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Polycystic ovary syndrome (PCOS), a common endocrine disorder, is associated with impaired oocyte development, leading to infertility. However, the pathogenesis of PCOS has not been completely elucidated. This study aimed to determine the differentially expressed genes (DEGs) and epigenetic changes in the oocytes from a PCOS mouse model to identify the etiological factors. RNA-sequencing analysis revealed that 90 DEGs were upregulated and 27 DEGs were downregulated in mice with PCOS compared with control mice. DNA methylation analysis revealed 30 hypomethylated and 10 hypermethylated regions in the PCOS group. However, the DNA methylation status did not correlate with differential gene expression. The pathway enrichment analysis revealed that five DEGs (Rps21, Rpl36, Rpl36a, Rpl37a, and Rpl22l1) were enriched in ribosome-related pathways in the oocytes of mice with PCOS, and the immunohistochemical analysis revealed significantly upregulated expression levels of Rps21 and Rpl36. These results suggest that differential gene expression in the oocytes of mice in PCOS is related to impaired folliculogenesis. These findings improve our understanding of PCOS pathogenesis. [ABSTRACT FROM AUTHOR]

Purpose: Human follicular fluid constitutes the microenvironment of follicles and includes various biological active proteins that can affect follicle growth and oocyte fertilization. Conducting proteomic evaluations of human follicular fluid may be helpful for identifying potential biomarkers possibly possessing a predictive value for oocyte quality and the success of in vitro fertilization. Method: We performed proteomic profiling of human follicular fluids containing oocytes that were fertilized and resulted in pregnancy and follicular fluids containing oocytes that were not fertilized in the same patients undergoing intracytoplasmic sperm injection using the LTQ Orbitrap coupled with liquid chromatography-tandem mass spectrometry (LC/MS/MS) analyses. Results: We identified a total of 503 proteins in human follicular fluids containing fertilized and non-fertilized oocytes obtained from 12 patients. We also found that 53 proteins exhibited significantly different spectral counts between the two groups, including heparan sulfate proteoglycan perlecan, which showed significant upregulation in the follicular fluids containing fertilized oocytes in comparison with that observed in the follicular fluids containing non-fertilized oocytes. Conclusion: Our results suggest a possibility that proteins identified by LC/MS/MS in follicular fluid might not only be involved in folliculogenesis, but also function as biomarkers possessing predictive potential for oocyte maturation and the success of IVF when their expression levels are significantly different between fertilized and non-fertilized oocytes, although no distinctive biomarkers were identified in the current study. [ABSTRACT FROM AUTHOR]

Our previous study found that 17β-estradiol (E2) suppresses primordial follicle activation and growth in cultured mouse ovaries. In this study, we administered tamoxifen, an estrogen receptor antagonist, into the abdominal cavity of mice to clarify the relationship between primordial follicle activation and the physiological concentration of E2 in mouse ovaries. The results showed that tamoxifen promoted primordial follicle activation. Administration of tamoxifen promoted degradation of the extracellular matrix surrounding primordial follicles in the ovaries. Furthermore, tamoxifen decreased the expression of stefin A, an inhibitor of cathepsins that digest some proteins and extracellular matrix, in the ovaries. Mechanical stress produced by the extracellular matrix reportedly suppresses the activation of primordial follicles. The collective results show that tamoxifen can promote primordial follicle activation through the degradation of the extracellular matrix surrounding primordial follicles. Our results indicate that E2 suppresses primordial follicle activation in vivo and that tamoxifen may be useful as a therapeutic agent against infertility. [ABSTRACT FROM AUTHOR]

Patients with primary ovarian insufficiency (POI) occasionally present with follicle growth; however, accurately predicting cycles accompanied by follicle growth is challenging. Early-stage follicles produce serum anti-Müllerian hormone (AMH), a useful marker of ovarian reserve. Therefore, serum AMH levels indicate growth of small follicles (which are difficult to detect ultrasonographically) and may predict follicle growth in patients with POI. Using an ultrasensitive enzyme-linked immunosorbent assay (ELISA) kit, we observed very low serum AMH levels in patients with POI. We further evaluated follicle growth in each patient during each cycle to determine the usefulness of measuring serum AMH levels as a predictor of follicle growth in patients with POI who receive hormone replacement therapy (HRT). We investigated 19 patients with POI in whom we analyzed 91 cycles; 14 cycles showed positive and 77 cycles showed negative results on serum AMH testing. The rate of cycles showing follicle growth in AMH-positive cycles was higher than that in AMH-negative cycles (64.3% vs. 6.5%, p = 0.0001). The median serum AMH level (7.7 pg/mL [25th and 75th percentiles 4.6 pg/mL and 22.3 pg/mL, respectively]) in AMH-positive cycles was lower than the lower limit of detection of conventional AMH ELISA kits. The positive predictive value of positive serum AMH levels for follicle growth was higher than that of follicle-stimulating hormone (< 10 mIU/mL). These results indicate that a very low level of serum AMH detected using picoAMH assays is a useful predictor of follicle growth in patients with POI receiving HRT. [ABSTRACT FROM AUTHOR]

Purpose: Follicle-stimulating hormone receptor (FSHR) expression in granulosa cells is critical in enabling follicles to achieve accelerated growth. Although FSHR expression has been reported to be epigenetically regulated, the mechanism is unclear. Cooperation between oocytes and granulosa cells is also essential for normal follicular growth. Among oocyte-derived factors, bone morphogenetic protein 15 (BMP15) promotes follicular growth and is suggested to have epigenetic effects. We examined the role of BMP15 in the acquirement of FSHR in human granulosa cells. Methods: Immortalized non-luteinized human granulosa (HGrC1) cells were stimulated with trichostatin A (TSA) or BMP15 to analyze FSHR expression, histone modifications, and USF1/2 binding at the FSHR promoter region. Histone acetyl transferase (HAT) activity and phosphorylation of Smad 1/5/8 and p38 MAPK were examined with or without BMP15, SB203580, and LDN193189. CYP19A1 expression and estradiol production were also studied. Results: TSA and BMP15 induced FSHR mRNA expression in a dose-dependent manner and histone modifications were observed with increased binding of USF1/2. BMP15 increased FSHR protein expression, which was suppressed by LDN193189. BMP15 increased phosphorylation of Smad 1/5/8 and significantly increased HAT activity, which was inhibited by LDN193189, but not by SB203580. BMP15 increased phosphorylation of p38 MAPK and USF1. LDN193189 suppressed BMP15-induced phosphorylation of both p38 MAPK and USF1, whereas SB203580 suppressed the phosphorylation of USF1. BMP15 increased CYP19A1 mRNA expression and estradiol production. Conclusion: BMP15 induced FSHR expression in human granulosa cells through Smad and non-Smad pathways. This mechanism of FSHR induction by BMP15 may be utilized for controlling follicular growth. [ABSTRACT FROM AUTHOR]

Growth differentiation factor 9 (GDF9), an oocyte-secreted factor, whose receptors exist in granulosa cells, is involved in follicle progression. Therefore, GDF9 is considered to potentially mediate signals necessary for follicular growth. However, the effect of GDF9 on human granulosa cells is not fully understood. Human immortalized nonluteinized granulosa cell line (HGrC1) which we have previously reported was stimulated with GDF9 and/or follicle-stimulating hormone (FSH). Granulosa cells obtained from in vitro fertilization (IVF) patients were also evaluated with quantitative reverse transcription polymerase chain reaction (RT-PCR). Real-time RT-PCR showed that GDF9 increased messenger RNA (mRNA) levels of enzymes required for cholesterol biosynthesis, such as 3-hydroxy-3-methylglutanyl-CoA synthase 1 (HMGCS1), farnesyl-diphosphate farnesyltransferase 1, squalene epoxidase, lanosterol synthase, and cytochrome P450, family 51, subfamily A, polypeptide 1 (CYP51A1). A greater increase in mRNA levels of HMGCS1 and CYP51A1 was observed by combined treatment with GDF9 and FSH. Clinical samples showed a significant increase in CYP51A1 mRNA in the group of granulosa cells connected with unfertilized oocytes. Our results suggest that GDF9, possibly with FSH, may play significant roles in the regulation of cholesterol biosynthesis and the expression of CYP51A1 which might be a predictor for unfertilization. [ABSTRACT FROM PUBLISHER]

Background: Endometriosis is a complex syndrome characterized by an estrogen-dependent chronic inflammatory process that affects 10% of women of reproductive age. Ovarian endometriosis (OE) is the most common lesion in endometriosis and may cause infertility, in addition to dysmenorrhea. Hormonal treatments, which are the conventional treatment methods for endometriosis, suppress ovulation and hence are not compatible with fertility. The inflammasome is a complex that includes Nod-like receptor (NLR) family proteins, which sense pathogen-associated molecular patterns and homeostasis-altering molecular processes. It has been reported that the nucleotide-binding oligomerization domain, leucine-rich repeat, and pyrin domain-containing (NLRP) 3 inflammasome, which contributes to the activation of interleukin-1 beta (IL-1β), might be related to the progression of endometriosis. Therefore, the aim of the present study was to evaluate non-hormonal therapies for OE, such as inhibitors of the NLRP3 inflammasome. Methods: The expression of NLRP3 was measured in the eutopic endometrium (EM) of patients with and without endometriosis and OE samples, as well as stromal cells derived from the endometrium of patients with and without endometriosis and OE samples (endometrial stromal cells with endometriosis [ESCs] and cyst-derived stromal cells [CSCs]). The effects of an NLRP3 inhibitor (MCC950) on ESCs and CSCs survival and IL-1β production were evaluated. We then administered MCC950 to a murine model of OE to evaluate its effects on OE lesions and ovarian function. Results: NLRP3 gene and protein expression levels were higher in OE and CSCs than in EM and ESCs, respectively. MCC950 treatment significantly reduced the survival of CSCs, but not that of ESCs. Moreover, MCC950 treatment reduced the co-localization of NLRP3 and IL-1β in CSCs, as well as IL-1β concentrations in CSCs supernatants. In the murine model, MCC950 treatment reduced OE lesion size compared to phosphate-buffered saline treatment (89 ± 15 vs. 49 ± 9.3 mm3 per ovary; P < 0.05). In the MCC950-treated group, IL-1β and Ki67 levels in the OE-associated epithelia were reduced along with the oxidative stress markers of granulosa cells. Conclusions: These results indicated that NLRP3/IL-1β is involved in the pathogenesis of endometriosis and that NLRP3 inhibitors may be useful for suppressing OE and improving the function of ovaries with endometriosis. [ABSTRACT FROM AUTHOR]

Background: Ovarian endometrioma is a common gynecological disease that is often treated with surgery or hormonal treatment. Ovarian cystectomy, a surgical procedure for ovarian endometrioma, can result in impaired ovarian reserve. Methods: We conducted a randomized controlled trial to evaluate the efficacy of hormonal treatment [gonadotropin-releasing hormone agonist (GnRHa) or dienogest (DNG)] for preserving ovarian reserve after cystectomy for ovarian endometrioma. The primary endpoint was the level of serum Anti-Müllerian hormone (AMH) as a marker of ovarian reserve. Results: Before and after laparoscopic surgery, 22 patients in the GnRHa group and 27 patients in the DNG group were administered hormonal treatment for a total of 4 months. After 1-year follow-up, >60% of the patients in the DNG group retained over 70% of their pretreatment AMH levels, whereas no patient in the GnRHa group retained their AMH levels after cystectomy (P < 0.01). Interleukin-6 (IL-6) is a key cytokine involved in inflammation. Compared with the GnRHa group, patients in the DNG group had lower IL-6 levels at the end of treatment. Conclusions: Our data revealed that DNG is more effective than GnRHa in preserving ovarian reserve after cystectomy of ovarian endometrioma. This is achieved through the reduction of the inflammatory response during the perioperative period and other endometriosis-related inflammatory reactions. Trial registration: The registration number of this trial is UMIN-CTR, UMIN000018569, registered 6 August 2015, https://upload.umin.ac.jp/cgi-open-bin/ctr%5fe/ctr%5fview.cgi?recptno=R000021492, and Japan Registry of Clinical Trials, jRCTs041180140, registered 29 March 2019, https://jrct.niph.go.jp/en-latest-detail/jRCTs041180140. This randomized controlled trial was conducted in accordance with the CONSORT guidelines. [ABSTRACT FROM AUTHOR]

BACKGROUND To assess the impact of ovarian cystectomy for endometriomas on the ovarian reserve, we evaluated the pre- and post-operative levels of serum anti-Müllerian hormone (AMH). We also analyzed the correlations between factors related to endometriosis and surgery for endometriomas and the serum AMH levels to investigate which factors affect ovarian reserve. METHODS Thirty-eight patients who were undergoing ovarian cystectomy for unilateral endometrioma (n = 20) and bilateral endometriomas (n = 18) participated. Preoperative and post-operative serum samples were collected and assayed for AMH levels, and changes between the two samples were analyzed in association with parameters of endometriosis and surgery for endometriomas. RESULTS The mean AMH level was 3.9 ng/ml prior to surgery, and was reduced to 2.1 ng/ml at 1 month post-surgery. The rate of decline of the serum AMH level was significantly higher in the bilateral group than the unilateral group (62.8 ± 29.6 versus 24.7 ± 32.5%, P < 0.001). The rate of decline in the serum AMH levels showed a significant correlation to the revised American Society for Reproductive Medicine (rASRM) score (P = 0.003), but not age, cyst diameter, blood loss during the operation or the number of follicles removed in the specimens. CONCLUSIONS Our results suggest that the decrease in ovarian reserve should be taken into account in patients indicated for cystectomy for bilateral endometriomas or unilateral endometrioma with high rASRM scores. [ABSTRACT FROM AUTHOR]

Endometriosis has several distinguishing features in the ectopic endometrium, including chronic inflammation and fibrosis. According to the retrograde menstruation theory, endometriotic cells are derived from eutopic endometrial cells, and adhesion of endometrial cells to the extracellular matrix can be the initial step in the development of endometriosis. Therefore, we hypothesized that cell adhesion, which mediates a sequence of events in the development of endometriosis triggering inflammatory responses and tissue fibrosis could be a possible therapeutic target for endometriosis. We found co-upregulation of focal adhesion kinase (FAK) and monocyte chemoattractant protein-1 (MCP-1) in the endometriotic tissues compared with that in the normal endometrium. MCP-1 secretion was significantly higher in the endometriotic stromal cells than in the eutopic endometrial stromal cells. Furthermore, co-culture of U937 cells and endometriotic stromal cells upregulated secretion of transforming growth factor-β1 (TGF-β1). A FAK inhibitor significantly inhibited the secretion of MCP-1 in the endometriotic stromal cells and TGF-β1 in the co-culture with macrophages. FAK inhibitor treatment in the murine endometriosis model demonstrated a decrease in the formation of endometriotic lesions as well as the expression of MCP-1 and TGF-β1. Our results suggest that the FAK-mediated sequential development of endometriosis, including inflammatory response and tissue fibrosis, can be a new therapeutic target in endometriosis. [ABSTRACT FROM AUTHOR]

Several features exist that distinguish endometriotic cells from eutopic endometrial cells. Progesterone resistance is one of the main distinguishing features, although how progesterone resistance affects the phenotype of endometriotic cells is not fully elucidated. Heart and neural crest derivatives expressed 2 (HAND2) is a transcriptional factor that plays an important role in maintaining endometrial function in a progesterone-dependent manner. Therefore, we explored whether progesterone-dependent HAND2 is implicated in the progression of endometriosis. HAND2 was less expressed by endometriotic tissues compared to endometrial tissues. Suppression of HAND2 expression induced fibroblast growth factor 1 (FGF1), FGF2, and FGF9 in endometriotic stromal cells and consequently enhanced migration and invasion capacity. AZD4547, a FGF receptor inhibitor, diminished the migration and invasion of endometriotic cells in vitro. In the murine model of endometriosis, AZD4547 showed suppressive effects on the development of endometriotic lesions at a relatively low concentration. In conclusion, we demonstrated that FGF1, FGF2, and FGF9 are downstream effectors of HAND2 in endometriotic cells. Since HAND2-dependent FGFs play roles in enhancing invasive capacity of endometriotic cells, our results suggest that FGF receptor inhibitors, such as AZD4547, can be promising therapeutic targets for endometriosis. [ABSTRACT FROM AUTHOR]

Patients with primary ovarian insufficiency (POI) have a high prevalence of thyroid autoimmune disorders. However, the extent of the contribution of thyroid autoantibodies or elevated thyroid-stimulating hormone (TSH) levels to decreased ovarian reserve is unclear. Therefore, we evaluated the serum levels of anti-Müllerian hormone (AMH) and thyroid autoantibodies [antithyroperoxidase antibody (TPOAb), and antithyroglobulin antibody (TgAb)] in euthyroid infertile women. One hundred and fifty-three women with normal menstrual cycles were recruited for this retrospective study. Serum levels of AMH were compared between patients with positive and negative thyroid autoantibodies. The correlation between serum levels of AMH and each thyroid autoantibody was also evaluated. Participants were observed to be either TPOAb or TgAb positive (n = 27), only TPOAb positive (n = 8), only TgAb positive (n = 7), TPOAb and TgAb positive (double positive; n = 12), and TPOAb and TgAb negative (double negative; n = 126). No significant differences were found in serum AMH levels between the TPOAb- or TgAb-positive women and the antibody-double negative women. Serum AMH levels did not show a significant correlation with the concentration of TgAb or TPOAb. On the other hand, serum AMH levels negatively correlated with TSH levels in patients who were either positive for TPOAb or TgAb. Thyroid autoantibodies are not likely to influence ovarian reserve in euthyroid women whose TSH levels fall within the normal range although elevated TSH levels may be involved in the decline of serum AMH levels. [ABSTRACT FROM AUTHOR]

Purpose To visualize and analyze follicle development in ovarian tissue culture using physiological concentrations of follicle-stimulating hormone (FSH) and luteinizing hormone (LH) in order to establish an ovarian tissue culture system that enables efficient in vitro growth of follicles. Methods Ovarian tissues from 4-week-old female ICR mice were sliced and cultured. Images of ovarian tissues in culture were obtained at 24-h or 30-min intervals by using a microscope. The area of each follicle observed in the ovarian tissue slices was tracked and analyzed in association with oocyte maturation. Results We were able to track the development of each follicle using this culture system. Follicle growth was associated with oocyte maturation. Meiotically matured oocytes (MII) were obtained from 33% of al l follicles investigated. Approximately, a quarter of follicles (24%) did not grow and resulted in atresia. Conclusion Follicle dynamics were successfully visualized and analyzed in murine ovarian tissue culture. We were able to obtain mature oocytes from the fully grown follicles in vitro. This culture system would be helpful for efficient in vitro culturing of ovarian tissues. [ABSTRACT FROM AUTHOR]

Background: Serum anti-Müllerian hormone (AMH) concentration has been used to assess ovarian reserve in patients with endometriosis, especially when endometrioma surgery is involved. Previously, we reported that decreased serum AMH levels after cystectomy for endometriomas can recover to preoperative levels in some cases. In this present study, we assessed the sequential changes in serum AMH levels before and after cystectomy in terms of the state of the mesosalpinx prior to surgery. Methods: The retrospective cohort study recruited 53 patients from a series of prospective studies conducted from 2009 to 2015. All patients underwent laparoscopic cystectomy for endometriomas. If either mesosalpinx was involved in the endometrioma or adnexal adhesion before cystectomy, the case was defined as 'involved mesosalpinx' (n = 14). If both mesosalpinx remained anatomically correct, the case was classified as 'intact mesosalpinx' (n = 39). Blood samples were obtained from the patients 2 weeks before surgery, and at 1 month and 1 year after surgery to assess serum AMH levels. Results: The serum AMH levels (the involved group vs. the intact group) were 1.92 vs. 0.98 (P = 0.552) preoperatively, 0.59 vs. 1.99 (P = 0.049) at 1 month postoperatively, and 0.48 vs. 2.37 ng/mL (P = 0.007) at 1 year postoperatively. The involved mesosalpinx group showed a further decrease in serum AMH levels at 1 year postoperatively, while serum AMH levels in the intact mesosalpinx group tended to recover. Conclusion: These results suggest that pre-existing mesosalpinx disturbance, in combination with adhesiolysis, may be involved in the medium- and long-term decrease in ovarian reserve after endometrioma surgery. A disturbance in ovarian blood supply via the mesosalpinx may underlie this. [ABSTRACT FROM AUTHOR]

Kisspeptin, a hypothalamic neuropeptide, is expressed in the arcuate nucleus (ARC) that is considered as the center of the gonadotropin-releasing hormone (GnRH)-pulse generator. We hypothesized that kisspeptin expressed in the ARC is implicated in the disturbance of the hypothalamus−pituitary−ovary axis observed in polycystic ovary syndrome (PCOS). To test this hypothesis, we evaluated the hormonal profiles, luteinizing hormone (LH) pulse, and ARC kisspeptin immunoreactivity in a PCOS rat model using the anti-progestin RU486. We found an alteration of the LH pulse, including a trend towards an increased mean LH concentration and area under the curve, and a significant upregulation of the mean LH pulse amplitude. Additionally, a higher number of kisspeptin-positive cells was observed in the ARC of RU486-treated rats than in the ARC of intact rats. These results suggest the possible involvement of hypothalamic kisspeptin in the hypothalamus−pituitary−ovary axis and therefore, in PCOS pathophysiology. [ABSTRACT FROM AUTHOR]

A reduced response to progesterone in the eutopic endometrium with endometriosis and in endometriotic tissues is considered to be the underlying factor for endometriosis. CD10 is known to be expressed by endometrial and endometriotic stromal cells and may be induced by progestins, although the function of CD10 is not fully revealed in endometrial or endometriotic tissues. In the current study, the expression of CD10 was significantly increased by treatment of the cells with progesterone, 17β-estradiol, and dibutyryl cyclic adenosine monophosphate (cAMP) in the endometrial stromal cells. On the other hand, the expression of CD10 following treatment with progesterone, 17β-estradiol, and dibutyryl cAMP was not significantly increased in endometriotic stromal cells. The adhesion assay for endometrial and endometriotic stromal cells to hyaluronan using 5- or 6-(N-succinimidyloxycarbonyl)-fluorescein 3', 6'-diacetate–labeled cells demonstrated that the CD44-dependent adhesion of stromal cells was inhibited by CD10. As far as the induction of CD10 is concerned, the effect of progesterone was different between endometrial stromal cells and endometriotic stromal cells. CD10 might be involved in the development of endometriosis due to its influence on CD44-dependent cell adhesion. [ABSTRACT FROM PUBLISHER]

With proper and careful selection of patients, fertility-preserving surgery may be feasible in patients with ovarian malignancies. However, the loss of follicles by oophorectomy and chemotherapy results in decreased ovarian reserve, which consecutively affects reproductive capacity. We evaluated postoperative levels of serum anti-Müllerian hormone (AMH) in women with ovarian malignancies to assess the impact of the fertility-preserving surgery with or without the administration of chemotherapy on ovarian reserve. In 13 patients who underwent the fertility-preserving surgery with ( n = 9) or without ( n = 4) the administration of chemotherapy, serum AMH levels were measured and compared with serum AMH levels measured in patients undergone cystectomy for benign ovarian tumors as a control. We found that the mean AMH level in the treatment group measured 0.9 ng/mL, which was significantly lower than that measured in the control group (4.70 ± 3.77 ng/mL). The possibility of decreased ovarian reserve occurring in patients with ovarian malignancies following treatment with fertility-preserving surgery with or without the administration of chemotherapy should be considered for fertility planning. [ABSTRACT FROM AUTHOR]

Purpose: Inflammatory mediators, including chemokines, may play crucial roles in the development of endometriosis. Therefore, we investigated the expression and localization of CXCL16 and its receptor, CXCR6, in ovarian endometriotic tissues. We also examined whether CXCL16 induces IL-8 production in endometriotic stromal cells. Methods: We performed immunohistochemical and Western blotting analyses of in vivo and in vitro samples. IL-8 production was assayed using an ELISA. Results: Both CXCL16 and CXCR6 were expressed by endometriotic epithelial cells and stromal cells, but not normal ovarian stroma. A Western blotting analysis using primary cultured endometriotic stromal cells showed a constant expression of CXCL16 and CXCR6 in the proliferative phase, secretory phase and during gonadotropin-releasing hormone agonist therapy. CXCL16 induced IL-8 production in several endometriotic stromal cells in vitro. Conclusions: CXCL16 and CXCR6 might be involved in the pathophysiology of endometriosis through regulation of the inflammatory response. [ABSTRACT FROM AUTHOR]

Background: Given the seriousness of chemotherapy-induced ovarian injury in female cancer patients, the preservation of fertility, including through the use of cryopreservation technology and pharmaceuticals, requires investigation. Previous studies have shown that damage to the ovaries is related to oxidative stress caused by anticancer drugs. Therefore, superoxide dismutase (SOD) may represent a key factor in the pharmacological protection of the ovaries. The aim of our study was to identify the effects of mangafodipir, a manganese chelate and SOD-mimetic, on suppression of apoptosis in granulosa cells and primordial follicle activation induced by anticancer drugs. Methods: Cell viability assays using methyltrichlorosilane solutions and immunoblotting for cleaved caspase-3 were performed in in vitro experiments with the simultaneous addition of mangafodipir to human non-luteinized granulosa cell line (HGrC) cultures treated with hydrogen peroxide (H2O2), cisplatin, or paclitaxel. Count and morphological analyses of follicles at each developing stage in the ovaries and immunohistochemistry for cleaved caspase-3, Ki67 and 4-hydroxynonenal, a marker for oxidative stress, were also performed using mangafodipir-injected 6-week-old female ICR mice treated with cisplatin or paclitaxel. Further, mangafodipir was injected into 6-week-old female BALB/c mice inoculated with ES-2 to analyze whether mangafodipir inhibits the anti-tumor effects of cisplatin or paclitaxel treatment. Results: Mangafodipir attenuated apoptosis induced by H2O2 and anticancer drugs in vitro. Mangafodipir also decreased the expression of 4-hydroxynonenal and reduced cisplatin- and paclitaxel-induced apoptosis in granulosa cells in vivo. In addition, mangafodipir inhibited the loss of primordial follicles. Tumor xenograft studies in mice showed that mangafodipir did not affect anticancer drug antitumor effects. Conclusions: Oxidative stress might be one of the mechanisms of cisplatin- and paclitaxel-induced the loss of primordial follicles. Mangafodipir can reduce cisplatin- and paclitaxel-induced apoptosis in granulosa cells and primordial follicle activation partially via its SOD activity. At the same time, mangafodipir might have other potential mechanisms to inhibit the activation of primordial follicles. Further, mangafodipir attenuated the ovarian damage caused by cisplatin and paclitaxel without affecting their antitumor activities. Mangafodipir, therefore, though its efficacy might be limited, may be a new option for the preservation of fertility during anticancer treatment. [ABSTRACT FROM AUTHOR]

Objective: To investigate the protective effect of sphingosine-1-phosphate (S1P) against H2O2-induced apoptosis in human granulosa cell cultures with freshly harvested granulosa cells. Design: Experimental study. Setting: Academic medical center for reproductive medicine. Patient(s): Cultures of primary granulosa cells isolated from women undergoing in vitro fertilization (IVF). Intervention(s): None. Main Outcome Measure(s): Cell apoptosis and Western blot analysis of signaling pathway proteins. Result(s): We found that S1P (1 and 10 mM) statistically significantly decreased granulosa cell apoptosis after H2O2 treatment. The decreased cell apoptosis induced by S1P was abolished after treatment with VPC23019, an inhibitor of S1P1 and S1P3 receptors, W146, an inhibitor of S1P1 receptors, and CAY10444, an inhibitor of S1P3 receptors. A Western blot analysis revealed that the level of phospho-Akt increased and peaked at 10 minutes after 10 mM S1P exposure. Conclusion(s): Treatment with S1P can inhibit the apoptosis of granulosa cells in response to oxidative stress induced by H2O2. The protective effect of S1P is mediated by activating the PI3K/Akt pathway, and the antiapoptotic effect of S1P is mainly mediated through the S1P1 and S1P3 receptor. [ABSTRACT FROM AUTHOR]