Histological analysis of (antral) follicle density in ovarian cortex tissue attached to stripped endometriomas.

PURPOSE: When resecting endometriomas with the stripping technique, in the majority of cases, a thin line of adjacent ovarian cortex is attached to the endometrioma. In this study, we performed histological analysis to determine (antral) follicle density in the ovarian cortex tissue attached to stripped endometriomas and assessed patient- and surgical characteristics that could affect this. METHODS: Histological slides of previously removed endometriomas were assessed. Follicles in the attached ovarian tissue were classified according to maturation, and follicular density was determined. Immunofluorescent staining of antral follicles in a subset of endometriomas was also performed. RESULTS: In 90 out of 96 included endometriomas (93.7%), ovarian tissue attached to the cyst wall was observed. One thousand nine hundred forty-four follicles at different maturation stages were identified (3 follicles/mm3). Follicle density was negatively associated with age (p < 0.001). Antral follicles (< 7-mm diameter) were present in the ovarian tissue attached to 35 endometriomas (36.5%) derived from younger patients compared to endometriomas where none were detected (30 versus 35 years, p = 0.003). Antral follicle density was 1 follicle/mm3. Based on immunofluorescence, healthy antral follicles were identified in two out of four examined endometriomas. CONCLUSIONS: Ovarian tissue attached to stripped endometriomas holds potential as a non-invasive source for antral follicles. In theory, application of IVM could be an interesting alternative FP option in young patients with endometriomas who undergo cystectomy in order to transform the surgical collateral damage to a potential oocyte source. Our results encourage future research with fresh tissue to further assess the quality and potential of these follicles. TRIAL REGISTRATION: Clinical Trials.gov Identifier: B21.055 (METC LDD), date of registration 12-08-2021, retrospectively registered.

Basal characteristics of patients who responded to Ovarian Fragmentation for Follicular Activation (OFFA) or In Vitro Activation (IVA): a systematic review and meta-analysis.

A systematic review and meta-analysis were performed to identify if there is a subset of patients with POI who are more likely to show follicular growth after ovarian fragmentation for follicular activation (OFFA) or in vitro activation (IVA). Five studies met inclusion criteria for meta-analysis with a total of 164 patients. Forty-three patients showed follicle development (26.21%). Of those, the pregnancy rate was 35.58% (11/43) and the live birth rate was 20.93% (9/43). Our meta-analysis showed that age was not associated with follicle growth. However, lower baseline FSH, lower duration of amenorrhea/diagnosis, and presence of follicles remaining in biopsy were statistically significant for follicle development. Patients with basal characteristics mentioned before may have more chances to show follicle growth after OFFA or IVA. Taking into account that approximately 20% of patients with follicle growth had live birth, these results are very promising. Given the overall certainty of evidence, future studies are needed to confirm said results.

Distocia em tigre d'água-de-orelha-vermelha, Trachemys scripta elegans, wied (1839): relato de caso / Dystocia in red-eared slider, Trachemys scripta elegans, wied (1839): case report / Distocia en tortuga de orejas rojas, Trachemys scripta elegans, wied (1839): reporte de caso

Atualmente muitos répteis se tornaram animais de companhia e são mantidos como pet's exóticos. A espécie Trachemys scripta elegans, Wied (1839) é um animal exótico da América do Norte, sua identificação é realizada pelas marcas avermelhadas encontradas lateralmente a sua cabeça. Na rotina clínica as principais enfermidades que acometem os quelônios são as de origem reprodutiva, como a estase folicular e distocia. O objetivo deste trabalho foi relatar um caso recorrente de distocia em um tigre d'água fêmea, para isso, a anamnese, o histórico da paciente, e seus sinais clínicos, em conjunto com os exames complementares de imagem foram essenciais para se obter diagnóstico definitivo. O tratamento foi realizado com a indução medicamentosa utilizando borogluconato de cálcio, seguida da aplicação de ocitocina, esta trouxe resultados positivos para a eliminação dos ovos. Porém devido ao histórico do paciente, optou-se pela intervenção cirúrgica de ovariossalpingectomia, sendo está a maneira permanente de resolução da patologia. O protocolo terapêutico escolhido proporcionou um resultado satisfatório e bem estar ao animal.(AU)

Currently, many reptiles have become companion animals and are kept as exotic pets. The species Trachemys scripta elegans, Wied (1839) is an exotic animal from North America, and its identification is based on the reddish markings found laterally on its head. In routine clinical practice, the main diseases that affect chelonians are those of reproductive origin, such as follicular stasis and dystocia. The aim of this study was to report a recurrent case of dystocia in a female red-eared slider turtle. For this purpose, the patient's anamnesis, history, and clinical signs, along with complementary imaging exams, were essential to obtain a definitive diagnosis. The treatment involved medical induction using calcium borogluconate, followed by the administration of oxytocin, which yielded positive results in egg elimination. However, due to the patient's history, surgical intervention in the form of ovariosalpingectomy was chosen as the permanent solution to the pathology. The chosen therapeutic protocol provided a satisfactory outcome and improved the animal's well-being.(AU)

Actualmente muchos reptiles se han convertido en animales de compañía y se mantienen como mascotas exóticas. La especie Trachemys scripta elegans, Wied (1839) es un animal exótico de América del Norte, su identificación se realiza por las marcas rojizas que se encuentran lateralmente a su cabeza. En la rutina clínica, las principales enfermedades que afectan a los quelonios son las de origen reproductivo, como la estasis folicular y la distocia. El objetivo de este trabajo fue reportar un caso recurrente de distocia en una hembra de tigre de agua, para ello la anamnesis, la historia de la paciente y sus signos clínicos, junto con los exámenes imagenológicos complementarios fueron fundamentales para obtener un diagnóstico definitivo. El tratamiento se realizó con inducción farmacológica con borogluconato de calcio, seguido de la aplicación de oxitocina, que arrojó resultados positivos con la eliminación de huevos. Sin embargo, debido a los antecedentes de la paciente, se optó por la intervención quirúrgica de ovarialpingectomía, que es la forma definitiva de resolución de la patología. El protocolo terapéutico elegido proporcionó un resultado satisfactorio y bienestar al animal.(AU)

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

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

Aged mice ovaries harbor stem cells and germ cell nests but fail to form follicles.

BACKGROUND: We recently published evidence to suggest that two populations of stem cells including very small embryonic-like stem cells (VSELs) and ovarian stem cells (OSCs) in ovary surface epithelium (OSE) undergo proliferation/differentiation, germ cell nests (GCN) formation, meiosis and eventually differentiate into oocytes that assemble as primordial follicles on regular basis during estrus cycle. Despite presence of stem cells, follicles get exhausted with advancing age in mice and result in senescence equivalent to menopause in women. Stem cells in aged ovaries can differentiate into oocytes upon transplantation into young ovaries, however, it is still not well understood why follicles get depleted with advancing age despite the presence of stem cells. The aim of the present study was to study stem cells and GCN in aged ovaries. METHODS: OSE cells from aged mice (> 18 months equivalent to > 55 years old women) were enzymatically separated and used to study stem cells. Viable (7-AAD negative) VSELs in the size range of 2-6 µm with a surface phenotype of Lin-CD45-Sca-1+ were enumerated by flow cytometry. Immuno-fluorescence and RT-PCR analysis were done to study stem/progenitor cells (OCT-4, MVH, SCP3) and transcripts specific for VSELs (Oct-4A, Sox-2, Nanog), primordial germ cells (Stella), germ cells (Oct-4, Mvh), early meiosis (Mlh1, Scp1) and ring canals (Tex14). RESULTS: Putative VSELs and OSCs were detected as darkly stained, spherical cells with high nucleo-cytoplasmic ratio along with germ cells nests (GCN) in Hematoxylin & Eosin stained OSE cells smears. Germ cells in GCN with distinct cytoplasmic continuity expressed OCT-4, MVH and SCP3. Transcripts specific for stem cells, early meiosis and ring canals were detected by RT-PCR studies. CONCLUSION: Rather than resulting as a consequence of accelerated loss of primordial follicle and their subsequent depletion, ovarian senescence/menopause occurs as a result of stem cells dysfunction. VSELs and OSCs exist along with increased numbers of GCNs arrested in pre-meiotic or early meiotic stage in aged ovaries and primordial follicle assembly is blocked possibly due to age-related changes in their microenvironment.

miR-450-5p and miR-202-5p Synergistically Regulate Follicle Development in Black Goat.

Follicle maturation is a complex biological process governed by numerous factors, and researchers have observed follicle development by studying the proliferation and apoptosis of follicular granulosa cells (GCs). However, the regulatory mechanisms of GCs proliferation and death during follicle development are largely unknown. To investigate the regulatory mechanisms of lncRNAs, mRNAs, and microRNAs, RNA sequencing (RNA-seq) and small RNA-seq were performed on large (>10 mm) and small follicles (<3 mm) of Leizhou black goat during estrus. We discovered two microRNAs, miR-450-5p and miR-202-5p, which can target GCs in goats and may be involved in follicle maturation, and the effects of miR-450-5p and miR-202-5p on ovarian granulosa cell lines were investigated (KGN). Using cell counting kit-8 (CCK-8) assays, 5-Ethynyl-2'-deoxyuridine (EdU) assay and flow cytometry, miR-202-5p overexpression could suppress the proliferation and induce apoptosis of GCs, whereas miR-450-5p overexpression induced the opposite effects. The dual-luciferase reporter assay confirmed that miR-450-5p could directly target the BMF gene (a BCL2 modifying factor), and miR-202-5p targeted the BCL2 gene. A considerable rise in phosphorylated Akt (p-AKT) protein was observed following the downregulation of BMF by miR-450-5p mimics. After BMF gene RNAi therapy, a notable elevation in p-AKT was detected. Mimics of miR-202-5p inhibited BCL2 protein expression, significantly decreasing p-AMPK protein expression. These results imply that during the follicular development in black goats, the miR-450-5p-BMF axis favored GC proliferation on a wide scale, while the miR-202-5p-BCL2 axis triggered GC apoptosis.

Evaluation of the Effects of Human Bone Marrow Mesenchymal Stem Cells Conditioned Medium on Growth and Maturation of Mouse Ovarian Follicle after Vitrification.

The aim of this research study is to evaluate the effect of human bone marrow mesenchymal stem cells conditioned medium (hBMSCs-CM) on growth and maturation of mouse ovarian follicle, and embryonic development after vitrification. The hBMSCs were cultured, and the derived CM was collected, concentrated, and stored. 14-day-old mice ovaries were collected and randomly divided into vitrified and non-vitrified groups. Then their isolated preantral follicles were cultured for 12 days in α-MEM supplemented with different concentrations of CM (2.5, 5, and 7.5%). Finally, the growth and diameter of follicles, maturation of oocytes, hormone level, and embryo developmental rate were assessed. The results showed the antrum formation, oocyte maturation, and hormone secretion were significantly higher in the presence of 7.5% CM (p < 0.001). In the vitrified group, the developmental rate of follicles was lower than the non-vitrified group, and the subgroup containing 7.5% CM showed better results than the 5%, and 2.5% CM subgroups. However, no changes in fertilization and embryonic development rates were observed. Supplementing follicle culture media with 7.5% CM could enhance follicle growth and oocyte maturation of follicles after vitrification.

Overexpression of Lin28a induces a primary ovarian insufficiency phenotype via facilitation of primordial follicle activation in mice.

Lin28a is an RNA binding protein and increasing evidence has indicated its role in regulating female fertility. Lin28a has been reported to be involved in ovarian follicle activation. However, its role and mechanisms in regulating primordial follicle activation have not yet been explored. To test whether overexpression of Lin28a activates ovarian primordial follicles, studies were conducted in wild type (WT) and Lin28a Tg mice. Female Lin28a Tg mice at 4-month old exhibited significantly smaller litter size and fewer ovulated oocytes when compared with the WT mice. By 6-month of age, these parameters in Lin28a Tg mice were less than 20% of the WT mice. At postnatal day (PD) 14, the number of primordial follicles was significantly decreased but the number of primary follicles was significantly increased in the transgenic mice. The number of primordial follicles, secondary and antral follicles in these mice were drastically reduced at PD21. In the ovary of Lin28a Tg mice, there were activation of Wnt/ß-catenin signaling and its downstream mTOR pathway. Interestingly, overexpression of Lin28a, which can also act as transcriptional activator, activated Wnt signaling through enhancing the transcription of Wnt co-receptor LRP5. In conclusion, overexpression of Lin28a induced a primary ovarian insufficiency phenotype in long term via facilitating Wnt/ß-catenin signaling leading to activation of primordial follicles.

Transcriptome analysis reveals that long noncoding RNAs contribute to developmental differences between medium-sized ovarian follicles of Meishan and Duroc sows.

Ovulation rate is an extremely important factor affecting litter size in sows. It differs greatly among pig breeds with different genetic backgrounds. Long non-coding RNAs (lncRNAs) can regulate follicle development, granulosa cell growth, and hormone secretion, which in turn can affect sow litter size. In this study, we identified 3554 lncRNAs and 25,491 mRNAs in M2 follicles of Meishan and Duroc sows. The lncRNA sequence and open reading frame lengths were shorter than mRNAs, and lncRNAs had fewer exons, were less abundant, and more conserved than protein-coding RNAs. Furthermore, 201 lncRNAs were differentially expressed (DE) between breeds, and quantitative trait loci analysis of DE lncRNAs were performed. A total of 127 DE lncRNAs were identified in 119 reproduction trait-related loci. In addition, the potential target genes of lncRNAs in cis or trans configurations were predicted. Gene ontology and Kyoto Encyclopedia of Genes and Genomes pathway analysis revealed that some potential target genes were involved in follicular development and hormone secretion-related biological processes or pathways, such as progesterone biosynthetic process, estrogen metabolic process, ovarian steroidogenesis, and PI3K-Akt signaling pathway. Furthermore, we also screened 19 differentially expressed lncRNAs in the PI3K-Akt signaling pathway as candidates. This study provides new insights into the roles of lncRNAs in follicular growth and development in pigs.

Role of Stem Cells in the Ovarian Tissue Cryopreservation and Transplantation for Fertility Preservation.

Although the cancer survival rate has increased, cancer treatments, including chemotherapy and radiotherapy, can cause ovarian failure and infertility in women of reproductive age. Preserving fertility throughout cancer treatment is critical for maintaining quality of life. Fertility experts should propose individualized fertility preservation methods based on the patient's marital status, pubertal status, partner status, and the urgency of treatment. Widely practiced fertility preservation methods, including ovarian transposition and embryo and oocyte cryopreservation, are inappropriate for prepubertal girls or those needing urgent initiation of cancer treatment. Ovarian tissue cryopreservation and transplantation, an emerging new technology, may be a solution for these cancer patients. The use of stem cells in ovarian tissue cryopreservation and transplantation increases oxygenation, angiogenesis, and follicle survival rates. This review discusses the recent advances in ovarian tissue cryopreservation and transplantation with special focus on the use of stem cells to improve fertilization techniques.

Effect of Exogenous Melatonin on the Development of Mice Ovarian Follicles and Follicular Angiogenesis.

In mammalian, the periodic growth and development of ovarian follicles constitutes the physiological basis of female estrus and ovulation. Concomitantly, follicular angiogenesis exerts a pivotal role in the growth of ovarian follicles. Melatonin (N-acetyl-5-methoxytryptamine, Mel), exists in follicle fluid, was suggested to affect the development of follicles and angiogenesis. This research was conducted to investigate the effects and mechanisms of Mel on the development of ovarian follicles and its angiogenesis. In total, 40 ICR mice at age of 3 weeks were allocated into four groups at liberty: control, Mel, FSH and FSH + Mel for a 12-day trial. Ovaries were collected at 8:00 a.m. on Day 13 for detecting the development of ovarian follicles and angiogenesis. Results indicated that Mel promoted the development of ovarian follicles of 50-250 µm (secondary follicles) and periphery angiogenesis, while FSH remarkably increased the number of antral follicles and periphery angiogenesis. Mechanically, Mel and FSH may regulate the expression of VEGF and antioxidant enzymes in different follicular stages. In conclusion, Mel primarily acted on the secondary follicles, while FSH mainly promoted the development of antral follicles. They both conduced to related periphery angiogenesis by increasing the expression of VEGF. These findings may provide new targets for the regulating of follicular development.

METTL3-mediated mRNA N6-methyladenosine is required for oocyte and follicle development in mice.

Proper follicle development is very important for the production of mature oocytes, which is essential for the maintenance of female fertility. This complex biological process requires precise gene regulation. The most abundant modification of mRNA, N6-methyladenosine (m6A), is involved in many RNA metabolism processes, including RNA splicing, translation, stability, and degradation. Here, we report that m6A plays essential roles during oocyte and follicle development. Oocyte-specific inactivation of the key m6A methyltransferase Mettl3 with Gdf9-Cre caused DNA damage accumulation in oocytes, defective follicle development, and abnormal ovulation. Mechanistically, combined RNA-seq and m6A methylated RNA immunoprecipitation sequencing (MeRIP-seq) data from oocytes revealed, that we found METTL3 targets Itsn2 for m6A modification and then enhances its stability to influence the oocytes meiosis. Taken together, our findings highlight the crucial roles of mRNA m6A modification in follicle development and coordination of RNA stabilization during oocyte growth.

A blueprint of the topology and mechanics of the human ovary for next-generation bioengineering and diagnosis.

Although the first dissection of the human ovary dates back to the 17th century, the biophysical characteristics of the ovarian cell microenvironment are still poorly understood. However, this information is vital to deciphering cellular processes such as proliferation, morphology and differentiation, as well as pathologies like tumor progression, as demonstrated in other biological tissues. Here, we provide the first readout of human ovarian fiber morphology, interstitial and perifollicular fiber orientation, pore geometry, topography and surface roughness, and elastic and viscoelastic properties. By determining differences between healthy prepubertal, reproductive-age, and menopausal ovarian tissue, we unravel and elucidate a unique biophysical phenotype of reproductive-age tissue, bridging biophysics and female fertility. While these data enable to design of more biomimetic scaffolds for the tissue-engineered ovary, our analysis pipeline is applicable for the characterization of other organs in physiological or pathological states to reveal their biophysical markers or design their bioinspired analogs.

Body status and blood metabolites profiles during resumption of postpartum ovarian activity in yak (Poephagus grunniens).

We examined the changes in body weight (BW), back-fat thickness (BFT) and blood metabolites in relation to postpartum (PP) ovarian activity status in twenty female yaks raised under semi-intensive system. BFT and ovarian activities, like follicle development, ovulation (OV) and corpus luteum (CL) development, were monitored from 4 to 15 weeks (wk) PP using ultrasonography. Resumption of ovarian activity was confirmed with ovulation of dominant follicle (DF) and subsequent CL development, and >1 ng/ml progesterone concentration in blood plasma sample after 1week of ovulation. Yaks were further classified as cyclic (with CL), acyclic (without CL), and cystic (with >25 mm follicular cyst; FC). Within 20 weeks PP, 60% yaks resumed cyclic ovarian activity, while 25% failed to initiate cycling activity, and 15% developed follicular cysts. In all categories of yak, BW gradually decreased (p < .05) till nadir; however, nadir reached earlier (p < .05) in acyclic yaks. BFT differed (p < .05) among the yak groups, but it tended to be higher in cyclic yaks as compared to acyclic and cystic. No difference (p > .05) in non-esterified fatty acids (NEFA) values was found among the different categories of yaks, whereas, beta-hydroxy butyrate (BHB) levels were higher in cystic animals as compared to acyclic and cyclic. Blood glucose levels decreased in all yaks during initial 2 weeks after calving. Our findings suggest that yaks with low BW, BFT and glucose levels, and higher BHB values were at risk of delayed resumption of ovarian activity and concomitant development of follicular cysts.

Leukaemia inhibitory factor modulates the differentiation of granulosa cells during sheep in vitro preantral to antral follicle development and improves oocyte meiotic competence.

In vitro follicle development from cryopreserved ovarian tissue could become an invaluable assisted reproduction technology for women with early ovarian failure. The challenge lies in producing, from small follicles present in the ovarian cortex, high-quality mature oocytes able to sustain embryo development. In vivo, an optimal combination of hormones and other factors coordinates the development of follicles and their enclosed oocyte. We have investigated the effect of the leukaemia inhibitory factor (LIF) cytokine, alone or in combination with FSH, on sheep in vitro follicle development from the preantral stage onwards. LIF did not alter follicle growth or antrum formation, but it modulated the differentiation of granulosa cells, as revealed by decreased production of anti-Müllerian hormone and abolished FSH-induced stimulation of oestradiol secretion. This modulatory role was also reflected in the abundance of mRNA from 35 genes, analysed by reverse-transcription coupled to microfluidic quantitative PCR. LIF stimulated or at least maintained the expression of genes involved in the dialogue between the oocyte and granulosa cells, through gap junctions (GJA4 encoding connexin 37) or paracrine signalling (Bone morphogenetic protein 15, KIT ligand and their receptors). Finally, the presence of both LIF and FSH during follicle growth strongly improved oocyte meiotic competence: most oocytes (56%) underwent subsequent nuclear maturation, a significant increase compared with their counterparts from follicles of similar size (550-900 µm) cultured with FSH only (28%) or developed in vivo (9%). Their ability to sustain embryo development remains to be evaluated. Combined supplementation with FSH and LIF certainly merits investigation with human follicles.

Perspectives on the development and future of oocyte IVM in clinical practice.

Oocyte in vitro maturation (IVM) is an assisted reproductive technology designed to obtain mature oocytes following culture of immature cumulus-oocyte complexes collected from antral follicles. Although IVM has been practiced for decades and is no longer considered experimental, the uptake of IVM in clinical practice is currently limited. The purpose of this review is to ensure reproductive medicine professionals understand the appropriate use of IVM drawn from the best available evidence supporting its clinical potential and safety in selected patient groups. This group of scientists and fertility specialists, with expertise in IVM in the ART laboratory and/or clinic, explore here the development of IVM towards acquisition of a non-experimental status and, in addition, critically appraise the current and future role of IVM in human ART.

Co-culture with peritoneum mesothelial stem cells supports the in vitro growth of mouse ovarian follicles.

The important roles played by the ovarian microenvironment and cell interactions in folliculogenesis suggest promising approaches for in vivo growth of ovarian follicles using appropriate scaffolds containing suitable cell sources. In this study, we have investigated the growth of early preantral follicles in the presence of decellularized mesenteric peritoneal membrane (MPM), peritoneum mesothelial stem cells (PMSCs), and conditioned medium (CM) of PMSCs. MPM of mouse was first decellularized; PMSCs were isolated from MPM and cultured and their conditioned medium (CM) was collected. Mouse follicles were separated into four groups: (1) culture in base medium (control), (2) culture in decellularized MPM (DMPM), (3) co-culture with PMSCs (Co-PMSCs), and (4) culture in CM of PMSCs (CM-PMSCs). Qualitative and quantitative assessments were performed to evaluate intact mesenteric peritoneal membrane (IMPM) as well as decellularized ones. After culturing the ovarian follicles, follicular and oocyte diameter, viability, eccentric oocyte percentage, and estradiol hormone amounts were evaluated. Quantitative and qualitative evaluations confirmed removal of cells and retention of the essential fibers in MPM after the decellularization process. Follicular parameters showed that Co-PMSCs better support in vitro growth and development of ovarian follicles than the other groups. The eccentric rate and estradiol production were statistically higher for the Co-PMSCs group than for the CM-PMSCs and control groups. Although the culture of early preantral follicles on DMPM and CM-PMSCs could improve in vitro follicular growth, co-culture of follicles with PMSCs showed even greater improvements in terms of follicular growth and diameter.

Integrated stress response control of granulosa cell translation and proliferation during normal ovarian follicle development.

Mechanisms that directly control mammalian ovarian primordial follicle (PF) growth activation and the selection of individual follicles for survival are largely unknown. Follicle cells produce factors that can act as potent inducers of cellular stress during normal function. Consistent with this, we show here that normal, untreated ovarian cells, including pre-granulosa cells of dormant PFs, express phenotype and protein markers of the activated integrated stress response (ISR), including stress-specific protein translation (phospho-Serine 51 eukaryotic initiation factor 2α; P-EIF2α), active DNA damage checkpoints, and cell-cycle arrest. We further demonstrate that mRNAs upregulated in primary (growing) follicles versus arrested PFs mostly include stress-responsive upstream open reading frames (uORFs). Treatment of a granulosa cell (GC) line with the PF growth trigger tumor necrosis factor alpha results in the upregulation of a 'stress-dependent' translation profile. This includes further elevated P-eIF2α and a shift of uORF-containing mRNAs to polysomes. Because the active ISR corresponds to slow follicle growth and PF arrest, we propose that repair and abrogation of ISR checkpoints (e.g. checkpoint recovery) drives the GC cell cycle and PF growth activation (PFGA). If cellular stress is elevated beyond a threshold(s) or, if damage occurs that cannot be repaired, cell and follicle death ensue, consistent with physiological atresia. These data suggest an intrinsic quality control mechanism for immature and growing follicles, where PFGA and subsequent follicle growth and survival depend causally upon ISR resolution, including DNA repair and thus the proof of genomic integrity.

Modulation of folliculogenesis in adult laying chickens by bisphenol A and bisphenol S: Perspectives on ovarian morphology and gene expression.

Both bisphenol A (BPA) and its analog bisphenol S (BPS) are industrial chemicals that have been used to make certain plastic products applied in chicken farms, including food and water containers. They are endocrine disrupting chemicals (EDCs) with xenoestrogenic activities and affect reproductive success in many ways. It was hypothesized that BPA and BPS could adversely affect the folliculogenesis in chickens due to their disruption of the estrogen responses, using either genomic or non-genomic mechanisms. This study investigated the deleterious effects of BPA and BPS on the ovaries when adult layer chickens were orally treated with these EDCs at 50 µg/kg body weight, the reference dose for chronic oral exposure of BPA established by the U.S. EPA. The chickens in both BPA and BPS-treated groups showed a decreased number of the preovulatory follicles. BPA-treated chickens showed a significant decrease in the diameter of F1. Additionally, both BPA and BPS treatments increased the infiltrations of lymphocytes and plasma cells in ovaries. Moreover, it was found that the ovaries of BPS-treated chickens weighed the most among the groups. RNA sequencing and subsequent pathway enrichment analysis of differentially expressed genes revealed that both BPA- and BPS-treatment groups showed significant changes in gene expression and pathways related to reproduction, immune function and carcinogenesis. Taken together, both BPA and BPS are potentially carcinogenic and have deleterious effects on the fertility of laying chickens by inducing inflammation, suggesting that BPS may not be a safe replacement for BPA.

The protective effect of small peptides from Periplaneta americana on hydrogen peroxide-induced apoptosis of granular cells.

This study investigates the protective effect of small peptides from Periplaneta americana (SPPA) on hydrogen peroxide (H2O2)-induced apoptosis of ovarian granular cells. H2O2 was applied to human ovarian granular cells (KGN cell strains). Cell viability was tested by cell counting Kit-8 (CCK-8). Cell apoptosis was tested by flow cytometry, and a cell apoptosis model was established. The model cells were treated with SPPA, and the cell survival rate was monitored using the CCK-8 method. The oxidative stress state of cells was examined using SOD, ROS, MDA, and NO kits. The protein expression levels of SIRT1, p53, and the apoptosis-related gene Caspase3 were measured using Western Blot methodology. Relative to the control group, cell viability declined significantly after the H2O2 treatment only (P < 0.01), while the apoptosis rate increased significantly (P < 0.01). The activity of SOD was weakened significantly (P < 0.01), while the cell levels of ROS, MDA, and NO increased dramatically (P < 0.01). Cell viability dramatically recovered (P < 0.01), and the SOD activity is hugely increased (P < 0.01) after SPPA treatment. In contrast, contents of ROS, MDA, and NO decreased sharply (P < 0.01), and significant dose-response relationships are characterized. Moreover, the H2O2 treatment group showed significantly downregulated expression of SIRT1 (P < 0.01) but significantly upregulated expressions of p53 and Caspase3 (P < 0.01) compared to the control group. Following the SPPA treatment of apoptosis cells, expression of SIRT1 increased significantly, while expressions of p53 and Caspase3 declined significantly (P < 0.01). This study suggests that SPPA inhibits H2O2-induced human KGN cell apoptosis through antioxidation, and the SIRT1/p53 signal pathway mediates the antioxidation.