Primary Follicles Research Articles

Effect of Moringa Leaf Extract (Moringa oleifera Lam.) on the Diameter of the Primary and Secondary Follicles in Female Mice (Mus musculus).

&lt;b&gt;Background and Objective:&lt;/b&gt; Infertility is still a phenomenon in the community, so consuming Moringa leaves (&lt;i&gt;Moringa oleifera&lt;/i&gt; Lam.) is expected to increase fertility. This study aimed to determine the effect of Moringa leaf extract (&lt;i&gt;Moringa oleifera&lt;/i&gt; Lam.) on the diameter of the primary and secondary follicles in female mice (&lt;i&gt;Mus musculus&lt;/i&gt;). &lt;b&gt;Materials and Methods:&lt;/b&gt; This study was an experiment using a Completely Randomized Design (CRD). The population of this study was 45 mice and samples were obtained by a simple random sampling technique from as many as 24 mice with the following criteria: Weight 20-25 g, 2-3 months old, female and in good health. Data analysis was performed through the ANOVA Test with a confidence level of α = 0.05 and further tested for the least significant difference (LSD). &lt;b&gt;Results:&lt;/b&gt; Moringa leaf extract significantly positively affects the diameter of primary and secondary follicles in female mice (p<0.05). The average primary follicle diameter was P&lt;sub&gt;0&lt;/sub&gt; (92.65 μm), P&lt;sub&gt;1&lt;/sub&gt; (124.92 μm), P&lt;sub&gt;2 &lt;/sub&gt;(150.72 μm), P&lt;sub&gt;3&lt;/sub&gt; (175.68 μm) and the average secondary follicle diameter was control (157.17 μm), P&lt;sub&gt;1 &lt;/sub&gt;(171.33 μm), P&lt;sub&gt;2&lt;/sub&gt; (204.57 μm), P&lt;sub&gt;3&lt;/sub&gt; (211.11 μm). Giving Moringa leaf extract (&lt;i&gt;Moringa oleifera&lt;/i&gt; Lam.) significantly increases the diameter of mice's primary and secondary follicles due to the presence of vitamin E in Moringa leaf extract (&lt;i&gt;Moringa oleifera&lt;/i&gt; Lam.). &lt;b&gt;Conclusion:&lt;/b&gt; This can stimulate granulosa cells to secrete the hormone estrogen, causing an increase in the diameter of the primary and secondary follicles.

Primordial follicles remaining in young cancer survivors who received chemotherapy.

Ovarian tissue cryopreservation (OTC) is performed for fertility preservation in cancer patients undergoing chemotherapy. Although anti-Müllerian hormone is used as a marker for ovarian reserve, serum levels do not always correlate with the number of follicles. Additionally, the follicle development stage most affected by chemotherapy is unclear. We examined the association between serum anti-Müllerian hormone levels and the number of remaining primordial follicles after chemotherapy, as well as which follicle stage is most affected by chemotherapy before ovarian cryopreservation. Thirty-three patients who underwent OTC were divided into the chemotherapy (n = 22) and non-chemotherapy (n = 11) groups; their ovarian tissues underwent histological examination. Pathological ovarian damage induced by chemotherapy was assessed. Ovarian volumes were estimated from weights. We compared the number of follicles at each developmental stage as a percentage of primordial follicles between the groups. The relationship between serum anti-Müllerian hormone level and primordial follicle density was analyzed. The chemotherapy group had a significantly lower serum anti-Müllerian hormone level, ovarian volume, and density of developing follicles than the non-chemotherapy group. Serum anti-Müllerian hormone levels correlated with primordial follicle density only in the non-chemotherapy group. The chemotherapy group had significantly lower numbers of primary and secondary follicles. Chemotherapy induces ovarian damage and follicle loss. However, serum anti-Müllerian hormone level does not always reflect the number of primordial follicles after chemotherapy, and chemotherapy more significantly affects primary and secondary follicles than primordial follicles. Many primordial follicles remain in the ovary after chemotherapy, supporting OTC for fertility preservation.

Clinician's perspective on the diagnosis of primary cutaneous B-cell lymphoma.

Of all cutaneous lymphomas, 25% are primary cutaneous B-cell lymphomas (PCBCLs). Of these, primary cutaneous follicle center lymphoma (PCFCL), primary cutaneous marginal zone B-cell lymphoma (PCMZL), and primary cutaneous diffuse large B-cell lymphoma, leg type (PCDLBCL-LT) are the most common subtypes. For the diagnosis of PCBCLs, a biopsy combined with immunohistochemistry and histological examination is the gold standard. PCBCLs are categorized into indolent or intermediate to aggressive subtypes based on their clinical behavior in a clinically oriented approach. PCDLBCL-LT has an aggressive course that spreads to extracutaneous sites in about 45% of cases, whereas PCFCL and PCMZL are indolent diseases. As a result, instrumental staging is advised for PCDLBCL-LT but not for extracutaneous disease after a diagnosis of PCMZL or PCFCL. Lastly, dermatoscopy may offer a novel diagnostic tool to improve the clinical recognition of various PCBCL subtypes when used in conjunction with a strong clinical suspicion.

Subacute exposure to Roundup® changes steroidogenesis and gene expression of the glutathione-glutaredoxin system in rat ovaries: Implications for ovarian toxicity of this glyphosate-based herbicide

Studies have indicated that glyphosate induces endocrine disruption and may adversely affect the male reproductive system. However, evidence of its effects on ovarian function is poorly understood so far, making further studies necessary on the mechanisms of the glyphosate toxicity in the female reproductive system. The aim of this work was to evaluate the effect of a subacute exposure (28 days) to the glyphosate-based formulation Roundup® (1.05, 10.5 and 105 μg/kg b.w. of glyphosate) on steroidogenesis, oxidative stress, systems involved in cell redox control and histopathological parameters in rat ovaries. Hence we quantify plasma estradiol and progesterone by chemiluminescence; non-protein thiol levels, TBARS, superoxide dismutase and catalase activity by spectrophotometry; gene expression of steroidogenic enzymes and redox systems by real-time PCR; and ovarian follicles by optical microscopy. Our results demonstrated that oral exposure increased progesterone levels and the mRNA expression of 3β-hydroxysteroid dehydrogenase. Histopathological analysis revealed a decrease in the number of primary follicles and an increase in the number of corpus luteum in rats exposed to Roundup®. An imbalance of the oxidative status was also evidenced by decreasing the catalase activity at all groups exposed to the herbicide. Increased lipid peroxidation and gene expression of glutarredoxin and decreased of glutathione reductase were also observed. Our results indicate that Roundup® causes endocrine disruption of hormones related to female fertility and reproduction and changes the oxidative status by altering antioxidant activity, inducing lipid peroxidation, as well as changing the gene expression of the glutathione-glutarredoxin system in rat ovaries.

Three-dimensional imaging and quantification of mouse ovarian follicles via optical coherence tomography.

Ovarian tissue cryopreservation has been successfully applied worldwide for fertility preservation. Correctly selecting the ovarian tissue with high follicle loading for freezing and reimplantation increases the likelihood of restoring ovarian function, but it is a challenging process. In this work, we explore the use of three-dimensional spectral-domain optical coherence tomography (SD-OCT) to identify different follicular stages, compare the identifications with H&E images, and measure the size and age-related follicular density distribution differences in mice ovaries. We use the thickness of the layers of granulosa cells to differentiate primordial and primary follicles from secondary follicles. The measured dimensions and age-related follicular distribution agree well with histological images and physiological aging. Finally, we apply attenuation coefficient map analyses to significantly improve the image contrast and the contrast-to-noise ratio (p < 0.001), facilitating follicle identification and quantification. We conclude that SD-OCT is a promising method to noninvasively evaluate ovarian follicles for ovarian tissue cryopreservation.

Primary cutaneous follicle centre lymphoma with secondary systemic evolution

AbstractPrimary cutaneous follicle centre lymphomas (PCFCLs) are cutaneous proliferation of centrocytes and centroblasts, with a generally indolent evolution. Secondary systemic spread of disease is a rare event, which significantly impairs prognosis. Recently published study by Zhou and al. proposed criteria aiming to predict such evolution at initial stage: (i) rearrangement of Bcl2, (ii) Ki67% index &lt;30% and (iii) 2 mutations among the CREBBP, KMT2D, EZH2 and EP300.1 genes. We herein report two cases of PCFCLs with systemic evolution. Both patients had been initially diagnosed with localised skin disease, and developed, several years after, a secondary systemic involvement. Skin biopsies at the initial stage and systemic spread were analysed by histology, immunohistochemistry, fluorescence in situ hybridisation, and by DNA sequencing of a panel of 54 genes frequently mutated in lymphomas. Both patients showed no rearrangement of Bcl2, &gt;30% Ki67% indexes and no mutations among the four proposed genes. These findings run contrary to these previously proposed criteria, highlighting the molecular heterogeneity of this rare disease.

CD30 expression in cutaneous B-cell lymphomas.

CD30 expression has been infrequently described in cutaneous B-cell lymphomas (CBCLs). We examined CD30 expression in reactive lymphoid hyperplasia (RLH) and CBCL and correlated expression with clinicopathologic features. CD30 was examined in 82 CBCL patients and 10 RLH patients that had been evaluated in our cutaneous lymphoma clinics. The CBCL patients included: primary cutaneous follicle center lymphoma (PCFCL), Grade 1/2 systemic/nodal follicular lymphoma (SFL); primary cutaneous marginal zone lymphoma/lymphoproliferative disorder (PCMZL/LPD); systemic marginal zone lymphoma (SMZL); primary cutaneous diffuse large B-cell lymphoma, leg type (PCDLBCL-LT); and extracutaneous/systemic diffuse large B-cell lymphoma (eDLBCL). We scored CD30 expression for intensity and extent and related CD30 expression to age at first diagnosis, sex, site of biopsy, clinical appearance, extracutaneous involvement, multiple cutaneous lesions, B-symptoms, lymphadenopathy, positive positron emission tomography/computed tomography (PET/CT), elevated lactate dehydrogenase (LDH), and positive bone marrow biopsy. CD30 expression was identified in 35% of CBCL, ranging from few, weak, scattered cells to strong and diffuse expression. It was most common in PCFCL and was not expressed in PCDLBCL-LT. Rare PCFCL expressed strong, diffuse CD30. Some cases of PCMZL/LPD, SMZL, FL, and RLH showed scattered, strongly positive cells. CD30 expression in CBCL was associated with favorable clinical features: younger age, negative PET/CT, and an LDH within normal limits. CD30 may be expressed in CBCL, possibly causing diagnostic confusion. CD30 expression was most commonly identified in PCFCL and is associated with favorable clinical features. In cases with strong and diffuse expression, CD30 could be a therapeutic target.

Stem cell therapy for premature ovarian insufficiency: a systematic review and meta-analysis of animal and clinical studies.

The aim of this systematic review and meta-analysis is to evaluate the efficacy of stem cell therapy in mouse models of POI and patients with POI. The PubMed, Web of Science, and Embase databases were searched from inception to February 2022 for relevant animal and clinical studies. The reference lists of the included reviews were manually searched to identify additional eligible studies. Data were independently extracted by two investigators, and disagreements were resolved by discussion. SYRCLE's risk of bias tool and the MINORS tool were used to assess the quality of animal and clinical studies by two independent investigators. All statistical analyses were conducted using Review Manager 5.3 software. A total of twenty animal studies and six clinical studies were included in this meta-analysis. In animal studies, the results showed that stem cells could improve hormone levels, follicle count, estrous cycle and pregnancy outcome. For hormone levels, stem cells increased serum E2 and AMH levels and decreased serum FSH and LH levels compared with the control group (serum E2 level: SMD: 5.05, 95% CI 4.21-5.90, P < 0.00001; serum AMH level: SMD: 4.42, 95% CI 3.06-5.79, P < 0.00001; serum FSH level: SMD: -3.79, 95% CI -4.87 to - 2.70, P < 0.00001; serum LH level: SMD: -1.31, 95% CI -1.65 to -0.96, P < 0.00001). All follicle counts, except for the antral follicle count, were significantly changed compared with the control group. (primordial follicle count: SMD: 4.61, 95% CI 3.65-5.56, P < 0.00001; primary follicle count: SMD: 3.35, 95% CI 1.08-5.63, P = 0.004; secondary follicle count: SMD: 3.23, 95% CI 1.92-4.55, P < 0.00001; total follicle count: SMD: 4.84, 95% CI 2.86-6.83, P < 0.00001; oocyte count: SMD: 7.56, 95% CI 5.92-9.20, P < 0.00001; atretic follicle count: SMD: -1.79, 95% CI -2.59 to -1.00, P < 0.00001). For the estrous cycle, stem cell therapy increased the number of estrous cycles (WMD: 2.72, 95% CI 2.07-3.37, P < 0.00001) and decreased the duration of the estrous cycle (WMD: -1.26, 95% CI -1.84 to -0.69, P < 0.0001) compared with the control group. For pregnancy outcomes, stem cell therapy increased the fertility rate (RR: 3.00, 95% CI 1.74-5.17, P < 0.0001) and litter size (WMD: 3.82, 95% CI 0.36-7.28, P = 0.03) compared with the control group. In animal studies, the asymmetric funnel plot of serum E2 and FSH levels indicated the possibility of publication bias. Unpublished and negative studies may be the source of publication bias. In clinical studies, the results showed that stem cell therapy could decrease serum FSH level (MD: -30.32, 95% CI -59.03 to -1.01, P = 0.04) and increase AFC (MD: 1.07, 95% CI 0.70-1.43, P < 0.00001), pregnancy rate (RD: 0.19, 95% CI 0.04-0.34, P = 0.01) and live birth rate (RD: 0.19, 95% CI 0.07-0.31, P = 0.001) in POI patients. In addition, there was no significant difference in menstrual function regained (RD: 0.22, 95% CI -0.03-0.46, P = 0.09), oocytes retrieved (MD: 1.00, 95% CI -0.64-2.64, P = 0.23) and embryos (MD: 0.80, 95% CI -0.15-1.76, P = 0.10) between different groups. This meta-analysis suggested that stem cell therapy might be effective in POI mouse models and patients and could be considered a potential treatment to restore fertility capability in POI patients.

Inhibition of phosphodiesterase PDE8B reduces activation of primordial follicles in mouse ovaries.

In the ovaries, cyclic adenosine 3',5'-monophosphate (cAMP) is a second messenger supporting the generation of steroids. Phosphodiesterases (PDEs) are regulators of intracellular cAMP, and therefore, potential regulators of ovarian function. Interestingly, the family of PDE genes are differentially expressed in human oocytes and granulosa cells from primordial and primary follicles, suggesting diverse roles. In this study, we addressed the functions of PDE3B and PDE8B in primordial follicle regulation using inhibitors of PDE3B and PDE8B in murine ovary primary in vitro cultures. Inhibition of PDE8B in ovarian cultures prevented primordial follicle activation, while inhibition of PDE3B had no effect on follicle distribution in the ovary, under the tested conditions. As cAMP levels may increase steroid levels, we assessed the protein levels of the steroidogenic acute regulatory protein (StAR) and aromatase enzymes, and found that inhibition of PDE3B reduced StAR protein levels, whereas inhibition of PDE8 did not alter StAR expression in our murine ovary culture system conditions. Our results showed that ketotifen-induced inhibition of PDE8B can decrease primordial follicle activation, whereas we observed no effect of follicle distribution, when PDE3B was inhibited. Expression of the StaR enzyme was not altered when PDE8B was inhibited, which might reflect not sufficient inhibition by ketotifen to induce StAR alterations, or redundant mechanisms.

What is new about primary cutaneous B cell lymphomas.

Primary cutaneous B-cell lymphomas (CBCL) are a heterogeneous group of B-cell lymphomas without evidence of extracutaneous disease at the time of diagnosis. The 2022 World Health Organization classification of mature lymphoid neoplasms differentiates the indolent primary cutaneous marginal zone lymphoproliferative disorder, primary cutaneous follicle center lymphoma and Epstein-Barr virus-positive mucocutaneous ulcer, from the more aggressive primary cutaneous diffuse large B-cell lymphoma, leg-type and intravascular large B-cell lymphoma. The new updates in the 2022 classification are based on recent scientific advances in the understanding and characterization of these entities. This article aims to review the main clinical, cellular and molecular features of the five CBCL subsets along with their management and treatment. The exponentially growing evidence for new treatment options for systemic B-cell lymphomas raises expectations for the field of CBCL as well. However, specific prospective high quality research on CBCL is still crucial to further define their management and update international guidelines.

Effects of Ethanol Extract of Sungkai (Peronema canescens Jack.) on Fertility of Female Wistar Mice (Mus musculus L.)

Sungkai leaf (Peronema canescens Jack.) is a natural source of flavonoids. Flavonoids have been implicated in increasing fertility in humans. This study aims to determine the effect of oral ethanol extract of Sungkai leaf on the fertility of female mice. Twenty-four female mice randomly divided into four groups were used in this study. The test (II-IV) groups were administered orally ethanolleaf extract of Peronema canescens at doses of 200 mg/kg bw, 400 mg/kg bw, and 800 mg/kg bw, respectively, for 14 days. The negative control (I) group received 0.5% Na-CMC only. On the 15th day, the mice were mated with male mice in one cage until pregnancy was established. On the 10th day of gestation, the mice were sacrificed by cervical dislocation, and both ovaries wereexcised. The result shows increased primary ovarian follicles, corpus luteum, and ovulation rate. The optimal increase of the ovaries occurred at the dose of 200 mg/kg bw. There was however a decrease in the above parameters at extract doses of 400 and 800 200 mg/kg bw. The study concludes that the leaf extract of Peronema canescens has the potential to treat infertility.

Effect of induced molting on ovarian function remodeling in laying hens

Induced molting (IM) can restore the laying rate of aged laying hens to the peak level of laying and rejuvenate ovarian function for the second laying cycle. To explore the mechanism of ovarian function remodeling during IM in laying hens, in this study, ninety 71-wk-old laying lady hens with 60% laying rate and uniform weight were selected for molting induction by fasting. Samples (serum and fresh ovarian tissue) were collected on the day before fasting (F0), the 3rd and 16th days of fasting (F3, F16), and the 6th, 15th, 32nd days of refeeding (R6, R15, and R32), and the number of follicles in each period was counted. Then, the reproductive hormone levels in serum and antioxidant levels in ovarian tissues were detected at different stages, and the gene expression of the KIT-PI3K-PTEN-AKT pathway and GDF-9 in ovaries was measured by qRT-PCR. The results showed that the laying rate increased rapidly after refeeding and returned to the prefasting level by R32. At F16 and R6, the number of mature follicles significantly decreased; the number of primary and secondary follicles significantly increased; the contents of follicle stimulating hormone (FSH), luteinizing hormone (LH), estradiol (E2), and progesterone (P4) in serum decreased; the relative expression of KIT, PI3K, AKT, and GDF-9 significantly increased; and that of PTEN significantly decreased. At R15 and R32, except for GDF-9, which maintained a high expression state, other indicators showed opposing trends to those observed at F16 and R6. In conclusion, IM activated the KIT-PI3K-PTEN-AKT signaling pathway and promoted the activation of primordial follicles during the fasting period and early resumption of feeding; gonadotropin secretion increased gradually, which promoted the rapid development of primary and secondary follicles to mature follicles and ovulation. This study explained the mechanism of ovarian function remodeling in the process of IM and provided a theoretical basis for improving the ovarian function of laying hens and optimizing the IM program.

The transcriptome reveals the molecular regulatory network of primordial follicle depletion in obese mice

To explore the dynamic transcriptional regulatory network of primordial follicle fate in obese mice to elucidate the potential mechanism of primordial follicle depletion. Experimental study and transcriptomic analysis. Healthy (n=15) and obese (n=15) female mice. Six-week-old CD-1 mice were divided into healthy and high-fat diet groups and fed continuously for 12 weeks. The diet of healthy mice contained 10% fat. The diet of high-fat mice contained 60% fat. Primordial to primary follicle transition rate, gene expression changes, enriched Kyoto Encyclopedia of Genes and Genomes pathways, and ferroptosis. Primordial follicle depletion was increased in the ovaries of obese mice. We found that deposited fat around primordial and primary follicles of obese mice was higher than that for healthy mice. The proliferation of granulosa cells around primary follicles was increased in obese mice. In addition, we uncovered specific gene signatures associated with the primordial to primary follicle transition (PPT) in obese mice using laser capture microdissection RNA sequencing analysis. Gene set enrichment analysis indicated that ferroptosis, cell oxidation, vascular endothelial growth factor, and mammalian target of rapamycin signaling were increased significantly in the primordial follicles of obese mice. Notably, the ferritin, acyl CoA synthetase long-chain family member 4, and solid carrier family 7 member 11 associated proteins of the ferroptosis signaling pathway were significantly increased in the PPT phase of obese mice. Our work suggests that ferroptosis is a key pathway activated within immature ovarian follicles in the context of obesity and that the process may be involved in the physiological regulation of the PPT as well.

Cutaneous B-cell lymphomas: 2023 update on diagnosis, risk-stratification, and management.

Approximately one-fourth of primary cutaneous lymphomas are B-cell derived and are generally classified into three distinct subgroups: primary cutaneous follicle center lymphoma (PCFCL), primary cutaneous marginal zone lymphoma (PCMZL), and primary cutaneous diffuse large B-cell lymphoma, leg type (PCDLBCL, LT). Diagnosis and disease classification is based on histopathologic review and immunohistochemical staining of an appropriate skin biopsy. Pathologic review and an appropriate staging evaluation are necessary to distinguish primary cutaneous B-cell lymphomas from systemic B-cell lymphomas with secondary skin involvement. Disease histopathology remains the most important prognostic determinant in primary cutaneous B-cell lymphomas. Both PCFCL and PCMZL are indolent lymphomas that infrequently disseminate to extracutaneous sites and are associated with 5-year survival rates that exceed 95%. In contrast, PCDLBCL, LT is an aggressive lymphoma with an inferior prognosis. PCFCL and PCMZL patients with solitary or relatively few skin lesions may be effectively managed with local radiation therapy. While single-agent rituximab may be employed for patients with more widespread skin involvement, multiagent chemotherapy is rarely appropriate. In contrast, management of patients with PCDLBCL, LT is comparable to the management of patients with systemic DLBCL.

The Gonadotoxic Effects of Conditioning Chemotherapy Regimens Prior to Hematopoietic Stem Cell Transplantation [ID: 1376168

INTRODUCTION: Primary ovarian insufficiency secondary to high-dose chemotherapy in females undergoing the conditioning regimen before hematopoietic stem cell transplantation is a challenging late complication. Limited studies have investigated the ovarian toxicity of various conditioning regimens. This study aimed to explore the gonadotoxic effects of cyclophosphamide (CY) and busulfan (BU) in CD-1 mice strain. METHODS: Sixteen 1-month-old female CD-1 mice (n=4/group) were subcutaneously injected with solvent, BU, CY, or BU+CY, respectively. The doses (144 mg/kg BU and 300 mg/kg CY) and injection plan were adjusted to human doses. The ovaries and female reproductive tracts were harvested 1 week after the last administration. Follicle counting was performed. The levels of serum antimüllerian hormone (AMH) and 17-beta-estradiol were measured. Animal use approval was obtained from the Institutional Animal Care and Use Committee at the University of Nebraska Medical Center. RESULTS: All mice survived the injections and grossly appeared normal. The average ovary weight was significantly lower in BU (P=.0007) and BU+CY (P&lt;.0001) groups than in the control and CY groups. The primordial (P&lt;.0001) and primary follicles (P=.01) in treatment groups were decreased compared to the control group. The serum AMH level in the BU group was also significantly lower than the control and CY group (P=.003). CONCLUSION: We show that conditioning chemotherapy with CY and BU can reduce the ovarian reserve in CD-1 mice strain and may have a synergic effect. Future studies to investigate the mechanism of chemotherapy-related ovarian follicle death are needed to improve fertility outcomes in patients receiving conditioning chemotherapy.

Review: Roles of follicle-stimulating hormone in preantral folliculogenesis of domestic animals: what can we learn from model species and where do we go from here?

The pituitary gonadotropin FSH is a glycoprotein critical for the development of ovarian follicles. Upon binding to its G protein-coupled membrane receptor located on the granulosa cells of ovarian follicles, FSH elicits a cascade of downstream intracellular responses to promote follicle growth, maturation and steroidogenic activity, leading to the acquisition of meiotic and developmental competence of the enclosed oocyte. The essential role of FSH for proper antral follicle development and fertility is indisputable; over the decades, increasing evidence has also pointed toward survival and growth-promoting effects elicited by FSH in earlier-stage preantral follicles, deeming these follicles FSH-responsive as opposed to the FSH-dependent antral follicles. Transgenic mouse models lacking GnRH1, Fshβ or Fshr clearly demonstrate this difference by showing that, morphologically, preantral follicles develop to the secondary stage without FSH signaling; however, exogenous expression or administration of FSH to hormone-deficient mice promotes preantral follicle development, with more pronounced effects seen in earlier stages (i.e., primary follicles). In hypophysectomized sheep, FSH administration also promotes the growth of primary-stage preantral follicles. However, in vivo studies in this area are more challenging to perform in domestic animals compared to rodents, and therefore most of the research to date has been done in vitro. Here, we present the existing evidence for a role of FSH in regulating the growth and survival of preantral follicles from data generated in rodents and domestic animals. We provide an overview of the process of folliculogenesis, FSH synthesis and cellular signaling, and the response to FSH by preantral follicles in vivo and in vitro, as well as interactions between FSH and other molecules to regulate preantral folliculogenesis. The widespread use of FSH in ovarian stimulation programs for assisted reproduction creates a real need for a better understanding of the effects of FSH beyond stimulation of antral follicle growth, and more research in this area could lead to the development of more effective fertility programs. In addition to its importance as an agricultural species, the cow provides a desirable model for humans regarding ovarian stimulation due to similar timing of folliculogenesis and follicle size, as well as similar ovarian architecture. The refinement of minimally invasive methods to allow the study of preantral folliculogenesis in live animals will be critical to understand the short- and long-term effects of FSH in ovarian folliculogenesis.

044 Gene expression profiling using next generation sequencing of primordial, primary and secondary bovine follicles

044 Gene expression profiling using next generation sequencing of primordial, primary and secondary bovine follicles

Age-related Changes in Humanin Expression in the Ovarian Tissue of Rat.

This study examined humanin expression in rat ovarian tissue, its cellular localization, and its correlation with rat age under physiological conditions. A total of 40 Sprague-Dawley rats of various ages (2, 12, 30, and 60 days old and 1 year old) were grouped by age. Immunofluorescence and immunohistochemical techniques were used to observe humanin expression and cellular location in the ovarian tissues of rats from each age group. In addition, Western blotting and Real-time quantitative reverse transcription PCR (qRT-PCR) techniques were used to measure humanin expression level in the ovarian tissues of rats from each age group. The immunofluorescence and immunohistochemical results confirmed that humanin was expressed in rat ovarian tissues. In addition, cellular localization analysis showed that humanin was expressed in the cytoplasm of oocytes, interstitial cells, granulosa cells and theca cells in all levels of follicles after the primary follicles, and in the corpus luteum. The qRT-PCR results revealed that the level of humanin expression in the ovarian tissues of 12-day-old rats was non-significantly higher than that in the ovarian tissues of 2-day-old rats (P>0.05), whereas the levels of humanin expression in the ovarian tissues of 30-day-old, 60-day-old, and 1-year-old rats were significantly lower than that in the ovarian tissues of 2-day-old rats (P<0.05). The Western blotting results demonstrated that the levels of humanin protein expression in the ovarian tissues of 60-day-old and 1-year-old rats were significantly lower than those of 2-day-old rats (P<0.01), whereas there was no significant difference in the level of humanin protein expression between the ovarian tissues of 12-day-old and 30-day-old rats. This study confirmed that humanin is expressed in the cytoplasm of various cells in rat ovarian tissues. Moreover, the level of humanin expression was highest in the ovarian tissues of 12-day-old rats, and it subsequently decreased with age. The changes in the expression of humanin in the ovary of rats at different ages will lay the foundation for the role of humanin in ovarian aging. The effect of humanin on ovarian function is worthy of further study in the future.

Preantral follicle numbers and size in heifers carrying Trio, a bovine high fecundity allele.

The bovine high fecundity allele, Trio, results in the occurrence of multiple ovulations and is characterized by antral follicles that develop slower and acquire ovulatory capacity at smaller sizes. This study provides novel information on the effect of the Trio allele on early folliculogenesis. The bovine high fecundity allele, Trio, causes overexpression in granulosa cells (GCs) of SMAD6, an inhibitor of BMP15-activated SMAD signalling. Furthermore, the Trio allele results in antral follicles that develop slower, acquire ovulatory capacity at smaller sizes, and have three-fold greater ovulation rate compared to half-sib non-carriers. The present study was designed to determine preantral follicle numbers and size in Trio carrier and non-carrier cattle testing the hypothesis that inhibition of SMAD signalling would alter preantral follicle activation and/or growth. Ovarian tissues from Trio carrier (n = 12) and non-carrier (n = 12) heifers were obtained by laparotomy after follicle wave synchronization. Follicle numbers and dimensions were determined for each stage of development (primordial, transitional, primary, and secondary) from paraffin-embedded sections. There were no differences in the number of primordial, transitional, or secondary follicles or in antral follicle count, circulating AMH, or ovarian volume between carriers and non-carriers. Trio carriers had ~2.5-fold greater (P < 0.01) number of primary follicles than non-carriers, and transitional and primary follicles were larger (~1.2-fold; P < 0.1) in Trio carriers. Oocyte volume of primordial and transitional follicles was not different between genotypes; however, oocytes were larger (P < 0.05) in primary (~1.3-fold) and secondary (~1.8-fold) follicles for Trio carriers. Granulosa cell numbers were not different (P > 0.3) between carriers and non-carriers, irrespective of the stage of development. These results suggest that, after primordial follicle activation, follicles in Trio carrier cattle have slower progression through the primary stage, hence the larger oocyte and greater number of primary follicles.

Pubertal lead exposure affects ovary development, folliculogenesis and steroidogenesis by activation of IRE1α-JNK signaling pathway in rat

Epidemic studies showed that lead exposures are associated with various female reproductive dysfunctions, including infertility, miscarriage, preterm delivery, and early menopause. However, the mechanism involved is still unclear. In the current study, SD rats were exposed to lead at doses of 0, 5, 25, 50 or 250 mg/L through drinking water from postnatal day 21–56. Lead exposures did not affect the body weight or ovary weight. However, the puberty initiation (ages by which vagina opens and estrous cycle occurs) was significantly delayed by as many as 5.8 and 6.8 days respectively (P < 0.05). Also, lead exposures disrupted the estrous cycles, reduced the numbers of primordial and primary follicles and increased the number of atretic follicles by adult. Furthermore, for the highest does group, serum levels of progesterone and testosterone decreased by 80.2% (P < 0.01) and 49.9% (P < 0.05) respectively, while estradiol level increased by 69.8% (P < 0.01). Western blot analyses indicated that lead exposures specifically down-regulated the expressions of steroidogenic protein STAR, CYP17A1, and HSD3B1, while up-regulated FSHR and CYP19A1. Also, the exposure stimulated the endoplasmic reticulum stress (ERS)-related IRE1α-JNK signaling pathway members. Such activation may also result in apoptosis since the death-signaling molecules CHOP and cleaved-CASP3 were up-regulated while BCL2 was down-regulated. In conclusion, lead exposure during juvenile and puberty significantly affected ovary development and functions. The effects may relate to ERS response since the 6 members related to the pathway were all consistently activated.