Role In Follicular Atresia Research Articles

Comparative miRNA expression profile analysis of porcine ovarian follicles: new insights into the initiation mechanism of follicular atresia.

Follicular atresia occurs in every stage of ovarian development, which is relevant to female fertility. In the past decade, increasing studies have confirmed that miRNAs, a class of short non-coding RNAs, play an important role in follicular atresia by post-transcription regulation of their target genes. However, the function of miRNAs on follicular atresia initiation is unknown. In the present study, high-throughput small RNA sequencing was performed to analyze differential miRNA expression profiles between healthy (HF) follicles and early atretic (EAF) follicles. A total of 237 conserved miRNA were detected, and the miR-143 is the highest expressed in follicles. Meanwhile, we also found wide sequence variations (isomiRs) in porcine ovarian miRNA, including in 5'un-translation region, core seed sequences and 3'untranslation region. Furthermore, we identified 22 differentially expressed miRNAs in EAF groups compared to HF group, of which 3 miRNAs were upregulated, as well as 19 miRNAs were downregulated, and then the RT-PCR was performed to validate these profiles. The target genes of these differentially expressed miRNAs were predicted by using miRwalk, miRDB, and Targetscan database, respectively. Moreover, the gene ontology and KEGG pathway enrichment established that the regulating functions and signaling pathways of these miRNAs contribute to follicular atresia initiation and cell fate. In conclusion, this study provides new insights into the changes of miRNAs in early atretic follicles to demonstrate their molecular regulation in ovarian follicular atretic initiation.

Impairment of steroidogenesis and follicle development after bisphenol A exposure during pregnancy and lactation in the ovaries of Mongolian gerbils aged females

The ovaries regulate fertility and hormonal control in females, and aging is a crucial factor in this process, when ovarian function is drastically impacted. Exogenous endocrine disruptors may accelerate this process, acting as the main agents in decreased female fertility and hormonal imbalance, since they impact different features related to reproduction. In the present study, we demonstrate the implications of exposure of adult mothers to the endocrine disruptor bisphenol A (BPA) during pregnancy and lactation on their ovarian function during the transition to later in life (aging). The follicle population of BPA exposed ovaries showed impairment in the development of follicles to the mature stages, with growing follicles being halted in the early stages. Atretic and early-atretic follicles were also enhanced. Expression of estrogen and androgen receptors in the follicle population demonstrated impairment in signaling function: ERβ was highly expressed in follicles from BPA exposed females, which also showed a higher incidence of early atresia of developed follicles. ERβ1 wild-type isoform was also enhanced in BPA-exposed ovaries, compared to its variant isoforms. In addition, steroidogenesis was targeted by BPA exposure: aromatase and 17-β-HSD were reduced, whereas 5-α reductase was enhanced. This modulation was reflected in serum levels of estradiol and testosterone, which decreased in BPA-exposed females. Imbalances in steroidogenesis impair the development of follicles and play an important role in follicular atresia. Our study demonstrated that BPA exposure in two windows of susceptibility – gestation and lactation – had implications during aging, enhancing perimenopausal and infertile features.

METTL3-mediated m6A methylation regulates granulosa cells autophagy during follicular atresia in pig ovaries

Follicular atresia is a normal physiological event in mammals, yet its mechanism remains to be studied. Granulosa cell (GC) autophagy is closely associated with follicular atresia. The N6-methyladenosine (m6A) modification is the most common post-transcriptional modification in eukaryotes, but its role in follicular atresia is still unknown. In this study, the possible relationship amongst follicular atresia, GC autophagy and m6A modification was studied. Our results showed that the level of autophagy in GCs increased with the degree of follicle atresia, whereas the overall m6A level decreased. Rapamycin treatment induced atresia in vitro cultured follicles, whereas 3-Methyladenine inhibited follicular atresia. Progressed atretic follicle (PAF) GCs had significantly lower METTL3 levels and significantly higher FTO levels than healthy follicle (HF) GCs. Differential gene expression analysis of GCs in PAF and HF by RNA sequencing was showed that the autophagy-related genes ULK1, ULK2, ATG2A, and ATG2B were significantly elevated in the PAF. In cultured GCs, overexpression of METTL3 significantly decreased the mRNA level of ULK1, as well as the autophagy level, whereas knockdown of METTL3 by RNAi significantly increased the mRNA level of ULK1, as well as the autophagy level. Our results indicate that m6A modification can regulate autophagy in GCs and play a role in the process of porcine follicular atresia.

MIR143 Inhibits Steroidogenesis and Induces Apoptosis Repressed by H3K27me3 in Granulosa Cells.

The granulosa cell growth factor and apoptotic factor are two factors to determine follicular apoptosis. Whether ssc-miR-143-3p (MIR143) plays as an apoptosis factor in porcine granulosa cells (pGCs) remain unclear. This study tries to investigate what function of MIR143 is and how MIR143 gets these functions in pGCs from 3 to 5 mm medium-sized follicles. Firstly, 5′ RACE was used to identify the structure of MIR143, and in situ hybridization, qPCR, and DNA pull-down were employed to exhibit the spatio-temporal expression and transcriptional regulation of MIR143. Furthermore, ELISA, Western blotting, and flow cytometry were adopted to explore the functions of MIR143 in pGCs. It was found that MIR143 was an exonic miRNA located in host gene LOC100514340 with an increasing expression during follicular growth. Moreover, MIR143 suppressed steroidogenesis related genes of HSD17β4, ER1, and PTGS2, negatively regulating estrogen, androgen, progesterone, and prostaglandin. MIR143 induced the apoptosis via activation of BAX-dependent Caspase 3 signaling. Furthermore, H3K27me3 influenced the recruitment of transcription factors and binding proteins to repress MIR143 transcription. At last, H3K27me3 agonist with MIR143 inhibition activated steroidogenesis but repressed apoptosis. These findings suggest that H3K27me3-mediated MIR143 inhibition play a critical role in follicular atresia by regulating cell apoptosis and steroidogenesis, which will provide useful information for further investigations of H3K27me3-miediated MIR143 epigenetic regulation in follicular growth in mammals.

Upregulation of miR-146b promotes porcine ovarian granulosa cell apoptosis by attenuating CYP19A1

MicroRNAs (miRNAs) are 21- to 24-nucleotide long small noncoding RNAs, which play an important role in follicular atresia and granulosa cell (GC) apoptosis in the mammalian ovary. Here, we report that miR-146b, a conserved and ovary-enriched miRNA, modulates estradiol (E2) secretion, GC apoptosis, and follicular atresia in pigs. Genome-wide analysis and quantitative real-time PCR revealed that miR-146b was significantly upregulated during follicular atresia, and fluorescence-activated cell sorting showed that miR-146b functioned as a proapoptotic factor to induce GC apoptosis. MicroRNA-mRNA network analysis and luciferase reporter assays showed that CYP19A1, the pivotal enzyme for E2 synthesis signaling, was directly targeted by miR-146b. Furthermore, miR-146b interacted with the 3′untranslated region of CYP19A1 to prevent translation, thereby regulating CYP19A1-mediated E2 secretion and GC apoptosis. However, miR-146b was not regulated by the transcription factor SMAD4 or oxidative stress, both of which are critical regulators of CYP19A1. We, thus, conclude that miR-146b is a novel epigenetic factor regulating GC functions, follicular development, and female reproduction.

Microanalysis of Fish Ovarian Follicular Atresia: A Possible Synergic Action of Somatic and Immune Cells.

The present study describes in detail the morphological characteristics of the process of ovarian follicular atresia in Redbelly tilapia (Coptodon zillii) during the nonbreeding season using light and electron microscopy and immunohistochemistry. The follicular regression process was initiated with shrinkage and disintegration of the nuclear membrane of oocytes resulting in dispersing of chromatin within the ooplasm, followed by marked hyperplasia and hypertrophy of follicular and granulosa cells, which exhibited a strong phagocytic activity to engulf the liquefied yolk particles. Rodlet cells and granulocytes were recorded on the follicular wall and invaded the regressed follicles. Rodlet cells expressed a strong immunoreactivity to matrix metalloperoxidase (MMP-9) and α-smooth muscle actin, while neutrophils expressed a strong reactivity to Myeloperoxidase-3 (MPO). In the advanced stage of follicular atresia, the yolk was almost phagocytized and resorbed and the regressed follicle lost its integrity and appeared to be formed of a cellular mass of phagocytic cells. Transmission electron microscopy revealed the presence of neutrophils, eosinophils, and dendritic cells within the atretic follicle in between these phagocytic cells. Moreover, numerous lysosomes, granules, and phagosomes were observed within the cytoplasm of both phagocytic cells and granulocytes. Telocytes were also demonstrated within the highly thickened richly vascularized theca layer during the late stages of follicular atresia. Immunohistochemical staining for caspase-3 established the participation of apoptosis in the advanced stages of follicular regression. Immune cells, rodlet cells, and telocytes in combination with follicular cells play an essential role in follicular atresia. In conclusion, the present study provides a new evidence on the role of both somatic and immune cells in the phenomenon of ovarian follicular atresia in Redbelly tilapia (Coptodon zillii) during the nonbreeding season.

Clinical correlation of apoptosis in human granulosa cells-A review.

The quality of oocytes depends on interactions with surrounding granulosa cells. Granulosa cells are essential in normal follicular maturation process since they produce steroidal hormones and growth factors, and they play a crucial role in follicular atresia. The success in reproductive biology and medicine depends on reliable assessment of oocyte and embryo viability which presently mainly bases on oocyte and embryo morphology. Recent investigations have tried to establish an evaluation system for oocyte quality and to predict outcome of in vitro fertilization (IVF) based on the incidence of granulosa cells and cumulus cells apoptosis. Apoptosis of granulosa cells seems to have a negative effect on conception and pregnancy rates in IFV programs. Thus, in this review we present a brief outline of clinical correlation of apoptosis in human granulosa cells and cumulus cells, and its influence upon oocyte quality and IFV outcome. Taken together, understanding the influence of granulosa cell apoptosis on oocyte quality and maturity as well as on embryo health may ultimately allow scientists and clinicians to harness better protocols of ovarian stimulation for infertility treatments.

Ovarian ageing: the role of mitochondria in oocytes and follicles.

There is a great inter-individual variability of ovarian ageing, and almost 20% of patients consulting for infertility show signs of premature ovarian ageing. This feature, taken together with delayed childbearing in modern society, leads to the emergence of age-related ovarian dysfunction concomitantly with the desire for pregnancy. Assisted reproductive technology is frequently inefficacious in cases of ovarian ageing, thus raising the economic, medical and societal costs of the procedures. Ovarian ageing is characterized by quantitative and qualitative alteration of the ovarian oocyte reserve. Mitochondria play a central role in follicular atresia and could be the main target of the ooplasmic factors determining oocyte quality adversely affected by ageing. Indeed, the oocyte is the richest cell of the body in mitochondria and depends largely on these organelles to acquire competence for fertilization and early embryonic development. Moreover, the oocyte ensures the uniparental transmission and stability of the mitochondrial genome across the generations. This review focuses on the role played by mitochondria in ovarian ageing and on the possible consequences over the generations. PubMed was used to search the MEDLINE database for peer-reviewed original articles and reviews concerning mitochondria and ovarian ageing, in animal and human species. Searches were performed using keywords belonging to three groups: 'mitochondria' or 'mitochondrial DNA'; 'ovarian reserve', 'oocyte', 'ovary' or 'cumulus cells'; and 'ageing' or 'ovarian ageing'. These keywords were combined with other search phrases relevant to the topic. References from these articles were used to obtain additional articles. There is a close relationship, in mammalian models and humans, between mitochondria and the decline of oocyte quality with ageing. Qualitatively, ageing-related mitochondrial (mt) DNA instability, which leads to the accumulation of mtDNA mutations in the oocyte, plays a key role in the deterioration of oocyte quality in terms of competence and of the risk of transmitting mitochondrial abnormalities to the offspring. In contrast, some mtDNA haplogroups are protective against the decline of ovarian reserve. Quantitatively, mitochondrial biogenesis is crucial during oogenesis for constituting a mitochondrial pool sufficiently large to allow normal early embryonic development and to avoid the untimely activation of mitochondrial biogenesis. Ovarian ageing also seriously affects the dynamic nature of mitochondrial biogenesis in the surrounding granulosa cells that may provide interesting alternative biomarkers of oocyte quality. A fuller understanding of the involvement of mitochondria in cases of infertility linked to ovarian ageing would contribute to a better management of the disorder in the future.

WITHDRAWN:Nitric oxide and ovarian folliculogenesis: a possible role in follicular atresia.

Ahead of Print article withdrawn by publisher.

MicroRNA-125a-5p induces mouse granulosa cell apoptosis by targeting signal transducer and activator of transcription 3.

Premature ovarian failure, a reproductive dysfunction characterized by follicle loss leads to premature menopause. Apoptosis of granulosa cells may be responsible for the associated follicle depletion. MicroRNAs are expressed abundantly in granulosa cells and play an important role in follicular atresia. Evidence suggests that signal transducer and activator of transcription 3 (STAT3) is involved in follicle growth and female fertility. We incubated cultured mouse granulosa cells (mGCs) with increasing doses of cisplatin (CP) for varying periods. Cell proliferation and apoptosis were measured by Cell Counting Kit-8 assay, flow cytometry, and protein expression of cleaved caspase-3. Western blot analysis was used to assess STAT3 and phospho-STAT3 after mGCs were transfected with a microRNA-125a-5p (miR-125a-5p) mimic and a miR-125a-5p inhibitor, respectively. Luciferase reporter assay was conducted to determine the relationship between miR-125a-5p and STAT3. CP reduced mGC viability, progesterone levels, and estradiol levels. miR-125a-5p was up-regulated in CP-treated mGCs, whereas STAT3 was down-regulated. Increased apoptosis and cleaved caspase-3 were observed in mGCs transfected with a miR-125a-5p mimic or STAT3 interference fragment. Protein expression of STAT3 and phospho-STAT3 was up-regulated or down-regulated when transfected with a miR-125a-5p inhibitor or miR-125a-5p mimic, respectively. Luciferase reporter assays indicated that miR-125a-5p targets the 3' untranslated region of STAT3. Overexpression of miR-125a-5p promotes mGC apoptosis by targeting STAT3. Our findings imply the important role of miR-125a-5p in the pathogenesis of premature ovarian failure.

Expression of Mitochondria-Associated Genes (PPARGC1A, NRF-1, BCL-2 and BAX) in Follicular Development and Atresia of Goat Ovaries.

Most follicles undergo atresia during the developmental process. Follicular atresia is predominantly regulated by apoptosis of granulosa cells, but the mechanism underlying apoptosis via the mitochondria-dependent apoptotic pathway is unclear. We aimed to investigate whether the mitochondria-associated genes peroxisome proliferator-activated receptor-gamma, coactivator1-alpha (PPARGC1A), nuclear respiratory factor-1 (NRF-1), B-cell CLL/lymphoma 2 (BCL-2) and BCL2-associated X protein (BAX) played a role in follicular atresia through this pathway. The four mitochondria-associated proteins (PGC-1α, which are encoded by the PPARGC1A gene, NRF-1, BCL-2 and BAX) mainly expressed in granulosa cells. The mRNA and protein levels of PPARGC1A/PGC-1α and NRF-1 in granulosa cells increased with the follicular development. These results showed that these genes may play a role in the regulation of the follicular development. In addition, compared with healthy follicles, the granulosa cell in atretic follicles had a reduced expression of NRF-1, increased BAX expression and increased ratio of BAX to BCL-2 expression. These results suggested that changes of the mitochondria-associated gene expression patterns in granulosa cells may lead to follicular atresia during goat follicle development.

Changes in the expression of Fox O1 and death ligand genes during follicular atresia in porcine ovary.

Follicular atresia, a key phenomenon in follicle development, eliminates most of the follicles in mammalian ovaries. To investigate the molecular mechanism of follicular atresia in porcine ovaries, we investigated the mRNA expression of three important cell death ligand-receptor systems and Fox O1 in follicles with a diameter of 3-5 mm. The phosphorylation and subcellular localization of Fox O1 during granulosa cell apoptosis was also determined. TRAIL and Fas L played an important role in follicular atresia at this stage. Fox O1 expression was upregulated during atresia, and was confined to the nucleus of granulosa cells; however, phosphorylated Fox O1 was localized to the cytoplasm. These results suggest Fox O1 involvement in the regulation of TRAIL and Fas L expression during follicular atresia in pigs.

English

Follicular atresia is the term used for the fate of follicles which undergo degenerative changes before rupturing during ovulation. Recent studies suggest that granulosa cell apoptosis play a major role in follicular atresia. The factors which lead the cell to apoptosis and which protect the cell death, still remain complicated and more studies are needed to elucidate the whole process. Here in this review, we aimed to simplify the factors and mechanisms taking place in granulosa cell apoptosis, to make the process more understandable.   Key words: Granulosa cell, apoptosis, follicular atresia.

Localization of interleukin-2 in goat ovary

––Distribution and relative frequencies of interleukin-2 in normal cycling goat ovaries were studied to explain local immunological defense mechanisms. IL-2 was observed in primordial, primary, secondary and antral follicles. Interleukins are known best for their involvement in the immune system and their role during inflammation. In the ovary, ovarian follicle is a site of inflammatory reaction. Thus ovarian cells could represent sources and targets of interleukins. Objective: To explore this, we have examined the localisation of interleukin-2 in goat ovarian follicles. Study design: Mature goat ovaries were collected from slaughter houses and brought to laboratory at 4 o C in normal saline. Mouse monoclonal antibody (mAb) was used at a 1:1000 dilution, Biotinylated goat anti polyvalent IgG was used as secondary antibody (TP-015-BN). A streptavidin alkaline phosphatase (lab vision, corporation, TP 015-AP) method was applied to detect IL-2+ cells. The specific color reaction was developed with napthol phosphate (fast red tablets). Results: IL-2 + cells were found in small numbers in developing follicles and newly formed corpora lutea, while atretic follicles and regressing corpora lutea contained these cells in abundance. Conclusion: The frequency of IL-2+ cells was higher in the atretic follicles of all stages. This suggests that interleukin 2 play an important role in follicular atresia rather than follicular growth in normal cycling ovaries. Keywords––goat, ovary, interleukin-2.

Do the Theca Layer and Fibroblast Growth Factors Have a Role in Follicular Atresia?1

Ovarian follicular development in the bovine estrous cycle is characterized by a wave-like pattern of successive periods of follicle growth and regression, with the majority of estrous cycles in cows comprising either two or three waves per cycle. Each wave involves recruitment of a cohort of antral follicles, followed by selection of a single dominant follicle that progresses to the pre-ovulatory stage, while the remaining follicles of the wave, the subordinate follicles, undergo atresia. The pituitary gonadotropic hormones, follicle-stimulating hormone (FSH) and luteinizing hormone, are the primary drivers of follicle recruitment and growth, with molecules produced locally within the follicle having secondary but essential roles in dominant follicle selection, while promoting atresia of subordinate follicles [1]. Bidirectional communication between the oocyte and adjacent cumulus granulosa cells results in expansion and mucification of the cumulus-oocyte complex necessary for ovum release at ovulation, mediated by oocyte-secreted peptides (growth differentiation factor 9, bone morphogenic protein [BMP] 15, fibroblast growth factor [FGF] 8) [2, 3]. In turn, molecules from cumulus cells (epidermal growth factor–like peptides, cAMP, and pyruvate) trigger meiotic and cytoplasmic maturation of the oocyte essential for fertilization and embryo development [4]. Similarly, paracrine signals and substrates passing between mural granulosa cells and theca cells of the follicle wall contribute to both granulosa and theca cell proliferation and differentiation essential for continued follicle growth and maturation leading to ovulation. Theca cells supply androgens as precursors for conversion to estrogens by aromatase located exclusively in granulosa cells, and growth factors (insulin-like growth factors [IGFs] and FGFs) that act synergistically with the estrogens to further enhance follicle growth. In turn, estrogens and other growth factors produced by granulosa cells exert regulatory actions back on the theca cells. These closely coordinated activities of the three follicle cell types—oocyte, granulosa cells, theca cells—comprise a regulatory network that results in an intrafollicular milieu necessary for follicle growth and ovulation of a developmentally competent ovum as culmination of the final follicular wave of the cycle [5,

Cortical Neural Precursors Inhibit Their Own Differentiation via N-Cadherin Maintenance of β-Catenin Signaling

Little is known about the architecture of cellular microenvironments that support stem and precursor cells during tissue development. Although adult stem cell niches are organized by specialized supporting cells, in the developing cerebral cortex, neural stem/precursor cells reside in a neurogenic niche lacking distinct supporting cells. Here, we find that neural precursors themselves comprise the niche and regulate their own development. Precursor-precursor contact regulates beta-catenin signaling and cell fate. In vivo knockdown of N-cadherin reduces beta-catenin signaling, migration from the niche, and neuronal differentiation in vivo. N-cadherin engagement activates beta-catenin signaling via Akt, suggesting a mechanism through which cells in tissues can regulate their development. These results suggest that neural precursor cell interactions can generate a self-supportive niche to regulate their own number.

Follicle Atresia Is Regulated by Fibroblast Growth Factor-18 in Cattle.

Understanding the paracrine events that regulate fertility in the cow is necessary not only because of the agricultural importance of this species, but also its potential use as a model for humans. Fibroblast growth factors (FGF) and their receptors are involved in the paracrine modulation of ovarian function. For example, FGF2, 7 and 10 inhibit estradiol secretion by granulosa cells in vitro. Another member of this family, FGF8, is an oocyte-derived protein that modulates glycolysis in cumulus cells. FGFs are grouped into subfamilies of closely-related proteins, and FGF18 is in the same subfamily as FGF8 and has also been detected in mouse oocytes. The existance of a subfamily of FGFs that are specifically derived from the oocyte are potentially interesting paracrine regulators of fertility. The objectives of the present study were to localize the expression of FGF18 in the bovine follicle, to gain insight into the control of FGF18 expression and to identify potential actions of FGF18 in the follicle. Bovine ovaries were obtained from an abattoir for localizing expression patterns by real-time PCR and immunohistochemistry. FGF18 mRNA was not detected in pooled oocytes by PCR, but was present in isolated theca and granulosa cells (GC), and protein was detected in the follicle wall by immunohistochemistry. Expression during follicle growth was measured by collecting the two largest follicles per cow at defined stages of the first follicle wave. The follicles were classified as dominant and subordinate according to estradiol levels in follicular fluid and aromatase mRNA abundance in GC. FGF18 mRNA abundance in both GC and theca cells was higher in subordinate follicles compared with dominant follicles (P<0.05). To determine regulation and function of FGF18, granulosa cells were isolated from 5-8 mm follicles and employed in an estrogenic, serum-free culture system. In vitro, FGF18 expression was down-regulated by IGF1, insulin, FGF2, FGF10 (P<0.05) in GC. In further experiments, the addition of graded doses of FGF18 (0, 1, 10, 100 ng/ml) to GC in culture inhibited estradiol secretion and abundance of mRNA encoding the estrogenic enzymes aromatase and 17β-hydroxysteroid dehydrogenase (17βHSD), and the FSH receptor. Progesterone secretion was also inhibited, which was associated with decreased abundance of mRNA encoding CYP11, StAR and 3βHSD. Assessment of apoptosis by flow cytometry showed that FGF18 significantly increased the proportion of apoptotic cells, and this was accompanied by decreased abundance of mRNA encoding antiapoptotic and proliferation genes (MIF, RFC4 and GADD45B). In summary, FGF18 is expressed in GC and theca cells but--in contrast to the subfamily prototype, FGF8--not in oocytes in cattle. As FGF18 expression is increased in atretic follicles, and as FGF18 protein inhibits estradiol secretion and induces apoptosis, we conclude that FGF18 plays a role in follicular atresia. Supported by NSERC Canada, CNPq and FAPESP, Brazil. (platform)

Role of Cell Death Ligand and Receptor System on Regulation of Follicular Atresia in Pig Ovaries

Several hundred thousand primordial follicles are present in the mammalian ovary, however, only a limited number develop to the pre-ovulatory stage, and then finally ovulate. The others, more than 99%, will be eliminated through a degenerative process called 'atresia'. The endocrinological regulatory mechanisms involved in follicular development and atresia have been characterized to a large extent, but the precise temporal and molecular mechanisms involved in the regulation of these events have remained unknown. From many recent studies, it is suggested that the apoptosis in ovarian granulosa cells plays a crucial role in follicular atresia. Notably, death ligand-receptor interaction and subsequent intracellular signalling have been demonstrated to be the key mechanisms regulating granulosa cell apoptosis. In this review, we provide an overview of granulosa cell apoptosis regulated by death ligand-receptor signalling. The roles of death ligands and receptors [Fas ligand (FasL)-Fas, tumour necrosis factor (TNF)alpha-TNF receptor (TNFR), and TNFalpha-related apoptosis-inducing ligand (TRAIL)-TRAIL receptor (TRAILR)] and intracellular death-signal mediators [Fas-associated death domain protein (FADD), TNF receptor 1-associated death domain protein (TRADD), caspases, apoptotic protease-activating factor 1 (Apaf1), TNFR-associated factor 2 (TRAF2), and cellular FLICE-like inhibitory protein (cFLIP), etc.] in granulosa cells will be discussed.

Expression and Localization of the Serine Proteases High-Temperature Requirement Factor A1, Serine Protease 23, and Serine Protease 35 in the Mouse Ovary

Proteolytic degradation of extracellular matrix components has been suggested to play an essential role in the occurrence of ovulation. Recent studies in our laboratory have indicated that the plasminogen activator and matrix metalloproteinase systems, which were previously believed to be crucial for ovulation, are not required in this process. In this study we have used a microarray approach to identify new proteases that are involved in ovulation. We found three serine proteases that were relatively highly expressed during ovulation: high-temperature requirement factor A1 (HtrA1), which was not regulated much during ovulation; serine protease 23 (PRSS23), which was down-regulated by gonadotropins; and serine protease 35 (PRSS35), which was up-regulated by gonadotropins. We have further investigated the expression patterns of these proteases during gonadotropin-induced ovulation in immature mice and in the corpus luteum (CL) of pseudopregnant mice. We found that HtrA1 was highly expressed in granulosa cells throughout follicular development and ovulation, as well as in the forming and regressing CL. PRSS23 was highly expressed in atretic follicles, and it was expressed in the ovarian stroma and theca tissues just before ovulation. PRSS35 was expressed in the theca layers of developing follicles. It was also highly induced in granulosa cells of preovulatory follicles. PRSS35 was also expressed in the forming and regressing CL. These data suggest that HtrA1 and PRSS35 may be involved in ovulation and CL formation and regression, and that PRSS23 may play a role in follicular atresia.

Regulation of granulosa cell apoptosis by death ligand–receptor signaling

ABSTRACTSeveral hundred thousand primordial follicles are present in the mammalian ovary, however, only 1% develop to the preovulatory stage and finally ovulate. The remainder will be eliminated via a degenerative process called ‘atresia’. The endocrinological regulatory mechanisms involved in follicular development and atresia have largely been characterized but the precise temporal and molecular mechanisms involved in the regulation of these events remain unknown. Many recent studies suggest that apoptosis in ovarian granulosa cells plays a crucial role in follicular atresia. Notably, death ligand‐receptor interaction and subsequent intracellular signaling have been demonstrated to be the key mechanisms regulating granulosa cell apoptosis. In this review we provide an overview of granulosa cell apoptosis regulated by death ligand‐receptor signaling. The roles of death ligands and receptors [Fas ligand (FasL)]‐Fas, tumor necrosis factor α (TNFα)‐TNF receptor and TNFα‐related apoptosis‐inducing ligand (TRAIL)‐TRAIL receptor (TRAILR)] and intracellular death‐signal mediating molecules (Fas‐associated death domain protein), TNF receptor 1‐associated death domain protein, caspases, apoptotic protease‐activating factor 1, TNFR‐associated factor 2 and cellular FLICE‐like inhibitory protein in granulosa cells are discussed.