Spatiotemporal transcriptomic changes of human ovarian aging and the regulatory role of FOXP1.

Limited understanding exists regarding how aging impacts the cellular and molecular aspects of the human ovary. This study combines single-cell RNA sequencing and spatial transcriptomics to systematically characterize human ovarian aging. Spatiotemporal molecular signatures of the eight types of ovarian cells during aging are observed. An analysis of age-associated changes in gene expression reveals that DNA damage response may be a key biological pathway in oocyte aging. Three granulosa cells subtypes and five theca and stromal cells subtypes, as well as their spatiotemporal transcriptomics changes during aging, are identified. FOXP1 emerges as a regulator of ovarian aging, declining with age and inhibiting CDKN1A transcription. Silencing FOXP1 results in premature ovarian insufficiency in mice. These findings offer a comprehensive understanding of spatiotemporal variability in human ovarian aging, aiding the prioritization of potential diagnostic biomarkers and therapeutic strategies.

Efficacy and safety of mesenchymal stem cell therapy for ovarian ageing in a mouse model.

BACKGROUND: Ovarian ageing is one of the major issues that impacts female fertility. Mesenchymal stem cell (MSC)-based therapy has made impressive progress in recent years. However, the efficacy and safety of MSCs, as nonautologous components, remain to be further verified. METHODS: Two common sources of MSCs, umbilical cord-derived MSCs (UC-MSCs) and adipose tissue-derived MSCs (AD-MSCs), were orthotopically transplanted into a mouse model of ovarian ageing to evaluate their therapeutic effects. The safety of the treatment was further evaluated, and RNA sequencing was performed to explore the underlying mechanisms involved. RESULTS: After orthotopic transplantation of MSCs into the ovary, the oestrous cycle, ovarian weight, number and proportion of primary follicles, granulosa cell proliferation, and angiogenesis were improved. The effects of AD-MSCs were superior to those of UC-MSCs in several indices, such as post-transplant granulosa cell proliferation, ovarian weight and angiogenesis. Moreover, the tumorigenesis, acute toxicity, immunogenicity and biodistribution of MSCs were evaluated, and both AD-MSCs and UC-MSCs were found to possess high safety profiles. Through RNA sequencing analysis, enhancement of the MAPK cascade was observed, and long-term effects were mainly linked to the activation of immune function. CONCLUSIONS: Orthotopic transplantation of MSCs displays significant efficacy and high safety for the treatment of ovarian ageing in mice.

The adipokines progranulin and omentin - novel regulators of basic ovarian cell functions.

The present study aimed to examine the effects of progranulin and omentin on basic ovarian cell functions. For this purpose, we investigated the effects of the addition of progranulin and omentin (0, 0.1, 1, or 10 ng/ml) on the viability, proliferation, apoptosis and steroidogenesis of cultured rabbit ovarian granulosa cells. To determine the importance of the interrelationships between granulosa cells and theca cells, we compared the influence of progranulin and omentin on progesterone and estradiol release in cultured granulosa cells and ovarian fragments containing both granulosa cells and theca cells. Cell viability, proliferation, cytoplasmic apoptosis and release of progesterone and estradiol were measured by Cell Counting Kit-8 (CCK-8), BrdU incorporation, cell death detection, and ELISA. Both progranulin and omentin increased granulosa cell viability and proliferation and decreased apoptosis. Progranulin increased progesterone release by granulosa cells but reduced progesterone output by ovarian fragments. Progranulin decreased estradiol release by granulosa cells but increased it in ovarian fragments. Omentin reduced progesterone release in both models. Omentin reduced estradiol release by granulosa cells but promoted this release in ovarian fragments. The present observations are the first to demonstrate that progranulin and omentin can be direct regulators of basic ovarian cell functions. Furthermore, the differences in the effects of these adipokines on steroidogenesis via granulosa and ovarian fragments indicate that these peptides could target both granulosa and theca cells.

Cellular atlas of the human ovary using morphologically guided spatial transcriptomics and single-cell sequencing.

The reproductive and endocrine functions of the ovary involve spatially defined interactions among specialized cell populations. Despite the ovary's importance in fertility and endocrine health, functional attributes of ovarian cells are largely uncharacterized. Here, we profiled >18,000 genes in 257 regions from the ovaries of two premenopausal donors to examine the functional units in the ovary. We also generated single-cell RNA sequencing data for 21,198 cells from three additional donors and identified four major cell types and four immune cell subtypes. Custom selection of sampling areas revealed distinct gene activities for oocytes, theca, and granulosa cells. These data contributed panels of oocyte-, theca-, and granulosa-specific genes, thus expanding the knowledge of molecular programs driving follicle development. Serial samples around oocytes and across the cortex and medulla uncovered previously unappreciated variation of hormone and extracellular matrix remodeling activities. This combined spatial and single-cell atlas serves as a resource for future studies of rare cells and pathological states in the ovary.

Long noncoding RNAs and mRNAs profiling in ovary during laying and broodiness in Taihe Black-Bone Silky Fowls (Gallus gallus Domesticus Brisson).

BACKGROUND: Broodiness significantly impacts poultry egg production, particularly notable in specific breeds such as the black-boned Silky, characterized by pronounced broodiness. An understanding of the alterations in ovarian signaling is essential for elucidating the mechanisms that influence broodiness. However, comparative research on the characteristics of long non-coding RNAs (lncRNAs) in the ovaries of broody chickens (BC) and high egg-laying chickens (GC) remains scant. In this investigation, we employed RNA sequencing to assess the ovarian transcriptomes, which include both lncRNAs and mRNAs, in eight Taihe Black-Bone Silky Fowls (TBsf), categorized into broody and high egg-laying groups. This study aims to provide a clearer understanding of the genetic underpinnings associated with broodiness and egg production. RESULTS: We have identified a total of 16,444 mRNAs and 18,756 lncRNAs, of which 349 mRNAs and 651 lncRNAs exhibited significantly different expression (DE) between the BC and GC groups. Furthermore, we have identified the cis-regulated and trans-regulated target genes of differentially abundant lncRNA transcripts and have constructed an lncRNA-mRNA trans-regulated interaction network linked to ovarian follicle development. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) annotation analyses have revealed that DE mRNAs and the target genes of DE lncRNAs are associated with pathways including neuroactive ligand-receptor interaction, CCR6 chemokine receptor binding, G-protein coupled receptor binding, cytokine-cytokine receptor interaction, and ECM-receptor interaction. CONCLUSION: Our research presents a comprehensive compilation of lncRNAs and mRNAs linked to ovarian development. Additionally, it establishes a predictive interaction network involving differentially abundant lncRNAs and differentially expressed genes (DEGs) within TBsf. This significantly contributes to our understanding of the intricate interactions between lncRNAs and genes governing brooding behavior.

ERß Regulation of Indian Hedgehog Expression in the First Wave of Ovarian Follicles.

Increased activation of ovarian primordial follicles in Erß knockout (ErßKO) rats becomes evident as early as postnatal day 8.5. To identify the ERß-regulated genes that may control ovarian primordial follicle activation, we analyzed the transcriptome profiles of ErßKO rat ovaries collected on postnatal days 4.5, 6.5, and 8.5. Compared to wildtype ovaries, ErßKO ovaries displayed dramatic downregulation of Indian hedgehog (Ihh) expression. IHH-regulated genes, including Hhip, Gli1, and Ptch1, were also downregulated in ErßKO ovaries. This was associated with a downregulation of steroidogenic enzymes Cyp11a1, Cyp19a1, and Hsd17b1. The expression of Ihh remained very low in ErßKO ovaries despite the high levels of Gdf9 and Bmp15, which are known upregulators of Ihh expression in the granulosa cells of activated ovarian follicles. Strikingly, the downregulation of the Ihh gene in ErßKO ovaries began to disappear on postnatal day 16.5 and recovered on postnatal day 21.5. In rat ovaries, the first wave of primordial follicles is rapidly activated after their formation, whereas the second wave of primordial follicles remains dormant in the ovarian cortex and slowly starts activating after postnatal day 12.5. We localized the expression of Ihh mRNA in postnatal day 8.5 wildtype rat ovaries but not in the age-matched ErßKO ovaries. In postnatal day 21.5 ErßKO rat ovaries, we detected Ihh mRNA mainly in the activated follicles in the ovaries' peripheral regions. Our findings indicate that the expression of Ihh in the granulosa cells of the activated first wave of ovarian follicles depends on ERß.

Changes in ovarian tissue structure and distribution of oestrogen receptors in Huanghuai goats at different ages.

To observe developmental changes in the ovarian tissue structure and distribution characteristics of oestrogen receptors (ERs) in the ovaries of Huanghuai goats at different ages, we selected healthy Huanghuai goats ewes and divided them into five groups (i.e. 3-, 30-, 60-, 90- and 120-day-old groups), with 10 animals in each group. The serum was separated after blood collection through the jugular vein, and the contents of oestrogen (E) and progesterone (P) in the serum of Huanghuai goats at each age were determined. Three goats were randomly selected from each group and sacrificed after anaesthesia, and the ovarian tissue was quickly obtained and placed in 4% paraformaldehyde fixative to prepare the tissue sections. Using HE, oestrogen receptors were immunohistochemically stained and observed. These results showed many primordial follicles and occasional secondary follicles in the ovaries of 3-day-old Huanghuai goats. Ovarian reticular structures were observed in 30-day-old ovarian medulla, with occasional near-mature growing follicles. Mature follicles and corpus luteum were occasionally detected in 60-day-old ovarian cortex. The 90-120-day-old ovarian cortices contained growing and mature follicles, and the number of mature follicles and corpora lutea increased, implying a significant luteal involution period. The E and P contents in the 120-day-old group were significantly higher than those in the 3-, 30-, 60- and 90-day-old groups. The levels of ERα and ERß in the 3- and 30-day-old groups were mainly distributed in the granulosa cells of ovarian reproductive epithelial cells, primordial follicles, atretic follicles, and primary and secondary follicles. The ERα and ERß levels of the 60-, 90- and 120-day-old groups were also distributed in the granulosa cells and luteal cells of mature follicles, especially in the 120-day-old endometrial cells of mature follicles, where ERß was distributed significantly. The overall expression of ERß in the ovary was higher than that of ERα. The results of this study provide basic data on the ovarian development and the specific expression of ERs and PRs in the ovaries of Huanghuai white goats, which play an important role in ovarian development and precocity.

Neurotensin modulates ovarian vascular permeability via adherens junctions.

Neurotensin (NTS) is a 13-amino acid peptide which is highly expressed in the mammalian ovary in response to the luteinizing hormone surge. Antibody neutralization of NTS in the ovulatory follicle of the cynomolgus macaque impairs ovulation and induces follicular vascular dysregulation, with excessive pooling of red blood cells in the follicle antrum. We hypothesize that NTS is an essential intrafollicular regulator of vascular permeability. In the present study, follicle injection of the NTS receptor antagonist SR142948 also resulted in vascular dysregulation. To measure vascular permeability changes in vitro, primary macaque ovarian microvascular endothelial cells (mOMECs) were enriched from follicle aspirates and studied in vitro. When treated with NTS, permeability of mOMECs decreased. RNA sequencing (RNA-Seq) of mOMECs revealed high mRNA expression of the permeability-regulating adherens junction proteins N-cadherin (CDH2) and K-cadherin (CDH6). Immunofluorescent detection of CDH2 and CDH6 confirmed expression and localized these cadherins to the cell-cell boundaries, consistent with function as components of adherens junctions. mOMECs did not express detectable levels of the typical vascular endothelial cadherin, VE-cadherin (CDH5) as determined by RNA-Seq, qPCR, western blot, and immunofluorescence. Knockdown of CDH2 or CDH6 via siRNA abrogated the NTS effect on mOMEC permeability. Collectively, these data suggest that NTS plays an ovulation-critical role in vascular permeability maintenance, and that CDH2 and CDH6 are involved in the permeability modulating effect of NTS on the ovarian microvasculature. NTS can be added to a growing number of angiogenic regulators which are critical for successful ovulation.

Transcriptome analysis during 4-vinylcyclohexene diepoxide exposure-induced premature ovarian insufficiency in mice.

The occupational chemical 4-Vinylcyclohexene diepoxide (VCD) is a reproductively toxic environmental pollutant that causes follicular failure, leading to premature ovarian insufficiency (POI), which significantly impacts a woman's physical health and fertility. Investigating VCD's pathogenic mechanisms can offer insights for the prevention of ovarian impairment and the treatment of POI. This study established a mouse model of POI through intraperitoneal injection of VCD into female C57BL/6 mice for 15 days. The results were then compared with those of the control group, including a comparison of phenotypic characteristics and transcriptome differences, at two time points: day 15 and day 30. Through a comprehensive analysis of differentially expressed genes (DEGs), key genes were identified and validated some using RT-PCR. The results revealed significant impacts on sex hormone levels, follicle number, and the estrous cycle in VCD-induced POI mice on both day 15 and day 30. The DEGs and enrichment results obtained on day 15 were not as significant as those obtained on day 30. The results of this study provide a preliminary indication that steroid hormone synthesis, DNA damage repair, and impaired oocyte mitosis are pivotal in VCD-mediated ovarian dysfunction. This dysfunction may have been caused by VCD damage to the primordial follicular pool, impairing follicular development and aggravating ovarian damage over time, making it gradually difficult for the ovaries to perform their normal functions.

Molybdenum and cadmium co-induce necroptosis through Th1/Th2 imbalance-mediated endoplasmic reticulum stress in duck ovaries.

Cadmium (Cd) and excess molybdenum (Mo) pose serious threats to animal health. Our previous study has determined that Cd and/or Mo exposure can cause ovarian damage of ducks, while the specific mechanism is still obscure. To further investigate the toxic mechanism of Cd and Mo co-exposure in the ovary, forty 8-day-old female ducks were randomly allocated into four groups for 16 weeks, and the doses of Cd and Mo in basic diet per kg were as follows: control group, Mo group (100 mg Mo), Cd group (4 mg Cd), and Mo + Cd group (100 mg Mo + 4 mg Cd). Cadmium sulfate 8/3-hydrate (CdSO4·8/3H2O) and hexaammonium molybdate ((NH4)6Mo7O24·4H2O) were the origins of Cd and Mo, respectively. At the 16th week of the experiment, all ovary tissues were collected for the detection of related indexes. The data indicated that Mo and/or Cd induced trace element disorders and Th1/Th2 balance to divert toward Th1 in the ovary, which activated endoplasmic reticulum (ER) stress and then provoked necroptosis through triggering RIPK1/RIPK3/MLKL signaling pathway, and eventually caused ovarian pathological injuries and necroptosis characteristics. The alterations of above indicators were most apparent in the joint group. Above all, this research illustrates that Mo and/or Cd exposure can initiate necroptosis through Th1/Th2 imbalance-modulated ER stress in duck ovaries, and Mo and Cd combined exposure aggravates ovarian injuries. This research explores the molecular mechanism of necroptosis caused by Mo and/or Cd, which reveals that ER stress attenuation may be a therapeutic target to alleviate necroptosis.

VCD-induced menopause mouse model reveals reprogramming of hepatic metabolism.

OBJECTIVE: Menopause adversely impacts systemic energy metabolism and increases the risk of metabolic disease(s) including hepatic steatosis, but the mechanisms are largely unknown. Dosing female mice with vinyl cyclohexene dioxide (VCD) selectively causes follicular atresia in ovaries, leading to a murine menopause-like phenotype. METHODS: In this study, we treated female C57BL6/J mice with VCD (160 mg/kg i.p. for 20 consecutive days followed by verification of the lack of estrous cycling) to investigate changes in body composition, energy expenditure (EE), hepatic mitochondrial function, and hepatic steatosis across different dietary conditions. RESULTS: VCD treatment induced ovarian follicular loss and increased follicle-stimulating hormone (FSH) levels in female mice, mimicking a menopause-like phenotype. VCD treatment did not affect body composition, or EE in mice on a low-fat diet (LFD) or in response to a short-term (1-week) high-fat, high sucrose diet (HFHS). However, the transition to a HFHS lowered cage activity in VCD mice. A chronic HFHS diet (16 weeks) significantly increased weight gain, fat mass, and hepatic steatosis in VCD-treated mice compared to HFHS-fed controls. In the liver, VCD mice showed suppressed hepatic mitochondrial respiration on LFD, while chronic HFHS resulted in compensatory increases in hepatic mitochondrial respiration. Also, liver RNA sequencing revealed that VCD promoted global upregulation of hepatic lipid/cholesterol synthesis pathways. CONCLUSION: Our findings suggest that the VCD-induced menopause model compromises hepatic mitochondrial function and lipid/cholesterol homeostasis that sets the stage for HFHS diet-induced steatosis while also increasing susceptibility to obesity.

Genome-Wide DNA Methylation Analysis and Functional Validation of Litter Size Traits in Jining Grey Goats.

DNA methylation (DNAm) is associated with the reproductive system. However, the genetic mechanism through which DNAm regulates gene expression and thus affects litter size in goats is unclear. Therefore, in the present work, genome-wide DNAm profiles of HP and LP Jining Grey goat ovary tissues were comprehensively analyzed via WGBS, and RNA-Seq data were combined to identify candidate genes associated with litter size traits in the Jining Grey goat. Finally, BSP and RT-qPCR were used to verify the sequencing results of the key genes. Notably, the DNMT genes were downregulated at the expression level in the HP group. Both groups exhibited comparable levels of methylation. A total of 976 differentially methylated regions (DMRs) (973 DMRs for CG and 3 DMRs for CHG) and 310 differentially methylated genes (DMGs) were identified in this study. Through integration of WGBS and RNA-Seq data, we identified 59 differentially methylated and differentially expressed genes (DEGs) and ultimately screened 8 key DMGs (9 DMRS) associated with litter size traits in Jining Grey goats (SERPINB2: chr24_62258801_62259000, NDRG4: chr18_27599201_27599400, CFAP43: chr26_27046601_27046800, LRP1B. chr2_79720201_79720400, EPHA6: chr1_40088601_40088800, TTC29: chr17_59385801_59386000, PDE11A: chr2_117418601_117418800 and PGF: chr10_ 16913801_16914000 and chr10_16916401_16916600). In summary, our research comprehensively analyzed the genome-wide DNAm profiles of HP and LP Jining Grey goat ovary tissues. The data findings suggest that DNAm in goat ovaries may play an important role in determining litter size.

Loss of ERß Disrupts Gene Regulation in Primordial and Primary Follicles.

Loss of ERß increases primordial follicle growth activation (PFGA), leading to premature ovarian follicle reserve depletion. We determined the expression and gene regulatory functions of ERß in dormant primordial follicles (PdFs) and activated primary follicles (PrFs) using mouse models. PdFs and PrFs were isolated from 3-week-old Erß knockout (Erßnull) mouse ovaries, and their transcriptomes were compared with those of control Erßfl/fl mice. We observed a significant (≥2-fold change; FDR p-value ≤ 0.05) deregulation of approximately 5% of genes (866 out of 16,940 genes, TPM ≥ 5) in Erßnull PdFs; ~60% (521 out of 866) of the differentially expressed genes (DEGs) were upregulated, and 40% were downregulated, indicating that ERß has both transcriptional enhancing as well as repressing roles in dormant PdFs. Such deregulation of genes may make the Erßnull PdFs more susceptible to increased PFGA. When the PdFs undergo PFGA and form PrFs, many new genes are activated. During PFGA of Erßfl/fl follicles, we detected a differential expression of ~24% genes (4909 out of 20,743; ≥2-fold change; FDR p-value ≤ 0.05; TPM ≥ 5); 56% upregulated and 44% downregulated, indicating the gene enhancing and repressing roles of Erß-activated PrFs. In contrast, we detected a differential expression of only 824 genes in Erßnull follicles during PFGA (≥2-fold change; FDR p-value ≤ 0.05; TPM ≥ 5). Moreover, most (~93%; 770 out of 824) of these DEGs in activated Erßnull PrFs were downregulated. Such deregulation of genes in Erßnull activated follicles may impair their inhibitory role on PFGA. Notably, in both Erßnull PdFs and PrFs, we detected a significant number of epigenetic regulators and transcription factors to be differentially expressed, which suggests that lack of ERß either directly or indirectly deregulates the gene expression in PdFs and PrFs, leading to increased PFGA.

A pattern recognition receptor interleukin-1 receptor is involved in reproductive immunity in Macrobrachium nipponense ovary.

The family of TIR domain-containing receptors includes numerous proteins involved in innate immunity. In this study, a member of this family was characterized from the ovary of the oriental river prawn Macrobrachium nipponense and identified as interleukin-1 receptor (MnIL-1R). Meanwhile, to elucidate the conservation of IL-1R, its orthologous were identified in several crustacean species as well. In addition, the expression pattern of MnIL-1R in various adult tissues and post different pathogen-associated molecular patterns (PAMPs) challenge in ovary was analyzed with qRT-PCR technology. Finally, the roles of MnIL-1R in the ovary were analyzed by RNAi technology. The main results are as follows: (1) MnIL-1R comprises a 1785 bp ORF encoding 594 amino acids and is structurally composed of five domains: a signal peptide, two immunoglobulin (IG) domains, a transmembrane region, and a TIR-2 domain; (2) the TIR domain showed a high conservation among analyzed crustacean species; (3) MnIL-1R is widely detected in all tested tissues including ovary; (4) MnIL-1R showed a positive response to challenges with LPS, PGN, and polyI:C in the ovary; (5) its IG domain showed strong binding ability to LPS and PGN, confirming its role as a pattern recognition receptor; (6) the expression patterns of several members of the Toll signaling pathway (Myd88, TRAF-6, Dorsal, and Relish) was similar to that of MnIL-1R after challenges with LPS, PGN, and polyI:C in the ovary; (7) the silencing of MnIL-1R resulted in down-regulation of theses gene' (Myd88, TRAF-6, Dorsal, and Relish) expression level in the ovary. These results suggest that MnIL-1R can activate the Toll signaling pathway in the ovary by directly recognizing LPS and PGN through its IG domain, thereby contributing to the immune response in the ovary of M. nipponense.

The spatio-temporal distribution of aromatase cytochrome in ovary throughout the canine oestrous cycle.

CONTEXT: New animal welfare legislation and ethical guidelines encourage alternative approaches for canine contraception, instead of surgical gonadectomy which is considered invasive and unjustified in healthy dogs. AIMS: Reversible contraception might be achieved by inhibition of aromatase (CYP19), an enzyme catalysing the conversion of androgens to oestrogens. This study provides insights into the spatio-temporal expression and distribution of aromatase in canine ovarian tissue. METHODS: Ovarian tissue was collected from 39 healthy and sexually mature bitches during different stages of the oestrous cycle: pro-oestrus (n =8), oestrus (n =12), dioestrus (n =9) (luteal phase) and anoestrus (n =10). Localisation of cytochrome P450 aromatase was determined by immunohistochemistry. KEY RESULTS: Aromatase activity in the dog is high during pro-oestrus, ovulation and early dioestrus. Comparing types of follicles and corpora lutea, the highest aromatase abundance was found in antral follicles and luteinising follicles, whereas corpora lutea and early antral follicles showed an intermediate presence of the enzyme. Interesting was the high abundance of aromatase in luteinising theca interna cells, prevailing over granulosa cells. CONCLUSIONS AND IMPLICATIONS: Understanding of cells involved in oestradiol production is important for targeted inhibition of oestradiol synthesis, possibly offering an approach for contraception and suppression of oestrus.

Single microcystin exposure impairs the hypothalamic-pituitary-gonadal axis at different levels in female rats.

Microcystin (MC) is most common cyanobacterial toxin. Few studies have evaluated the MC effects on the hypothalamic-pituitary-gonadal (HPG) axis and metabolic function. In this study, we assessed whether MC exposure results in HPG axis and metabolic changes. Female rats were exposed to a single dose of MC at environmentally relevant levels (5, 20 and 40 µg/kg). After 24 h, we evaluated reproductive and metabolic parameters for 15 days. MC reduced the hypothalamic GnRH protein expression, increased the pituitary protein expression of GnRHr and IL-6. MC reduced LH levels and increased FSH levels. MC reduced the primary follicles, increased the corpora lutea, elevated levels of anti-Müllerian hormone (AMH) and progesterone, and decreased estrogen levels. MC increased ovarian VEGFr, LHr, AMH, ED1, IL-6 and Gp91-phox protein expression. MC increased uterine area and reduced endometrial gland number. A blunted estrogen-negative feedback was observed in MC rats after ovariectomy, with no changes in LH levels compared to intact MC rats. Therefore, these data suggest that a MC leads to abnormal HPG axis function in female rats.

Communication between the stem cell niche and an adjacent differentiation niche through miRNA and EGFR signaling orchestrates exit from the stem cell state in the Drosophila ovary.

The signaling environment, or niche, often governs the initial difference in behavior of an adult stem cell and a derivative that initiates a path towards differentiation. The transition between an instructive stem cell niche and differentiation niche must generally have single-cell resolution, suggesting that multiple mechanisms might be necessary to sharpen the transition. Here, we examined the Drosophila ovary and found that Cap cells, which are key constituents of the germline stem cell (GSC) niche, express a conserved microRNA (miR-124). Surprisingly, loss of miR-124 activity in Cap cells leads to a defect in differentiation of GSC derivatives. We present evidence that the direct functional target of miR-124 in Cap cells is the epidermal growth factor receptor (EGFR) and that failure to limit EGFR expression leads to the ectopic expression of a key anti-differentiation BMP signal in neighboring somatic escort cells (ECs), which constitute a differentiation niche. We further found that Notch signaling connects EFGR activity in Cap cells to BMP expression in ECs. We deduce that the stem cell niche communicates with the differentiation niche through a mechanism that begins with the selective expression of a specific microRNA and culminates in the suppression of the major anti-differentiation signal in neighboring cells, with the functionally important overall role of sharpening the spatial distinction between self-renewal and differentiation environments.

FOXL2 interaction with different binding partners regulates the dynamics of ovarian development.

The transcription factor FOXL2 is required in ovarian somatic cells for female fertility. Differential timing of Foxl2 deletion, in embryonic versus adult mouse ovary, leads to distinctive outcomes, suggesting different roles across development. Here, we comprehensively investigated FOXL2's role through a multi-omics approach to characterize gene expression dynamics and chromatin accessibility changes, coupled with genome-wide identification of FOXL2 targets and on-chromatin interacting partners in somatic cells across ovarian development. We found that FOXL2 regulates more targets postnatally, through interaction with factors regulating primordial follicle formation and steroidogenesis. Deletion of one interactor, ubiquitin-specific protease 7 (Usp7), results in impairment of somatic cell differentiation, germ cell nest breakdown, and ovarian development, leading to sterility. Our datasets constitute a comprehensive resource for exploration of the molecular mechanisms of ovarian development and causes of female infertility.

miR-210 promotes immune- and suppresses oocyte meiosis-related genes in the zebrafish ovarian cells.

microRNA-210 (miRNA), a well-documented miRNA, has been implicated in a myriad of biological processes, including responses to hypoxia, angiogenesis, cell proliferation, and male infertility in humans. However, a comprehensive understanding of its functions in fish requires further investigation. This study pursued to elucidate the downstream effect of dre-miR-210-5p on primary ovarian cell culture in zebrafish (Danio rerio), an animal model. A protocol was settled down by incubations with either an miR-210 mimic or a scrambled miRNA in the isolated ovaries. RNA-sequencing analysis identified ∼6000 differentially expressed target genes revealing that downregulated genes were associated with reproduction-related pathways while immune-related pathways displayed an upregulated pattern. To identify molecular markers, predicted target genes were classified into reproduction and immune cell types. These findings underscore the existence of a profound interplay between the reproductive and immune systems, with miR-210 emerging as a pivotal player in orchestrating transcriptomic alterations within fish ovaries.

Corpus luteum presence in the bovine ovary increase intrafollicular progesterone concentration: consequences in follicular cells gene expression and follicular fluid small extracellular vesicles miRNA contents.

BACKGROUND: It is well described that circulating progesterone (P4) plays a key role in several reproductive events such as oocyte maturation. However, during diestrus, when circulating P4 is at the highest concentrations, little is known about its local impact on the follicular cells such as intrafollicular P4 concentration due to corpus luteum (CL) presence within the same ovary. Based on that, our hypothesis is that the CL presence in the ovary during diestrus alters intrafollicular P4 concentrations, oocyte competence acquisition, follicular cells gene expression, and small extracellular vesicles (sEVs) miRNAs contents. RESULTS: P4 hormonal analysis revealed that ipsilateral to the CL follicular fluid (iFF) presented higher P4 concentration compared to contralateral follicular fluid (cFF). Furthermore, oocyte maturation and miRNA biogenesis pathways transcripts (ADAMTS-1 and AGO2, respectively) were increased in cumulus and granulosa cells of iFF, respectively. Nevertheless, a RT-PCR screening of 382 miRNAs showed that three miRNAs were upregulated and two exclusively expressed in sEVs from iFF and are predicted to regulate cell communication pathways. Similarly, seven miRNAs were higher and two exclusively expressed from cFF sEVs and are predicted to modulate proliferation signaling pathways. CONCLUSION: In conclusion, intrafollicular P4 concentration is influenced by the presence of the CL and modulates biological processes related to follicular cell development and oocyte competence, which may influence the oocyte quality. Altogether, these results are crucial to improve our knowledge about the follicular microenvironment involved in oocyte competence acquisition.