species preservation. Nevertheless, together with an efficient culture system, the most significant limitation to implementing in vitro follicle growth is the lack of an efficient method to isolate viable and homogeneous subpopulations of primordial, primary, and secondary follicles suitable for in vitro culture. Our study provides a strategy for high-yielding mechanical isolation of primordial, primary, and early secondary follicles from a limited portion of the ovarian cortex in the bovine animal model. In the first part of the study, we refined a mechanical isolation protocol of preantral follicles, adopting specific methodological strategies to separate viable and distinct subpopulations of primordial (oblate and prolate forms), primary, and early secondary follicles from 0.16 cm3 of the ovarian cortex. In the second part of the study, we tested the effectiveness of the isolation protocol, considering the individual's age as a critical factor, bearing in mind the progressive decrease in the ovarian reserve that naturally accompanies the reproductive life span. Our study provides a way for designing quantitative and conservative fertility preservation approaches to preserve organ function and minimize the invasiveness of the interventions, also considering age-related differences.

Melatonin protects oogenesis from hypobaric hypoxia-induced fertility damage in mice.

Environmental hypoxia adversely affects reproductive health in humans and animals at high altitudes. Therefore, how to alleviate the follicle development disorder caused by hypoxia exposure and to improve the competence of fertility in plateau non-habituated female animals are important problems to be solved urgently. In this study, a hypobaric hypoxic chamber was used for 4 weeks to simulate hypoxic conditions in female mice, and the effects of hypoxia on follicle development, proliferation and apoptosis of granulosa cells, reactive oxygen species (ROS) levels in MII oocyte and 2-cell rate were evaluated. At the same time, the alleviating effect of melatonin on hypoxic exposure-induced oogenesis damage was evaluated by feeding appropriate amounts of melatonin daily under hypoxia for 4 weeks. The results showed that hypoxia exposure significantly increased the proportion of antral follicles in the ovary, the number of proliferation and apoptosis granulosa cells in the follicle, and the level of ROS in MII oocytes, eventually led to the decline of oocyte quality. However, these defects were alleviated when melatonin was fed under hypoxia conditions. Together, these findings suggest that hypoxia exposure impaired follicular development and reduced oocyte quality, and that melatonin supplementation alleviated the fertility reduction induced by hypoxia exposure.

Cell polarity opposes Jak/STAT-mediated Escargot activation that drives intratumor heterogeneity in a Drosophila tumor model.

In proliferating neoplasms, microenvironment-derived selective pressures promote tumor heterogeneity by imparting diverse capacities for growth, differentiation, and invasion. However, what makes a tumor cell respond to signaling cues differently from a normal cell is not well understood. In the Drosophila ovarian follicle cells, apicobasal-polarity loss induces heterogeneous epithelial multilayering. When exacerbated by oncogenic-Notch expression, this multilayer displays an increased consistency in the occurrence of morphologically distinguishable cells adjacent to the polar follicle cells. Polar cells release the Jak/STAT ligand Unpaired (Upd), in response to which neighboring polarity-deficient cells exhibit a precursor-like transcriptomic state. Among the several regulons active in these cells, we could detect and further validate the expression of Snail family transcription factor Escargot (Esg). We also ascertain a similar relationship between Upd and Esg in normally developing ovaries, where establishment of polarity determines early follicular differentiation. Overall, our results indicate that epithelial-cell polarity acts as a gatekeeper against microenvironmental selective pressures that drive heterogeneity.

Follicle cell contact maintains main body axis polarity in the Drosophila melanogaster oocyte.

In Drosophila melanogaster, the anterior-posterior body axis is maternally established and governed by differential localization of partitioning defective (Par) proteins within the oocyte. At mid-oogenesis, Par-1 accumulates at the oocyte posterior end, while Par-3/Bazooka is excluded there but maintains its localization along the remaining oocyte cortex. Past studies have proposed the need for somatic cells at the posterior end to initiate oocyte polarization by providing a trigger signal. To date, neither the molecular identity nor the nature of the signal is known. Here, we provide evidence that mechanical contact of posterior follicle cells (PFCs) with the oocyte cortex causes the posterior exclusion of Bazooka and maintains oocyte polarity. We show that Bazooka prematurely accumulates exclusively where posterior follicle cells have been mechanically detached or ablated. Furthermore, we provide evidence that PFC contact maintains Par-1 and oskar mRNA localization and microtubule cytoskeleton polarity in the oocyte. Our observations suggest that cell-cell contact mechanics modulates Par protein binding sites at the oocyte cortex.

Knockdown of E1- and E2-ubiquitin enzymes triggers defective chorion biogenesis and modulation of autophagy-related genes in the follicle cells of the vector Rhodnius prolixus.

In insects, the last stage of oogenesis is the process where the chorion layers (eggshell) are synthesized and deposited on the surface of the oocytes by the follicle cells. Protein homeostasis is determined by the fine-tuning of translation and degradation pathways, and the ubiquitin-proteasome system is one of the major degradative routes in eukaryotic cells. The conjugation of ubiquitin to targeted substrates is mediated by the ordered action of E1-activating, E2-conjugating, and E3-ligase enzymes, which covalently link ubiquitin to degradation-targeted proteins delivering them to the proteolytic complex proteasome. Here, we found that the mRNAs encoding polyubiquitin (pUbq), E1, and E2 enzymes are highly expressed in the ovaries of the insect vector of Chagas Disease Rhodnius prolixus. RNAi silencing of pUbq was lethal whereas the silencing of E1 and E2 enzymes resulted in drastic decreases in oviposition and embryo viability. Eggs produced by the E1- and E2-silenced insects presented particular phenotypes of altered chorion ultrastructure observed by high-resolution scanning electron microscopy as well as readings for dityrosine cross-linking and X-ray elemental microanalysis, suggesting a disruption in the secretory routes responsible for the chorion biogenesis. In addition, the ovaries from silenced insects presented altered levels of autophagy-related genes as well as a tendency of upregulation in ER chaperones, indicating a disturbance in the general biosynthetic-secretory pathway. Altogether, we found that E1 and E2 enzymes are essential for chorion biogenesis and that their silencing triggers the modulation of autophagy genes suggesting a coordinated function of both pathways for the progression of choriogenesis.

Expression of steroidogenic enzymes and TGFß superfamily members in follicular cells of prepubertal gilts with distinct endocrine profiles.

Regulation of the transforming growth factor beta (TGFß) superfamily by gonadotrophins in swine follicular cells is not fully understood. This study evaluated the expression of steroidogenic enzymes and members of the TGFß superfamily in prepubertal gilts allocated to three treatments: 1200 IU eCG at D -3 (eCG); 1200 IU eCG at D -6 plus 500 IU hCG at D -3 (eCG + hCG); and the control, composed of untreated gilts. Blood samples and ovaries were collected at slaughter (D0) and follicular cells were recovered thereafter. Relative gene expression was determined by real-time PCR. Serum progesterone levels were greater in the eCG + hCG group compared with the other groups (P < 0.01). No differences were observed in the expression of BMP15, BMPR1A, BMPR2, FSHR, GDF9, LHCGR and TGFBR1 (P > 0.05). Gilts from the eCG group presented numerically greater mean expression of CYP11A1 mRNA than in the control group that approached statistical significance (P = 0.08) and greater expression of CYP19A1 than in both the eCG and the control groups (P < 0.05). Expression of BMPR1B was lower in the eCG + hCG treatment group compared with the control (P < 0.05). In conclusion, eCG treatment increased the relative expression of steroidogenic enzymes, whereas treatment with eCG + hCG increased serum progesterone levels. Although most of the evaluated TGFß members were not regulated after gonadotrophin treatment, the downregulation of BMPR1B observed after treatment with eCG + hCG and suggests a role in luteinization regulation.

Molecular characterization, expression profile and transcriptional regulation of the CYP19 gene in goose ovarian follicles.

Cytochrome P450 Family 19 (CYP19) is a crucial enzyme to catalyze the conversion of androgens to estrogens. However, the regulatory mechanism of goose CYP19 gene remains poorly understood. The present study attempted to obtain the full-length coding sequence (CDS) and 5'-flanking sequence of CYP19 gene, to investigate its expression and distribution profiles in different sized follicles, and to analyze the transcriptional regulatory mechanism of CYP19 gene in goose. Results showed that its CDS consisted of 1512 nucleotides and the encoded amino acid sequence contained a classical P450 structural domain. Homology analysis showed that there were high homologies of nucleotide and amino acid sequences between goose and other avian species. Its promoter sequence spanned from -1925 bp to the transcription start site (ATG) and several transcriptional factors were predicted in this region. Further analysis from luciferase assay showed that the luciferase activity was the highest spanning from -118 to -1 bp by constructing deletion promoter reporter vector. In addition, result from quantitative real-time polymerase chain reaction indicated that the mRNA level of CYP19 gene were highly expressed in theca layer of the fifth largest follicle, and the cellular location was in the theca externa cells by immunohistochemistry. Taken together, it could be concluded that the transcription activity of CYP19 gene was activated by transcriptional factors in its proximal region of promoter to promote the synthesis of estrogens, regulating the selection of pre-hierarchical into hierarchical follicle in goose.

Evaluation of ImageJ software in ultrasonic image analysis: Follicular and luteal morphological characteristics of cattle.

This study was conducted to compare the effectiveness of two methodologies in evaluating B- and Doppler-mode ultrasonic images: analysis using ultrasonic software and utilizing a computer with ImageJ software. To determine if ImageJ software utilization is an efficacious alternative to ultrasonic software device- analysis, there were comparisons of values when using the two methods for morphological and vascular characteristics of follicular dynamics and luteal function in 18 crossbred cattle. From day 8 of an ovarian dynamics synchronization treatment regimen period until the time of ovulation (Day 10), B-mode and power-flow ultrasonic cineloops were obtained every 12 h to assess follicular diameter, wall area, and wall blood perfusion area. On Day 14 after ovulation, US cineloops of ovaries were obtained in B mode and power flow to evaluate various morphological and vascular characteristics of the corpus luteum (CL), including luteal diameter, luteal area, and CL blood perfusion area. Cineloops were evaluated and analyzed using ultrasonic software, and in a computer with ImageJ software. To evaluate consistency in results between the two methods, there was evaluation utilizing paired t-test, Pearson correlation coefficient, Bland-Altman plot, and Linear Regression Test procedures to calculate proportion of bias between values for measurements of variables evaluated. Results indicated none of the values for variables before and after ovulation differed (P > 0.05). It, therefore, was concluded that utilization of ImageJ software is an efficacious biomedical technique to analyze ultrasonic images of morphological and vascular characteristics before and after ovulation in cattle.

El consumo semicrónico moderado de alcohol altera la foliculogénesis y la calidad núcleo-citoplasmática oocitaria, en el ratón / Moderate semi-chronic alcohol consumption alters folliculogenesis and oocyte nuclear-cytoplasmic quality in mice

Resumen El consumo crónico de alcohol es un problema de salud mundial que afecta particularmente a la población femenina. Sin embargo, los efectos de la ingesta semicrónica en cantidades moderadas a bajas en el ovario y el oocito son poco conocidos. En un modelo murino, se administró etanol al 10% en agua de bebida (hembras tratadas) o agua (hembras control) por 15 días, y luego de la superovulación o no (ovulación espontánea), se analizó el ciclo estral y la calidad ovárico-gamética. En las hembras tratadas, la frecuencia y duración del diestro aumentó, y las frecuencias de folículos y cuerpos lúteos disminuyeron vs hembras controles, valores que se restauraron luego de la superovulación. Sin embargo, en las hembras tratadas, la tasa de proliferación celular folicular y el desbalance de la expresión ovárica de VEGF (factor de crecimiento endotelial) persistieron luego de la superovulación. El número de ovocitos ovulados con metafase II anormal, fragmentados y activados partenogenéticamente fue mayor en las hembras tratadas respecto las controles. En conclusión, el consumo semicrónico moderado de alcohol produce anestro, ciclo estral irregular, foliculogénesis deficiente y anomalías núcleo-citoplasmáticas en los oocitos ovulados. Estas alteraciones podrían constituirse en un factor etiológico de pérdida gestacional temprana y desarrollo embrionario anormal luego del consumo de alcohol.

Abstract Chronic alcohol consumption is a global health problem that particularly affects the female population. However, the ef-fects of semi-chronic ethanol intake in low-moderate amounts on the ovary and oocyte are poorly understood. In a mouse model, 10% ethanol was administered in drinking water (treated females) or water (control females) for 15 days, and after superovulation or not (spontaneous ovulation), the estrous cycle and ovarian-gametic quality were analyzed. In treated females, the frequency and duration of the diestrus increased, and the frequencies of follicles and corpus luteum decreased vs control females, values that restored after superovulation. However, in treated females, the follicular cell proliferation rate and the imbalance in ovarian expression of VEGF (endothelial growth factor) persisted after superovulation. The number of ovulated oocytes with abnormal metaphase II, fragmented and parthenogenetically activated was higher in treated females than in control ones. In conclusion, moderate semi-chronic alcohol consumption produces anestrum, irregular estrous cycle, poor folliculogenesis, and nuclear-cytoplasmic abnormalities in ovulated oocytes. These alterations could constitute an etiological factor of early gestational loss and abnormal embryonic development after alcohol consumption.

Maintenance of quiescent oocytes by noradrenergic signals.

All females adopt an evolutionary conserved reproduction strategy; under unfavorable conditions such as scarcity of food or mates, oocytes remain quiescent. However, the signals to maintain oocyte quiescence are largely unknown. Here, we report that in four different species - Caenorhabditis elegans, Caenorhabditis remanei, Drosophila melanogaster, and Danio rerio - octopamine and norepinephrine play an essential role in maintaining oocyte quiescence. In the absence of mates, the oocytes of Caenorhabditis mutants lacking octopamine signaling fail to remain quiescent, but continue to divide and become polyploid. Upon starvation, the egg chambers of D. melanogaster mutants lacking octopamine signaling fail to remain at the previtellogenic stage, but grow to full-grown egg chambers. Upon starvation, D. rerio lacking norepinephrine fails to maintain a quiescent primordial follicle and activates an excessive number of primordial follicles. Our study reveals an evolutionarily conserved function of the noradrenergic signal in maintaining quiescent oocytes.

Equine Chorionic Gonadotropin as an Effective FSH Replacement for In Vitro Ovine Follicle and Oocyte Development.

The use of assisted reproductive technologies (ART) still requires strategies through which to maximize individual fertility chances. In vitro folliculogenesis (ivF) may represent a valid option to convey the large source of immature oocytes in ART. Several efforts have been made to set up ivF cultural protocols in medium-sized mammals, starting with the identification of the most suitable gonadotropic stimulus. In this study, Equine Chorionic Gonadotropin (eCG) is proposed as an alternative to Follicle Stimulating Hormone (FSH) based on its long superovulation use, trans-species validation, long half-life, and low costs. The use of 3D ivF on single-ovine preantral (PA) follicles allowed us to compare the hormonal effects and to validate their influence under two different cultural conditions. The use of eCG helped to stimulate the in vitro growth of ovine PA follicles by maximizing its influence under FBS-free medium. Higher performance of follicular growth, antrum formation, steroidogenic activity and gap junction marker expression were recorded. In addition, eCG, promoted a positive effect on the germinal compartment, leading to a higher incidence of meiotic competent oocytes. These findings should help to widen the use of eCG to ivF as a valid and largely available hormonal support enabling a synchronized in vitro follicle and oocyte development.

Single-Cell Transcriptomics Analysis of Human Small Antral Follicles.

Human ovarian folliculogenesis is a highly regulated and complex process. Characterization of follicular cell signatures during this dynamic process is important to understand follicle fate (to grow, become dominant, or undergo atresia). The transcriptional signature of human oocytes and granulosa cells (GCs) in early-growing and ovulatory follicles have been previously described; however, that of oocytes with surrounding GCs in small antral follicles have not been studied yet. Here, we have generated a unique dataset of single-cell transcriptomics (SmartSeq2) consisting of the oocyte with surrounding GCs from several individual (non-dominant) small antral follicles isolated from adult human ovaries. We have identified two main types of (healthy) follicles, with a distinct oocyte and GC signature. Using the CellphoneDB algorithm, we then investigated the bi-directional ligand-receptor interactions regarding the transforming growth factor-ß (TGFß)/bone morphogenetic protein (BMP), wingless-type (MMTV)-integration site (WNT), NOTCH, and receptor tyrosine kinases (RTK) signaling pathways between oocyte and GCs within each antral follicle type. Our work not only revealed the diversity of small antral follicles, but also contributes to fill the gap in mapping the molecular landscape of human folliculogenesis and oogenesis.

Ovarian Folliculogenesis and Uterine Endometrial Receptivity after Intermittent Vaginal Injection of Recombinant Human Follicle-Stimulating Hormone in Infertile Women Receiving In Vitro Fertilization and in Immature Female Rats.

The uterine first-pass effect occurs when drugs are delivered vaginally. However, the effect of vaginally administered recombinant human follicle-stimulating hormone (rhFSH) on ovarian folliculogenesis and endometrial receptivity is not well established. We aimed to compare the efficacy of rhFSH administered vaginally and abdominally in clinical in vitro fertilization (IVF) treatment, pharmacokinetic study, and animal study. In IVF treatment, the number of oocytes retrieved, endometrial thickness and uterine artery blood perfusion were not different between women who received the rhFSH either vaginally or abdominally. For serum pharmacokinetic parameters, significantly lower Tmax, clearance, and higher AUC and T1/2_elimination of rhFSH were observed in women who received rhFSH vaginally, but urine parameters were not different. Immature female rats that received daily abdominal or vaginal injections (1 IU twice daily for 4 days) or intermittent vaginal injections (4 IU every other day for two doses) of rhFSH had more total follicles than the control group. In addition, the serum progesterone and progesterone receptors in the local endometrium were significantly higher in the groups treated with intermittent abdominal or vaginal injection of rhFSH, compared with those who recieved daily injection. In summary, vaginal administration of rhFSH may provide an alternative treatment regimen in women receiving IVF.